Liver X Receptor (LXR) activation negatively regulates visfatin expression in macrophages

Energy Technology Data Exchange (ETDEWEB)

Mayi, Therese Hervee; Rigamonti, Elena [Univ Lille Nord de France, F-59000 Lille (France); INSERM UR1011, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France); Pattou, Francois [Univ Lille Nord de France, F-59000 Lille (France); Department of Endocrine Surgery, University Hospital, Lille (France); U859 Biotherapies for Diabetes, INSERM, Lille (France); Staels, Bart, E-mail: bart.staels@pasteur-lille.fr [Univ Lille Nord de France, F-59000 Lille (France); INSERM UR1011, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France); Chinetti-Gbaguidi, Giulia [Univ Lille Nord de France, F-59000 Lille (France); INSERM UR1011, F-59000 Lille (France); UDSL, F-59000 Lille (France); Institut Pasteur de Lille, F-59019 Lille (France)

2011-01-07

Research highlights: {yields} Synthetic LXR ligands decreased visfatin expression in human macrophages. {yields} LXR activation leads to a modest and transient decrease of NAD{sup +} concentration. {yields} LXR activation decreased PPAR{gamma}-induced visfatin in human macrophages. -- Abstract: Adipose tissue macrophages (ATM) are the major source of visfatin, a visceral fat adipokine upregulated during obesity. Also known to play a role in B cell differentiation (pre-B cell colony-enhancing factor (PBEF)) and NAD biosynthesis (nicotinamide phosphoribosyl transferase (NAMPT)), visfatin has been suggested to play a role in inflammation. Liver X Receptor (LXR) and Peroxisome Proliferator-Activated Receptor (PPAR){gamma} are nuclear receptors expressed in macrophages controlling the inflammatory response. Recently, we reported visfatin as a PPAR{gamma} target gene in human macrophages. In this study, we examined whether LXR regulates macrophage visfatin expression. Synthetic LXR ligands decreased visfatin gene expression in a LXR-dependent manner in human and murine macrophages. The decrease of visfatin mRNA was paralleled by a decrease of protein secretion. Consequently, a modest and transient decrease of NAD{sup +} concentration was observed. Interestingly, LXR activation decreased the PPAR{gamma}-induced visfatin gene and protein secretion in human macrophages. Our results identify visfatin as a gene oppositely regulated by the LXR and PPAR{gamma} pathways in human macrophages.

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

Science.gov (United States)

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.

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

Directory of Open Access Journals (Sweden)

Emily M Eshleman

2017-05-01

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

PPARγ regulates the expression of cholesterol metabolism genes in alveolar macrophages

International Nuclear Information System (INIS)

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

2010-01-01

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

Epigenetic Regulation of Monocyte and Macrophage Function

NARCIS (Netherlands)

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

2016-01-01

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

TLR Stimulation Dynamically Regulates Heme and Iron Export Gene Expression in Macrophages

Directory of Open Access Journals (Sweden)

Mary Philip

2016-01-01

Full Text Available Pathogenic bacteria have evolved multiple mechanisms to capture iron or iron-containing heme from host tissues or blood. In response, organisms have developed defense mechanisms to keep iron from pathogens. Very little of the body’s iron store is available as free heme; rather nearly all body iron is complexed with heme or other proteins. The feline leukemia virus, subgroup C (FeLV-C receptor, FLVCR, exports heme from cells. It was unknown whether FLVCR regulates heme-iron availability after infection, but given that other heme regulatory proteins are upregulated in macrophages in response to bacterial infection, we hypothesized that macrophages dynamically regulate FLVCR. We stimulated murine primary macrophages or macrophage cell lines with LPS and found that Flvcr is rapidly downregulated in a TLR4/MD2-dependent manner; TLR1/2 and TLR3 stimulation also decreased Flvcr expression. We identified several candidate TLR-activated transcription factors that can bind to the Flvcr promoter. Macrophages must balance the need to sequester iron from systemic circulating or intracellular pathogens with the macrophage requirement for heme and iron to produce reactive oxygen species. Our findings underscore the complexity of this regulation and point to a new role for FLVCR and heme export in macrophages responses to infection and inflammation.

Mitogen-activated protein kinase phosphatase 1 (MKP-1) in macrophage biology and cardiovascular disease. A redox-regulated master controller of monocyte function and macrophage phenotype.

Science.gov (United States)

Kim, Hong Seok; Asmis, Reto

2017-08-01

MAPK pathways play a critical role in the activation of monocytes and macrophages by pathogens, signaling molecules and environmental cues and in the regulation of macrophage function and plasticity. MAPK phosphatase 1 (MKP-1) has emerged as the main counter-regulator of MAPK signaling in monocytes and macrophages. Loss of MKP-1 in monocytes and macrophages in response to metabolic stress leads to dysregulation of monocyte adhesion and migration, and gives rise to dysfunctional, proatherogenic monocyte-derived macrophages. Here we review the properties of this redox-regulated dual-specificity MAPK phosphatase and the role of MKP-1 in monocyte and macrophage biology and cardiovascular diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

DMPD: The atrial natriuretic peptide regulates the production of inflammatorymediators in macrophages. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 11890659 The atrial natriuretic peptide regulates the production of inflammatorymed...tml) (.csml) Show The atrial natriuretic peptide regulates the production of inflammatorymediators in macrop...hages. PubmedID 11890659 Title The atrial natriuretic peptide regulates the produ

Pro-Resolving Mediators in Regulating and Conferring Macrophage Function

Directory of Open Access Journals (Sweden)

Jesmond Dalli

2017-11-01

Full Text Available Macrophages are central in coordinating the host response to both sterile and infective insults. Clearance of apoptotic cells and cellular debris is a key biological action preformed by macrophages that paves the way to the resolution of local inflammation, repair and regeneration of damaged tissues, and re-establishment of function. The essential fatty acid-derived autacoids termed specialized pro-resolving mediators (SPM play central roles in promoting these processes. In the present article, we will review the role of microvesicles in controlling macrophage efferocytosis and SPM production. We will also discuss the role of both apoptotic cells and microvesicles in providing substrate for transcellular biosynthesis of several SPM families during efferocyotsis. In addition, this article will discuss the biological actions of the recently uncovered macrophage-derived SPM termed maresins. These mediators are produced via 14-lipoxygenation of docosahexaenoic acid that is either enzymatically converted to mediators carrying two hydroxyl groups or to autacoids that are peptide-lipid conjugates, coined maresin conjugates in tissue regeneration. The formation of these mediators is temporally regulated during acute self-limited infectious-inflammation where they promote the uptake and clearance of apoptotic cells, regulate several aspects of the tissue repair and regeneration, and display potent anti-nociceptive actions.

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

Energy Technology Data Exchange (ETDEWEB)

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

2010-03-19

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

Intratumoral macrophages contribute to epithelial-mesenchymal transition in solid tumors

International Nuclear Information System (INIS)

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

2012-01-01

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

Promotion of Tumor Invasion by Cooperation of Granulocytes and Macrophages Activated by Anti-tumor Antibodies

Directory of Open Access Journals (Sweden)

Emilio Barbera-Guillem

1999-11-01

Full Text Available We investigated the potential role of anti-tumor antibodies and tumor antigens in the formation of immune complexes which promote matrix degradation and angiogenesis. B-cell deficient or B-cell depleted mice showed a reduction in tumor invasion and metastasis. In vitro invasion assays and in vivo models of metastasis showed that anti-sTn antibodies and sTn tumor antigens form complexes which induce granulocytes and macrophages together to mediate tumor invasion and metastasis by processes including extracellular matrix degradation and angiogenesis. These results suggest the existence of a tumor promoting role of a B-cell immune response induced by shed tumor associated antigens of solid, nonlymphoid tumors.

Hacking macrophage-associated immunosuppression for regulating glioblastoma angiogenesis.

Science.gov (United States)

Cui, Xin; Morales, Renee-Tyler Tan; Qian, Weiyi; Wang, Haoyu; Gagner, Jean-Pierre; Dolgalev, Igor; Placantonakis, Dimitris; Zagzag, David; Cimmino, Luisa; Snuderl, Matija; Lam, Raymond H W; Chen, Weiqiang

2018-04-01

Glioblastoma (GBM) is the most lethal primary adult brain tumor and its pathology is hallmarked by distorted neovascularization, diffuse tumor-associated macrophage infiltration, and potent immunosuppression. Reconstituting organotypic tumor angiogenesis models with biomimetic cell heterogeneity and interactions, pro-/anti-inflammatory milieu and extracellular matrix (ECM) mechanics is critical for preclinical anti-angiogenic therapeutic screening. However, current in vitro systems do not accurately mirror in vivo human brain tumor microenvironment. Here, we engineered a three-dimensional (3D), microfluidic angiogenesis model with controllable and biomimetic immunosuppressive conditions, immune-vascular and cell-matrix interactions. We demonstrate in vitro, GL261 and CT-2A GBM-like tumors steer macrophage polarization towards a M2-like phenotype for fostering an immunosuppressive and proangiogenic niche, which is consistent with human brain tumors. We distinguished that GBM and M2-like immunosuppressive macrophages promote angiogenesis, while M1-like pro-inflammatory macrophages suppress angiogenesis, which we coin "inflammation-driven angiogenesis." We observed soluble immunosuppressive cytokines, predominantly TGF-β1, and surface integrin (α v β 3 ) endothelial-macrophage interactions are required in inflammation-driven angiogenesis. We demonstrated tuning cell-adhesion receptors using an integrin (α v β 3 )-specific collagen hydrogel regulated inflammation-driven angiogenesis through Src-PI3K-YAP signaling, highlighting the importance of altered cell-ECM interactions in inflammation. To validate the preclinical applications of our 3D organoid model and mechanistic findings of inflammation-driven angiogenesis, we screened a novel dual integrin (α v β 3 ) and cytokine receptor (TGFβ-R1) blockade that suppresses GBM tumor neovascularization by simultaneously targeting macrophage-associated immunosuppression, endothelial-macrophage interactions, and

Niacin and its metabolites as master regulators of macrophage activation.

Science.gov (United States)

Montserrat-de la Paz, Sergio; Naranjo, M Carmen; Lopez, Sergio; Abia, Rocio; Muriana, Francisco J Garcia; Bermudez, Beatriz

2017-01-01

Niacin is a broad-spectrum lipid-regulating drug used for clinical therapy of chronic high-grade inflammatory diseases. However, the mechanisms by which either niacin or the byproducts of its catabolism ameliorate these inflammatory diseases are not clear yet. Human circulating monocytes and mature macrophages were used to analyze the effects of niacin and its metabolites (NAM, NUA and 2-Pyr) on oxidative stress, plasticity and inflammatory response by using biochemical, flow cytometry, quantitative real-time PCR and Western blot technologies. Niacin, NAM and 2-Pyr significantly decreased ROS, NO and NOS2 expression in LPS-treated human mature macrophages. Niacin and NAM skewed macrophage polarization toward antiinflammatory M2 macrophage whereas a trend toward proinflammatory M1 macrophage was noted following treatment with NUA. Niacin and NAM also reduced the inflammatory competence of LPS-treated human mature macrophages and promoted bias toward antiinflammatory CD14 + CD16 ++ nonclassical human primary monocytes. This study reveals for the first time that niacin and its metabolites possess antioxidant, reprogramming and antiinflammatory properties on human primary monocytes and monocyte-derived macrophages. Our findings imply a new understanding of the mechanisms by which niacin and its metabolites favor a continuous and gradual plasticity process in the human monocyte/macrophage system. Copyright © 2016 Elsevier Inc. All rights reserved.

Mechanisms of Hypoxic Up-Regulation of Versican Gene Expression in Macrophages.

Directory of Open Access Journals (Sweden)

Fattah Sotoodehnejadnematalahi

Full Text Available Hypoxia is a hallmark of many pathological tissues. Macrophages accumulate in hypoxic sites and up-regulate a range of hypoxia-inducible genes. The matrix proteoglycan versican has been identified as one such gene, but the mechanisms responsible for hypoxic induction are not fully characterised. Here we investigate the up-regulation of versican by hypoxia in primary human monocyte-derived macrophages (HMDM, and, intriguingly, show that versican mRNA is up-regulated much more highly (>600 fold by long term hypoxia (5 days than by 1 day of hypoxia (48 fold. We report that versican mRNA decay rates are not affected by hypoxia, demonstrating that hypoxic induction of versican mRNA is mediated by increased transcription. Deletion analysis of the promoter identified two regions required for high level promoter activity of luciferase reporter constructs in human macrophages. The hypoxia-inducible transcription factor HIF-1 has previously been implicated as a key potential regulator of versican expression in hypoxia, however our data suggest that HIF-1 up-regulation is unlikely to be principally responsible for the high levels of induction observed in HMDM. Treatment of HMDM with two distinct specific inhibitors of Phosphoinositide 3-kinase (PI3K, LY290042 and wortmannin, significantly reduced induction of versican mRNA by hypoxia and provides evidence of a role for PI3K in hypoxic up-regulation of versican expression.

Pharmacological Regulation of Neuropathic Pain Driven by Inflammatory Macrophages

Directory of Open Access Journals (Sweden)

Norikazu Kiguchi

2017-11-01

Full Text Available Neuropathic pain can have a major effect on quality of life but current therapies are often inadequate. Growing evidence suggests that neuropathic pain induced by nerve damage is caused by chronic inflammation. Upon nerve injury, damaged cells secrete pro-inflammatory molecules that activate cells in the surrounding tissue and recruit circulating leukocytes to the site of injury. Among these, the most abundant cell type is macrophages, which produce several key molecules involved in pain enhancement, including cytokines and chemokines. Given their central role in the regulation of peripheral sensitization, macrophage-derived cytokines and chemokines could be useful targets for the development of novel therapeutics. Inhibition of key pro-inflammatory cytokines and chemokines prevents neuroinflammation and neuropathic pain; moreover, recent studies have demonstrated the effectiveness of pharmacological inhibition of inflammatory (M1 macrophages. Nicotinic acetylcholine receptor ligands and T helper type 2 cytokines that reduce M1 macrophages are able to relieve neuropathic pain. Future translational studies in non-human primates will be crucial for determining the regulatory mechanisms underlying neuroinflammation-associated neuropathic pain. In turn, this knowledge will assist in the development of novel pharmacotherapies targeting macrophage-driven neuroinflammation for the treatment of intractable neuropathic pain.

Macrophages and fibroblasts : key regulators in wound healing, fibrosis and the foreign body reaction

NARCIS (Netherlands)

Ploeger, Diana

2017-01-01

Macrophages and fibroblasts are key regulators in wound healing, fibrosis and foreign body reaction (FBR). After injury macrophages migrate through the extracellular matrix (ECM) towards the wounded area, and adopt a M1 or M2 phenotype. M1 macrophages are associated with tissue injury and

Epigenetic suppression of neprilysin regulates breast cancer invasion.

Science.gov (United States)

Stephen, H M; Khoury, R J; Majmudar, P R; Blaylock, T; Hawkins, K; Salama, M S; Scott, M D; Cosminsky, B; Utreja, N K; Britt, J; Conway, R E

2016-03-07

In women, invasive breast cancer is the second most common cancer and the second cause of cancer-related death. Therefore, identifying novel regulators of breast cancer invasion could lead to additional biomarkers and therapeutic targets. Neprilysin, a cell-surface enzyme that cleaves and inactivates a number of substrates including endothelin-1 (ET1), has been implicated in breast cancer, but whether neprilysin promotes or inhibits breast cancer cell progression and metastasis is unclear. Here, we asked whether neprilysin expression predicts and functionally regulates breast cancer cell invasion. RT-PCR and flow cytometry analysis of MDA-MB-231 and MCF-7 breast cancer cell lines revealed decreased neprilysin expression compared with normal epithelial cells. Expression was also suppressed in invasive ductal carcinoma (IDC) compared with normal tissue. In addition, in vtro invasion assays demonstrated that neprilysin overexpression decreased breast cancer cell invasion, whereas neprilysin suppression augmented invasion. Furthermore, inhibiting neprilysin in MCF-7 breast cancer cells increased ET1 levels significantly, whereas overexpressing neprilysin decreased extracellular-signal related kinase (ERK) activation, indicating that neprilysin negatively regulates ET1-induced activation of mitogen-activated protein kinase (MAPK) signaling. To determine whether neprilysin was epigenetically suppressed in breast cancer, we performed bisulfite conversion analysis of breast cancer cells and clinical tumor samples. We found that the neprilysin promoter was hypermethylated in breast cancer; chemical reversal of methylation in MDA-MB-231 cells reactivated neprilysin expression and inhibited cancer cell invasion. Analysis of cancer databases revealed that neprilysin methylation significantly associates with survival in stage I IDC and estrogen receptor-negative breast cancer subtypes. These results demonstrate that neprilysin negatively regulates the ET axis in breast cancer

Macrophage pro-inflammatory response to Francisella novicida infection is regulated by SHIP.

Directory of Open Access Journals (Sweden)

Kishore V L Parsa

2006-07-01

Full Text Available Francisella tularensis, a Gram-negative facultative intracellular pathogen infecting principally macrophages and monocytes, is the etiological agent of tularemia. Macrophage responses to F. tularensis infection include the production of pro-inflammatory cytokines such as interleukin (IL-12, which is critical for immunity against infection. Molecular mechanisms regulating production of these inflammatory mediators are poorly understood. Herein we report that the SH2 domain-containing inositol phosphatase (SHIP is phosphorylated upon infection of primary murine macrophages with the genetically related F. novicida, and negatively regulates F. novicida-induced cytokine production. Analyses of the molecular details revealed that in addition to activating the MAP kinases, F. novicida infection also activated the phosphatidylinositol 3-kinase (PI3K/Akt pathway in these cells. Interestingly, SHIP-deficient macrophages displayed enhanced Akt activation upon F. novicida infection, suggesting elevated PI3K-dependent activation pathways in absence of SHIP. Inhibition of PI3K/Akt resulted in suppression of F. novicida-induced cytokine production through the inhibition of NFkappaB. Consistently, macrophages lacking SHIP displayed enhanced NFkappaB-driven gene transcription, whereas overexpression of SHIP led to decreased NFkappaB activation. Thus, we propose that SHIP negatively regulates F. novicida-induced inflammatory cytokine response by antagonizing the PI3K/Akt pathway and suppressing NFkappaB-mediated gene transcription. A detailed analysis of phosphoinositide signaling may provide valuable clues for better understanding the pathogenesis of tularemia.

Plasmid Negative Regulation of CPAF Expression Is Pgp4 Independent and Restricted to Invasive Chlamydia trachomatis Biovars

Directory of Open Access Journals (Sweden)

Michael John Patton

2018-01-01

Full Text Available Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes blinding trachoma and sexually transmitted disease. C. trachomatis isolates are classified into 2 biovars—lymphogranuloma venereum (LGV and trachoma—which are distinguished biologically by their natural host cell infection tropism. LGV biovars infect macrophages and are invasive, whereas trachoma biovars infect oculo-urogenital epithelial cells and are noninvasive. The C. trachomatis plasmid is an important virulence factor in the pathogenesis of these infections. Central to its pathogenic role is the transcriptional regulatory function of the plasmid protein Pgp4, which regulates the expression of plasmid and chromosomal virulence genes. As many gene regulatory functions are post-transcriptional, we employed a comparative proteomic study of cells infected with plasmid-cured C. trachomatis serovars A and D (trachoma biovar, a L2 serovar (LGV biovar, and the L2 serovar transformed with a plasmid containing a nonsense mutation in pgp4 to more completely elucidate the effects of the plasmid on chlamydial infection biology. Our results show that the Pgp4-dependent elevations in the levels of Pgp3 and a conserved core set of chromosomally encoded proteins are remarkably similar for serovars within both C. trachomatis biovars. Conversely, we found a plasmid-dependent, Pgp4-independent, negative regulation in the expression of the chlamydial protease-like activity factor (CPAF for the L2 serovar but not the A and D serovars. The molecular mechanism of plasmid-dependent negative regulation of CPAF expression in the LGV serovar is not understood but is likely important to understanding its macrophage infection tropism and invasive infection nature.

Plexin-B2 negatively regulates macrophage motility, Rac, and Cdc42 activation.

Directory of Open Access Journals (Sweden)

Kelly E Roney

Full Text Available Plexins are cell surface receptors widely studied in the nervous system, where they mediate migration and morphogenesis though the Rho family of small GTPases. More recently, plexins have been implicated in immune processes including cell-cell interaction, immune activation, migration, and cytokine production. Plexin-B2 facilitates ligand induced cell guidance and migration in the nervous system, and induces cytoskeletal changes in overexpression assays through RhoGTPase. The function of Plexin-B2 in the immune system is unknown. This report shows that Plexin-B2 is highly expressed on cells of the innate immune system in the mouse, including macrophages, conventional dendritic cells, and plasmacytoid dendritic cells. However, Plexin-B2 does not appear to regulate the production of proinflammatory cytokines, phagocytosis of a variety of targets, or directional migration towards chemoattractants or extracellular matrix in mouse macrophages. Instead, Plxnb2(-/- macrophages have greater cellular motility than wild type in the unstimulated state that is accompanied by more active, GTP-bound Rac and Cdc42. Additionally, Plxnb2(-/- macrophages demonstrate faster in vitro wound closure activity. Studies have shown that a closely related family member, Plexin-B1, binds to active Rac and sequesters it from downstream signaling. The interaction of Plexin-B2 with Rac has only been previously confirmed in yeast and bacterial overexpression assays. The data presented here show that Plexin-B2 functions in mouse macrophages as a negative regulator of the GTPases Rac and Cdc42 and as a negative regulator of basal cell motility and wound healing.

The macrophage A2B adenosine receptor regulates tissue insulin sensitivity.

Directory of Open Access Journals (Sweden)

Hillary Johnston-Cox

Full Text Available High fat diet (HFD-induced type 2 diabetes continues to be an epidemic with significant risk for various pathologies. Previously, we identified the A2b adenosine receptor (A2bAR, an established regulator of inflammation, as a regulator of HFD-induced insulin resistance. In particular, HFD was associated with vast upregulation of liver A2bAR in control mice, and while mice lacking this receptor showed augmented liver inflammation and tissue insulin resistance. As the A2bAR is expressed in different tissues, here, we provide the first lead to cellular mechanism by demonstrating that the receptor's influence on tissue insulin sensitivity is mediated via its expression in macrophages. This was shown using a newly generated transgenic mouse model expressing the A2bAR gene in the macrophage lineage on an otherwise A2bAR null background. Reinstatement of macrophage A2bAR expression in A2bAR null mice fed HFD restored insulin tolerance and tissue insulin signaling to the level of control mice. The molecular mechanism for this effect involves A2bAR-mediated changes in cyclic adenosine monophosphate in macrophages, reducing the expression and release of inflammatory cytokines, which downregulate insulin receptor-2. Thus, our results illustrate that macrophage A2bAR signaling is needed and sufficient for relaying the protective effect of the A2bAR against HFD-induced tissue inflammation and insulin resistance in mice.

Tumor-Associated Macrophages and Neutrophils in Tumor Microenvironment

Directory of Open Access Journals (Sweden)

Jaehong Kim

2016-01-01

Full Text Available Distinct tumor microenvironment forms in each progression step of cancer and has diverse capacities to induce both adverse and beneficial consequences for tumorigenesis. It is now known that immune cells can be activated to favor tumor growth and progression, most probably influenced by the tumor microenvironment. Tumor-associated macrophages and tumor-associated neutrophils can exert protumoral functions, enhancing tumor cell invasion and metastasis, angiogenesis, and extracellular matrix remodeling, while inhibiting the antitumoral immune surveillance. Considering that neutrophils in inflammatory environments recruit macrophages and that recruited macrophages affect neutrophil functions, there may be various degrees of interaction between tumor-associated macrophages and tumor-associated neutrophils. Platelets also play an important role in the recruitment and regulation of monocytic and granulocytic cells in the tumor tissues, suggesting that platelet function may be essential for generation of tumor-associated macrophages and tumor-associated neutrophils. In this review, we will explore the biology of tumor-associated macrophages and tumor-associated neutrophils and their possible interactions in the tumor microenvironment. Special attention will be given to the recruitment and activation of these tumor-associated cells and to the roles they play in maintenance of the tumor microenvironment and progression of tumors.

Intracellular glutathione status regulates mouse bone marrow monocyte-derived macrophage differentiation and phagocytic activity

International Nuclear Information System (INIS)

Kim, Jin-Man; Kim, Hyunsoo; Kwon, Soon Bok; Lee, Soo Young; Chung, Sung-Chang; Jeong, Dae-Won; Min, Byung-Moo

2004-01-01

Although a redox shift can regulate the development of cells, including proliferation, differentiation, and survival, the role of the glutathione (GSH) redox status in macrophage differentiation remains unclear. In order to elucidate the role of a redox shift, macrophage-like cells were differentiated from the bone marrow-derived monocytes that were treated with a macrophage colony stimulating factor (M-CSF or CSF-1) for 3 days. The macrophagic cells were characterized by a time-dependent increase in three major symptoms: the number of phagocytic cells, the number of adherent cells, and the mRNA expression of c-fms, a M-CSF receptor that is one of the macrophage-specific markers and mediates development signals. Upon M-CSF-driven macrophage differentiation, the GSH/GSSG ratio was significantly lower on day 1 than that observed on day 0 but was constant on days 1-3. To assess the effect of the GSH-depleted and -repleted status on the differentiation and phagocytosis of the macrophages, GSH depletion by BSO, a specific inhibitor of the de novo GSH synthesis, inhibited the formation of the adherent macrophagic cells by the down-regulation of c-fms, but did not affect the phagocytic activity of the macrophages. To the contrary, GSH repletion by the addition of NAC, which is a GSH precursor, or reduced GSH in media had no effect on macrophage differentiation, and led to a decrease in the phagocytic activity. Furthermore, we observed that there is checkpoint that is capable of releasing from the inhibition of the formation of the adherent macrophagic cells according to GSH depletion by BSO. Summarizing, these results indicate that the intracellular GSH status plays an important role in the differentiation and phagocytosis of macrophages

Regulation of antioxidant enzyme activities in male and female rat macrophages by sex steroids

Directory of Open Access Journals (Sweden)

Azevedo R.B.

2001-01-01

Full Text Available Human and animal immune functions present sex dimorphism that seems to be mainly regulated by sex hormones. In the present study, the activities of the antioxidant enzymes total superoxide dismutase (SOD, catalase (CAT, and glutathione peroxidase (GSH-Px were measured in intraperitoneal resident macrophages from adult male and female rats. In addition to comparing males and females, we also examined the regulation of these enzyme activities in macrophages by sex steroids. GSH-Px activity did not differ between male and female macrophages. However, both total SOD and CAT activities were markedly higher in females than in males (83 and 180%. Removal of the gonads in both males and females (comparison between castrated groups increased the difference in SOD activity from 83 to 138% and reduced the difference in CAT activity from 180 to 86%. Castration and testosterone administration did not significantly modify the activities of the antioxidant enzymes in male macrophages. Ovariectomy did not affect SOD or GSH-Px activity but markedly reduced (48% CAT activity. This latter change was fully reversed by estrogen administration, whereas progesterone had a smaller effect. These results led us to conclude that differences in the SOD and CAT activities may partially explain some of the differences in immune function reported for males and females. Also, estrogen is a potent regulator of CAT in macrophages and therefore this enzyme activity in macrophages may vary considerably during the menstrual cycle.

Tumor-Associated Macrophages as Major Players in the Tumor Microenvironment

International Nuclear Information System (INIS)

Chanmee, Theerawut; Ontong, Pawared; Konno, Kenjiro; Itano, Naoki

2014-01-01

During tumor progression, circulating monocytes and macrophages are actively recruited into tumors where they alter the tumor microenvironment to accelerate tumor progression. Macrophages shift their functional phenotypes in response to various microenvironmental signals generated from tumor and stromal cells. Based on their function, macrophages are divided broadly into two categories: classical M1 and alternative M2 macrophages. The M1 macrophage is involved in the inflammatory response, pathogen clearance, and antitumor immunity. In contrast, the M2 macrophage influences an anti-inflammatory response, wound healing, and pro-tumorigenic properties. Tumor-associated macrophages (TAMs) closely resemble the M2-polarized macrophages and are critical modulators of the tumor microenvironment. Clinicopathological studies have suggested that TAM accumulation in tumors correlates with a poor clinical outcome. Consistent with that evidence, experimental and animal studies have supported the notion that TAMs can provide a favorable microenvironment to promote tumor development and progression. In this review article, we present an overview of mechanisms responsible for TAM recruitment and highlight the roles of TAMs in the regulation of tumor angiogenesis, invasion, metastasis, immunosuppression, and chemotherapeutic resistance. Finally, we discuss TAM-targeting therapy as a promising novel strategy for an indirect cancer therapy

Tumor-Associated Macrophages as Major Players in the Tumor Microenvironment

Energy Technology Data Exchange (ETDEWEB)

Chanmee, Theerawut [Institute of Advanced Technology, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan); Ontong, Pawared [Division of Engineering (Biotechnology), Graduate School of Engineering, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan); Konno, Kenjiro [Department of Animal Medical Sciences, Faculty of Life Sciences, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan); Itano, Naoki, E-mail: itanon@cc.kyoto-su.ac.jp [Institute of Advanced Technology, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan); Division of Engineering (Biotechnology), Graduate School of Engineering, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan); Department of Molecular Biosciences, Faculty of Life Sciences, Kyoto Sangyo University, Kita-ku, Kyoto 603-8555 (Japan)

2014-08-13

During tumor progression, circulating monocytes and macrophages are actively recruited into tumors where they alter the tumor microenvironment to accelerate tumor progression. Macrophages shift their functional phenotypes in response to various microenvironmental signals generated from tumor and stromal cells. Based on their function, macrophages are divided broadly into two categories: classical M1 and alternative M2 macrophages. The M1 macrophage is involved in the inflammatory response, pathogen clearance, and antitumor immunity. In contrast, the M2 macrophage influences an anti-inflammatory response, wound healing, and pro-tumorigenic properties. Tumor-associated macrophages (TAMs) closely resemble the M2-polarized macrophages and are critical modulators of the tumor microenvironment. Clinicopathological studies have suggested that TAM accumulation in tumors correlates with a poor clinical outcome. Consistent with that evidence, experimental and animal studies have supported the notion that TAMs can provide a favorable microenvironment to promote tumor development and progression. In this review article, we present an overview of mechanisms responsible for TAM recruitment and highlight the roles of TAMs in the regulation of tumor angiogenesis, invasion, metastasis, immunosuppression, and chemotherapeutic resistance. Finally, we discuss TAM-targeting therapy as a promising novel strategy for an indirect cancer therapy.

A TNF-Regulated Recombinatorial Macrophage Immune Receptor Implicated in Granuloma Formation in Tuberculosis

Science.gov (United States)

Streich, Roswita; Breysach, Caroline; Raddatz, Dirk; Oniga, Septimia; Peccerella, Teresa; Findeisen, Peter; Kzhyshkowska, Julia; Gratchev, Alexei; Schweyer, Stefan; Saunders, Bernadette; Wessels, Johannes T.; Möbius, Wiebke; Keane, Joseph; Becker, Heinz; Ganser, Arnold; Neumaier, Michael; Kaminski, Wolfgang E.

2011-01-01

Macrophages play a central role in host defense against mycobacterial infection and anti- TNF therapy is associated with granuloma disorganization and reactivation of tuberculosis in humans. Here, we provide evidence for the presence of a T cell receptor (TCR) αβ based recombinatorial immune receptor in subpopulations of human and mouse monocytes and macrophages. In vitro, we find that the macrophage-TCRαβ induces the release of CCL2 and modulates phagocytosis. TNF blockade suppresses macrophage-TCRαβ expression. Infection of macrophages from healthy individuals with mycobacteria triggers formation of clusters that express restricted TCR Vβ repertoires. In vivo, TCRαβ bearing macrophages abundantly accumulate at the inner host-pathogen contact zone of caseous granulomas from patients with lung tuberculosis. In chimeric mouse models, deletion of the variable macrophage-TCRαβ or TNF is associated with structurally compromised granulomas of pulmonary tuberculosis even in the presence of intact T cells. These results uncover a TNF-regulated recombinatorial immune receptor in monocytes/macrophages and demonstrate its implication in granuloma formation in tuberculosis. PMID:22114556

A TNF-regulated recombinatorial macrophage immune receptor implicated in granuloma formation in tuberculosis.

Directory of Open Access Journals (Sweden)

Alexander W Beham

2011-11-01

Full Text Available Macrophages play a central role in host defense against mycobacterial infection and anti- TNF therapy is associated with granuloma disorganization and reactivation of tuberculosis in humans. Here, we provide evidence for the presence of a T cell receptor (TCR αβ based recombinatorial immune receptor in subpopulations of human and mouse monocytes and macrophages. In vitro, we find that the macrophage-TCRαβ induces the release of CCL2 and modulates phagocytosis. TNF blockade suppresses macrophage-TCRαβ expression. Infection of macrophages from healthy individuals with mycobacteria triggers formation of clusters that express restricted TCR Vβ repertoires. In vivo, TCRαβ bearing macrophages abundantly accumulate at the inner host-pathogen contact zone of caseous granulomas from patients with lung tuberculosis. In chimeric mouse models, deletion of the variable macrophage-TCRαβ or TNF is associated with structurally compromised granulomas of pulmonary tuberculosis even in the presence of intact T cells. These results uncover a TNF-regulated recombinatorial immune receptor in monocytes/macrophages and demonstrate its implication in granuloma formation in tuberculosis.

Postprandial triglyceride-rich lipoproteins regulate perilipin-2 and perilipin-3 lipid-droplet-associated proteins in macrophages.

Science.gov (United States)

Varela, Lourdes M; López, Sergio; Ortega-Gómez, Almudena; Bermúdez, Beatriz; Buers, Insa; Robenek, Horst; Muriana, Francisco J G; Abia, Rocío

2015-04-01

Lipid accumulation in macrophages contributes to atherosclerosis. Within macrophages, lipids are stored in lipid droplets (LDs); perilipin-2 and perilipin-3 are the main LD-associated proteins. Postprandial triglyceride (TG)-rich lipoproteins induce LD accumulation in macrophages. The role of postprandial lipoproteins in perilipin-2 and perilipin-3 regulation was studied. TG-rich lipoproteins (TRLs) induced the levels of intracellular TGs, LDs and perilipin-2 protein expression in THP-1 macrophages and in Apoe(-/-) mice bone-marrow-derived macrophages with low and high basal levels of TGs. Perilipin-3 was only synthesized in mice macrophages with low basal levels of TGs. The regulation was dependent on the fatty acid composition of the lipoproteins; monounsaturated and polyunsaturated fatty acids (PUFAs) more strongly attenuated these effects compared with saturated fatty acids. In THP-1 macrophages, immunofluorescence microscopy and freeze-fracture immunogold labeling indicated that the lipoproteins translocated perilipin-3 from the cytoplasm to the LD surface; only the lipoproteins that were rich in PUFAs suppressed this effect. Chemical inhibition showed that lipoproteins induced perilipin-2 protein expression through the peroxisome proliferator-activated nuclear receptor (PPAR) PPARα and PPARγ pathways. Overall, our data indicate that postprandial TRLs may be involved in atherosclerotic plaque formation through the regulation of perilipin-2 and perilipin-3 proteins in macrophages. Because the fatty acid composition of the lipoproteins is dependent on the type of fat consumed, the ingestion of olive oil, which is rich in monounsaturated fatty acids, and fish oil, which is rich in omega-3 fatty acids, can be considered a good nutritional strategy to reduce the risk of atherosclerosis by LD-associated proteins decrease. Copyright © 2015 Elsevier Inc. All rights reserved.

Monocytes/macrophages support mammary tumor invasivity by co-secreting lineage-specific EGFR ligands and a STAT3 activator

International Nuclear Information System (INIS)

Vlaicu, Philip; Mertins, Philipp; Mayr, Thomas; Widschwendter, Peter; Ataseven, Beyhan; Högel, Bernhard; Eiermann, Wolfgang; Knyazev, Pjotr; Ullrich, Axel

2013-01-01

Tumor-associated macrophages (TAM) promote malignant progression, yet the repertoire of oncogenic factors secreted by TAM has not been clearly defined. We sought to analyze which EGFR- and STAT3-activating factors are secreted by monocytes/macrophages exposed to tumor cell-secreted factors. Following exposure of primary human monocytes and macrophages to supernatants of a variety of tumor cell lines, we have analyzed transcript and secreted protein levels of EGFR family ligands and of STAT3 activators. To validate our findings, we have analyzed TAM infiltration levels, systemic and local protein levels as well as clinical data of primary breast cancer patients. Primary human monocytes and macrophages respond to tumor cell-derived factors by secreting EGFR- and STAT3-activating ligands, thus inducing two important oncogenic pathways in carcinoma cells. Tumor cell-secreted factors trigger two stereotype secretory profiles in peripheral blood monocytes and differentiated macrophages: monocytes secrete epiregulin (EREG) and oncostatin-M (OSM), while macrophages secrete heparin-binding EGF-like growth factor (HB-EGF) and OSM. HB-EGF and OSM cooperatively induce tumor cell chemotaxis. HB-EGF and OSM are co-expressed by TAM in breast carcinoma patients, and plasma levels of both ligands correlate strongly. Elevated HB-EGF levels accompany TAM infiltration, tumor growth and dissemination in patients with invasive disease. Our work identifies systemic markers for TAM involvement in cancer progression, with the potential to be developed into molecular targets in cancer therapy

CD36 Differently Regulates Macrophage Responses to Smooth and Rough Lipopolysaccharide.

Directory of Open Access Journals (Sweden)

Rafał Biedroń

Full Text Available Lipopolysaccharide (LPS is the major pathogen-associated molecular pattern of Gram-negative bacterial infections, and includes smooth (S-LPS and rough (R-LPS chemotypes. Upon activation by LPS through CD14, TLR4/MD-2 heterodimers sequentially induce two waves of intracellular signaling for macrophage activation: the MyD88-dependent pathway from the plasma membrane and, following internalization, the TRIF-dependent pathway from endosomes. We sought to better define the role of scavenger receptors CD36 and CD204/SR-A as accessory LPS receptors that can contribute to pro-inflammatory and microbicidal activation of macrophages. We have found that CD36 differently regulates activation of mouse macrophages by S-LPS versus R-LPS. The ability of CD36 to substitute for CD14 in loading R-LPS, but not S-LPS onto TLR4/MD-2 allows CD14-independent macrophage responses to R-LPS. Conversely, S-LPS, but not R-LPS effectively stimulates CD14 binding to CD36, which favors S-LPS transfer from CD14 onto TLR4/MD-2 under conditions of low CD14 occupancy with S-LPS in serum-free medium. In contrast, in the presence of serum, CD36 reduces S-LPS binding to TLR4/MD-2 and the subsequent MyD88-dependent signaling, by mediating internalization of S-LPS/CD14 complexes. Additionally, CD36 positively regulates activation of TRIF-dependent signaling by both S-LPS and R-LPS, by promoting TLR4/MD-2 endocytosis. In contrast, we have found that SR-A does not function as a S-LPS receptor. Thus, by co-operating with CD14 in both R- and S-LPS loading onto TLR4/MD-2, CD36 can enhance the sensitivity of tissue-resident macrophages in detecting infections by Gram-negative bacteria. However, in later phases, following influx of serum to the infection site, the CD36-mediated negative regulation of MyD88-dependent branch of S-LPS-induced TLR4 signaling might constitute a mechanism to prevent an excessive inflammatory response, while preserving the adjuvant effect of S-LPS for adaptive

Classification of M1/M2-polarized human macrophages by label-free hyperspectral reflectance confocal microscopy and multivariate analysis.

Science.gov (United States)

Bertani, Francesca R; Mozetic, Pamela; Fioramonti, Marco; Iuliani, Michele; Ribelli, Giulia; Pantano, Francesco; Santini, Daniele; Tonini, Giuseppe; Trombetta, Marcella; Businaro, Luca; Selci, Stefano; Rainer, Alberto

2017-08-21

The possibility of detecting and classifying living cells in a label-free and non-invasive manner holds significant theranostic potential. In this work, Hyperspectral Imaging (HSI) has been successfully applied to the analysis of macrophagic polarization, given its central role in several pathological settings, including the regulation of tumour microenvironment. Human monocyte derived macrophages have been investigated using hyperspectral reflectance confocal microscopy, and hyperspectral datasets have been analysed in terms of M1 vs. M2 polarization by Principal Components Analysis (PCA). Following PCA, Linear Discriminant Analysis has been implemented for semi-automatic classification of macrophagic polarization from HSI data. Our results confirm the possibility to perform single-cell-level in vitro classification of M1 vs. M2 macrophages in a non-invasive and label-free manner with a high accuracy (above 98% for cells deriving from the same donor), supporting the idea of applying the technique to the study of complex interacting cellular systems, such in the case of tumour-immunity in vitro models.

Metformin reduces the endotoxin-induced down-regulation of apolipoprotein E gene expression in macrophages

Energy Technology Data Exchange (ETDEWEB)

Stavri, Simona; Trusca, Violeta G.; Simionescu, Maya; Gafencu, Anca V., E-mail: anca.gafencu@icbp.ro

2015-05-29

The atheroprotective role of macrophage-derived apolipoprotein E (apoE) is well known. Our previous reports demonstrated that inflammatory stress down-regulates apoE expression in macrophages, aggravating atherogenesis. Metformin, extensively used as an anti-diabetic drug, has also anti-inflammatory properties, and thus confers vascular protection. In this study, we questioned whether metformin could have an effect on apoE expression in macrophages in normal conditions or under lipopolysaccharide (LPS)-induced stress. The results showed that metformin slightly increases the apoE expression only at high doses (5–10 mM). Low doses of metformin (1–3 mM) significantly reduce the LPS down-regulatory effect on apoE expression in macrophages. Our experiments demonstrated that LPS-induced NF-κB binds to the macrophage-specific distal regulatory element of apoE gene, namely to the multienhancer 2 (ME.2) and its 5′-deletion fragments. The NF-κB binding on ME.2 and apoE promoter has a down-regulatory effect. In addition, data revealed that metformin impairs NF-κB nuclear translocation, and thus, improves the apoE levels in macrophages under inflammatory stress. The positive effect of metformin in the inflammatory states, its clinical safety and low cost, make this drug a potential adjuvant in the therapeutic strategies for atherosclerosis. - Highlights: • High doses of metformin slightly increase apoE expression in macrophages. • Low doses of metformin up-regulate apoE gene in endotoxin-stressed macrophages. • Metformin reduces the negative effect of LPS on apoE expression by NF-κB inhibition.

Metformin reduces the endotoxin-induced down-regulation of apolipoprotein E gene expression in macrophages

International Nuclear Information System (INIS)

Stavri, Simona; Trusca, Violeta G.; Simionescu, Maya; Gafencu, Anca V.

2015-01-01

The atheroprotective role of macrophage-derived apolipoprotein E (apoE) is well known. Our previous reports demonstrated that inflammatory stress down-regulates apoE expression in macrophages, aggravating atherogenesis. Metformin, extensively used as an anti-diabetic drug, has also anti-inflammatory properties, and thus confers vascular protection. In this study, we questioned whether metformin could have an effect on apoE expression in macrophages in normal conditions or under lipopolysaccharide (LPS)-induced stress. The results showed that metformin slightly increases the apoE expression only at high doses (5–10 mM). Low doses of metformin (1–3 mM) significantly reduce the LPS down-regulatory effect on apoE expression in macrophages. Our experiments demonstrated that LPS-induced NF-κB binds to the macrophage-specific distal regulatory element of apoE gene, namely to the multienhancer 2 (ME.2) and its 5′-deletion fragments. The NF-κB binding on ME.2 and apoE promoter has a down-regulatory effect. In addition, data revealed that metformin impairs NF-κB nuclear translocation, and thus, improves the apoE levels in macrophages under inflammatory stress. The positive effect of metformin in the inflammatory states, its clinical safety and low cost, make this drug a potential adjuvant in the therapeutic strategies for atherosclerosis. - Highlights: • High doses of metformin slightly increase apoE expression in macrophages. • Low doses of metformin up-regulate apoE gene in endotoxin-stressed macrophages. • Metformin reduces the negative effect of LPS on apoE expression by NF-κB inhibition

Macrophage activation and differentiation signals regulate schlafen-4 gene expression: evidence for Schlafen-4 as a modulator of myelopoiesis.

Directory of Open Access Journals (Sweden)

Wendy J van Zuylen

Full Text Available BACKGROUND: The ten mouse and six human members of the Schlafen (Slfn gene family all contain an AAA domain. Little is known of their function, but previous studies suggest roles in immune cell development. In this report, we assessed Slfn regulation and function in macrophages, which are key cellular regulators of innate immunity. METHODOLOGY/PRINCIPAL FINDINGS: Multiple members of the Slfn family were up-regulated in mouse bone marrow-derived macrophages (BMM by the Toll-like Receptor (TLR4 agonist lipopolysaccharide (LPS, the TLR3 agonist Poly(I∶C, and in disease-affected joints in the collagen-induced model of rheumatoid arthritis. Of these, the most inducible was Slfn4. TLR agonists that signal exclusively through the MyD88 adaptor protein had more modest effects on Slfn4 mRNA levels, thus implicating MyD88-independent signalling and autocrine interferon (IFN-β in inducible expression. This was supported by the substantial reduction in basal and LPS-induced Slfn4 mRNA expression in IFNAR-1⁻/⁻ BMM. LPS causes growth arrest in macrophages, and other Slfn family genes have been implicated in growth control. Slfn4 mRNA levels were repressed during macrophage colony-stimulating factor (CSF-1-mediated differentiation of bone marrow progenitors into BMM. To determine the role of Slfn4 in vivo, we over-expressed the gene specifically in macrophages in mice using a csf1r promoter-driven binary expression system. Transgenic over-expression of Slfn4 in myeloid cells did not alter macrophage colony formation or proliferation in vitro. Monocyte numbers, as well as inflammatory macrophages recruited to the peritoneal cavity, were reduced in transgenic mice that specifically over-expressed Slfn4, while macrophage numbers and hematopoietic activity were increased in the livers and spleens. CONCLUSIONS: Slfn4 mRNA levels were up-regulated during macrophage activation but down-regulated during differentiation. Constitutive Slfn4 expression in the

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

International Nuclear Information System (INIS)

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

2016-01-01

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

Nogo-B Facilitates LPS-Mediated Immune Responses by Up-Regulation of TLR4-Signaling in Macrophage RAW264.7

Directory of Open Access Journals (Sweden)

Ying Zhu

2017-01-01

Full Text Available Background/Aims: Nogo-B, a member of the reticulon family of proteins, is mainly located in the endoplasmic reticulum (ER. Here, we investigate the function and mechanism of Nogo-B in the regulation of TLR4-associated immune responses in the macrophage cell line of RAW264.7. Methods: Nogo-B was up- and down-regulated through the use of appropriate adenoviral vectors or siRNA, and the effects of Nogo-B on macrophages under liposaccharide (LPS stimulation were evaluated via western blotting, immunofluorescence, enzyme-linked immunosorbent assay (ELISA, flow cytometric analysis, and transwell assay. Results: Our data indicates that the protein of Nogo-B was down-regulated in a time- and dose-dependent manner following LPS administration in the macrophage. Nogo-B overexpression increased the production of inflammatory cytokines (MCP-1, TNF-α, IL-1β, and TGF-β, enhanced macrophage migration activities, activated major histocompatibility complex II (MHC II, and elevated the expression of macrophage scavenger receptor 1(MSR1, all of which suggest that Nogo-B is necessary for immune responses and plays an important role in regulating macrophage recruitment. Mechanistically, Nogo-B may enhance TLR4 expression in macrophage surfaces, activate mitogen-activated protein kinase (MAPK pathways, and initiate inflammatory responses. Conclusion: These findings illustrate the key regulatory functions of Nogo-B in facilitating LPS-mediated immune responses through promoting the phosphorylation of MAP kinase.

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

Directory of Open Access Journals (Sweden)

Groot-Kormelink Paul J

2012-10-01

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

Targeting tumor-associated macrophages by anti-tumor Chinese materia medica.

Science.gov (United States)

Pu, Wei-Ling; Sun, Li-Kang; Gao, Xiu-Mei; Rüegg, Curzio; Cuendet, Muriel; Hottiger, Micheal O; Zhou, Kun; Miao, Lin; Zhang, Yun-Sha; Gebauer, Margaret

2017-10-01

Tumor-associated macrophages (TAMs) play a key role in all stages of tumorigenesis and tumor progression. TAMs secrete different kinds of cytokines, chemokines, and enzymes to affect the progression, metastasis, and resistance to therapy depending on their state of reprogramming. Therapeutic benefit in targeting TAMs suggests that macrophages are attractive targets for cancer treatment. Chinese materia medica (CMM) is an important approach for treating cancer in China and in the Asian region. According to the theory of Chinese medicine (CM) and its practice, some prescriptions of CM regulate the body's internal environment possibly including the remodeling the tumor microenvironment (TME). Here we briefly summarize the pivotal effects of TAMs in shaping the TME and promoting tumorigenesis, invasion, metastasis and immunosuppression. Furthermore, we illustrate the effects and mechanisms of CMM targeting TAMs in antitumor therapy. Finally, we reveal the CMM's dual-regulatory and multi-targeting functions on regulating TAMs, and hopefully, provide the theoretical basis for CMM clinical practice related to cancer therapy.

Notch signaling regulates expression of Mcl-1 and apoptosis in PPD-treated macrophages.

Science.gov (United States)

Palaga, Tanapat; Ratanabunyong, Siriluk; Pattarakankul, Thitiporn; Sangphech, Naunpun; Wongchana, Wipawee; Hadae, Yukihiro; Kueanjinda, Patipark

2013-09-01

Macrophages are cellular targets for infection by bacteria and viruses. The fate of infected macrophages plays a key role in determining the outcome of the host immune response. Apoptotic cell death of macrophages is considered to be a protective host defense that eliminates pathogens and infected cells. In this study, we investigated the involvement of Notch signaling in regulating apoptosis in macrophages treated with tuberculin purified protein derivative (PPD). Murine bone marrow-derived macrophages (BMMs) treated with PPD or infected with Mycobacterium bovis Bacillus Calmette-Guérin (BCG) induced upregulation of Notch1. This upregulation correlated well with the upregulation of the anti-apoptotic gene mcl-1 both at the transcriptional and translational levels. Decreased levels of Notch1 and Mcl-1 were observed in BMM treated with PPD when a gamma secretase inhibitor (GSI), which inhibits the processing of Notch receptors, was used. Moreover, silencing Notch1 in the macrophage-like cell line RAW264.7 decreased Mcl-1 protein expression, suggesting that Notch1 is critical for Mcl-1 expression in macrophages. A significant increase in apoptotic cells was observed upon treatment of BMM with PPD in the presence of GSI compared to the vehicle-control treated cells. Finally, analysis of the mcl-1 promoter in humans and mice revealed a conserved potential CSL/RBP-Jκ binding site. The association of Notch1 with the mcl-1 promoter was confirmed by chromatin immunoprecipitation. Taken together, these results indicate that Notch1 inhibits apoptosis of macrophages stimulated with PPD by directly controlling the mcl-1 promoter.

Orphan Nuclear Receptor ERRα Controls Macrophage Metabolic Signaling and A20 Expression to Negatively Regulate TLR-Induced Inflammation.

Science.gov (United States)

Yuk, Jae-Min; Kim, Tae Sung; Kim, Soo Yeon; Lee, Hye-Mi; Han, Jeongsu; Dufour, Catherine Rosa; Kim, Jin Kyung; Jin, Hyo Sun; Yang, Chul-Su; Park, Ki-Sun; Lee, Chul-Ho; Kim, Jin-Man; Kweon, Gi Ryang; Choi, Hueng-Sik; Vanacker, Jean-Marc; Moore, David D; Giguère, Vincent; Jo, Eun-Kyeong

2015-07-21

The orphan nuclear receptor estrogen-related receptor α (ERRα; NR3B1) is a key metabolic regulator, but its function in regulating inflammation remains largely unknown. Here, we demonstrate that ERRα negatively regulates Toll-like receptor (TLR)-induced inflammation by promoting Tnfaip3 transcription and fine-tuning of metabolic reprogramming in macrophages. ERRα-deficient (Esrra(-/-)) mice showed increased susceptibility to endotoxin-induced septic shock, leading to more severe pro-inflammatory responses than control mice. ERRα regulated macrophage inflammatory responses by directly binding the promoter region of Tnfaip3, a deubiquitinating enzyme in TLR signaling. In addition, Esrra(-/-) macrophages showed an increased glycolysis, but impaired mitochondrial respiratory function and biogenesis. Further, ERRα was required for the regulation of NF-κB signaling by controlling p65 acetylation via maintenance of NAD(+) levels and sirtuin 1 activation. These findings unravel a previously unappreciated role for ERRα as a negative regulator of TLR-induced inflammatory responses through inducing Tnfaip3 transcription and controlling the metabolic reprogramming. Copyright © 2015 Elsevier Inc. All rights reserved.

Prognostic significance of macrophage invasion in hilar cholangiocarcinoma

International Nuclear Information System (INIS)

Atanasov, Georgi; Hau, Hans-Michael; Dietel, Corinna; Benzing, Christian; Krenzien, Felix; Brandl, Andreas; Wiltberger, Georg; Matia, Ivan; Prager, Isabel; Schierle, Katrin; Robson, Simon C.; Reutzel-Selke, Anja; Pratschke, Johann; Schmelzle, Moritz; Jonas, Sven

2015-01-01

Tumor-associated macrophages (TAMs) promote tumor progression and have an effect on survival in human cancer. However, little is known regarding their influence on tumor progression and prognosis in human hilar cholangiocarcinoma. We analyzed surgically resected tumor specimens of hilar cholangiocarcinoma (n = 47) for distribution and localization of TAMs, as defined by expression of CD68. Abundance of TAMs was correlated with clinicopathologic characteristics, tumor recurrence and patients’ survival. Statistical analysis was performed using SPSS software. Patients with high density of TAMs in tumor invasive front (TIF) showed significantly higher local and overall tumor recurrence (both ρ < 0.05). Furthermore, high density of TAMs was associated with decreased overall (one-year 83.6 % vs. 75.1 %; three-year 61.3 % vs. 42.4 %; both ρ < 0.05) and recurrence-free survival (one-year 93.9 % vs. 57.4 %; three-year 59.8 % vs. 26.2 %; both ρ < 0.05). TAMs in TIF and tumor recurrence, were confirmed as the only independent prognostic variables in the multivariate survival analysis (all ρ < 0.05). Overall survival and recurrence free survival of patients with hilar cholangiocarcinoma significantly improved in patients with low levels of TAMs in the area of TIF, when compared to those with a high density of TAMs. These observations suggest their utilization as valuable prognostic markers in routine histopathologic evaluation, and might indicate future therapeutic approaches by targeting TAMs

Primary Cilium-Regulated EG-VEGF Signaling Facilitates Trophoblast Invasion.

Science.gov (United States)

Wang, Chia-Yih; Tsai, Hui-Ling; Syu, Jhih-Siang; Chen, Ting-Yu; Su, Mei-Tsz

2017-06-01

Trophoblast invasion is an important event in embryo implantation and placental development. During these processes, endocrine gland-derived vascular endothelial growth factor (EG-VEGF) is the key regulator mediating the crosstalk at the feto-maternal interface. The primary cilium is a cellular antenna receiving environmental signals and is crucial for proper development. However, little is known regarding the role of the primary cilium in early human pregnancy. Here, we demonstrate that EG-VEGF regulates trophoblast cell invasion via primary cilia. We found that EG-VEGF activated ERK1/2 signaling and subsequent upregulation of MMP2 and MMP9, thereby facilitating cell invasion in human trophoblast HTR-8/SVneo cells. Inhibition of ERK1/2 alleviated the expression of MMPs and trophoblast cell invasion after EG-VEGF treatment. In addition, primary cilia were observed in all the trophoblast cell lines tested and, more importantly, in human first-trimester placental tissue. The receptor of EG-VEGF, PROKR1, was detected in primary cilia. Depletion of IFT88, the intraflagellar transporter required for ciliogenesis, inhibited primary cilium growth, thereby ameliorating ERK1/2 activation, MMP upregulation, and trophoblast cell invasion promoted by EG-VEGF. These findings demonstrate a novel function of primary cilia in controlling EG-VEGF-regulated trophoblast invasion and reveal the underlying molecular mechanism. J. Cell. Physiol. 232: 1467-1477, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

GM-CSF and IL-4 produced by NKT cells inversely regulate IL-1β production by macrophages.

Science.gov (United States)

Ahn, Sehee; Jeong, Dongjin; Oh, Sae Jin; Ahn, Jiye; Lee, Seung Hyo; Chung, Doo Hyun

2017-02-01

Natural Killer T (NKT) cells are distinct T cell subset that link innate and adaptive immune responses. IL-1β, produced by various immune cells, plays a key role in the regulation of innate immunity in vivo. However, it is unclear whether NKT cells regulate IL-1β production by macrophages. To address this, we co-cultured NKT cells and peritoneal macrophages in the presence of TCR stimulation and inflammasome activators. Among cytokines secreted from NKT cells, GM-CSF enhanced IL-1β production by macrophages via regulating LPS-mediated pro-IL-1β expression and NLRP3-dependent inflammasome activation, whereas IL-4 enhanced M2-differentiation of macrophages and decreased IL-1β production. Together, our findings suggest the NKT cells have double-sided effects on IL-1β-mediated innate immune responses by producing IL-4 and GM-CSF. These findings may be helpful for a comprehensive understanding of NKT cell-mediated regulatory mechanisms of the pro-inflammatory effects of IL-1β in inflammatory diseases in vivo. Copyright © 2017 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

Hypoxic stress up-regulates the expression of Toll-like receptor 4 in macrophages via hypoxia-inducible factor.

Science.gov (United States)

Kim, So Young; Choi, Yong Jun; Joung, Sun Myung; Lee, Byung Ho; Jung, Yi-Sook; Lee, Joo Young

2010-04-01

Toll-like receptors (TLRs) are germline-encoded innate immune receptors that recognize invading micro-organisms and induce immune and inflammatory responses. Deregulation of TLRs is known to be closely linked to various immune disorders and inflammatory diseases. Cells at sites of inflammation are exposed to hypoxic stress, which further aggravates inflammatory processes. We have examined if hypoxic stress modulates the TLR activity of macrophages. Hypoxia and CoCl(2) (a hypoxia mimetic) enhanced the expression of TLR4 messenger RNA and protein in macrophages (RAW264.7 cells), whereas the messenger RNA of other TLRs was not increased. To determine the underlying mechanism, we investigated the role of hypoxia-inducible factor 1 (HIF-1) in the regulation of TLR4 expression. Knockdown of HIF-1alpha expression by small interfering RNA inhibited hypoxia-induced and CoCl(2)-induced TLR4 expression in macrophages, while over-expression of HIF-1alpha potentiated TLR4 expression. Chromatin immunoprecipitation assays revealed that HIF-1alpha binds to the TLR4 promoter region under hypoxic conditions. In addition, deletion or mutation of a putative HIF-1-binding motif in the TLR4 promoter greatly attenuated HIF-1alpha-induced TLR4 promoter reporter expression. Up-regulation of TLR4 expression by hypoxic stress enhanced the response of macrophages to lipopolysaccharide, resulting in increased expression of cyclooxygenase-2, interleukin-6, regulated on activation normal T cell expressed and secreted, and interferon-inducible protein-10. These results demonstrate that TLR4 expression in macrophages is up-regulated via HIF-1 in response to hypoxic stress, suggesting that hypoxic stress at sites of inflammation enhances susceptibility to subsequent infection and inflammatory signals by up-regulating TLR4.

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

Directory of Open Access Journals (Sweden)

Norifumi Urao

2010-10-01

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

The AP-1 Transcription Factor c-Jun Promotes Arthritis by Regulating Cyclooxygenase-2 and Arginase-1 Expression in Macrophages.

Science.gov (United States)

Hannemann, Nicole; Jordan, Jutta; Paul, Sushmita; Reid, Stephen; Baenkler, Hanns-Wolf; Sonnewald, Sophia; Bäuerle, Tobias; Vera, Julio; Schett, Georg; Bozec, Aline

2017-05-01

Activation of proinflammatory macrophages is associated with the inflammatory state of rheumatoid arthritis. Their polarization and activation are controlled by transcription factors such as NF-κB and the AP-1 transcription factor member c-Fos. Surprisingly, little is known about the role of the AP-1 transcription factor c-Jun in macrophage activation. In this study, we show that mRNA and protein levels of c-Jun are increased in macrophages following pro- or anti-inflammatory stimulations. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment cluster analyses of microarray data using wild-type and c-Jun-deleted macrophages highlight the central function of c-Jun in macrophages, in particular for immune responses, IL production, and hypoxia pathways. Mice deficient for c-Jun in macrophages show an amelioration of inflammation and bone destruction in the serum-induced arthritis model. In vivo and in vitro gene profiling, together with chromatin immunoprecipitation analysis of macrophages, revealed direct activation of the proinflammatory factor cyclooxygenase-2 and indirect inhibition of the anti-inflammatory factor arginase-1 by c-Jun. Thus, c-Jun regulates the activation state of macrophages and promotes arthritis via differentially regulating cyclooxygenase-2 and arginase-1 levels. Copyright © 2017 by The American Association of Immunologists, Inc.

IL-4 Receptor Alpha Signaling through Macrophages Differentially Regulates Liver Fibrosis Progression and Reversal

Directory of Open Access Journals (Sweden)

Shih-Yen Weng

2018-03-01

Full Text Available Chronic hepatitis leads to liver fibrosis and cirrhosis. Cirrhosis is a major cause of worldwide morbidity and mortality. Macrophages play a key role in fibrosis progression and reversal. However, the signals that determine fibrogenic vs fibrolytic macrophage function remain ill defined. We studied the role of interleukin-4 receptor α (IL-4Rα, a potential central switch of macrophage polarization, in liver fibrosis progression and reversal. We demonstrate that inflammatory monocyte infiltration and liver fibrogenesis were suppressed in general IL-4Rα−/− as well as in macrophage-specific IL-4Rα−/− (IL-4RαΔLysM mice. However, with deletion of IL-4RαΔLysM spontaneous fibrosis reversal was retarded. Results were replicated by pharmacological intervention using IL-4Rα-specific antisense oligonucleotides. Retarded resolution was linked to the loss of M2-type resident macrophages, which secreted MMP-12 through IL-4 and IL-13-mediated phospho-STAT6 activation. We conclude that IL-4Rα signaling regulates macrophage functional polarization in a context-dependent manner. Pharmacological targeting of macrophage polarization therefore requires disease stage-specific treatment strategies. Research in Context: Alternative (M2-type macrophage activation through IL-4Rα promotes liver inflammation and fibrosis progression but speeds up fibrosis reversal. This demonstrates context dependent, opposing roles of M2-type macrophages. During reversal IL-4Rα induces fibrolytic MMPs, especially MMP-12, through STAT6. Liver-specific antisense oligonucleotides efficiently block IL-4Rα expression and attenuate fibrosis progression. Keywords: Fibrosis, IL-4 receptor alpha, Liver, Macrophage, MMP12, Progression, Reversal

MicroRNA-365 in macrophages regulates Mycobacterium tuberculosis-induced active pulmonary tuberculosis via interleukin-6.

Science.gov (United States)

Song, Qingzhang; Li, Hui; Shao, Hua; Li, Chunling; Lu, Xiao

2015-01-01

The present study is to investigate the relationship between microRNA (miR)-365 expression and the levels of interleukin (IL)-6 mRNA and protein in patients with active tuberculosis. From June 2011 to June 2014, 48 patients with active pulmonary tuberculosis induced by Mycobacterium tuberculosis were included in the study. In addition, 23 healthy subjects were enrolled as control. Macrophages were collected by pulmonary alveolus lavage. In addition, serum and mononuclear cells were isolated from peripheral blood. The levels of miR-365 and IL-6 in macrophages, mononuclear cells and serum were determined using quantitative real-time polymerase chain reaction. The protein expression of IL-6 in macrophages and mononuclear cells was measured using Western blotting, while that in serum was detected by enzyme-linked immunoabsorbent assay. Expression of IL-6 mRNA and protein was significantly enhanced in patients with active pulmonary tuberculosis. Increase of IL-6 protein concentration in serum was probably due to the release of IL-6 protein from mononuclear cells in the blood. In addition, miR-365 levels were significantly lowered in patients with active pulmonary tuberculosis. Up-regulated IL-6 expression in macrophages, mononuclear cells and serum in patients with active pulmonary tuberculosis is related to the down-regulation of miR-365, suggesting that miR-365 may regulate the occurrence and immune responses of active pulmonary tuberculosis via IL-6.

Nitric oxide production from macrophages is regulated by arachidonic acid metabolites.

Science.gov (United States)

Imai, Y; Kolb, H; Burkart, V

1993-11-30

In activated macrophages the inducible form of the enzyme nitric oxide (NO) synthase generates high amounts of the toxic mediator NO. After 20 h of treatment with LPS rat peritoneal macrophages release 12-16 nmol NO2-/10(5) cells which is detectable in the culture supernatant by the Griess reaction as a measure of NO formation. The addition of aminoguanidine (1 mM), a preferential inhibitor of the inducible NO-synthase, completely abolished NO2-accumulation. Incubation with indomethacin or acetyl-salicylic acid, preferential inhibitors of the cyclooxygenase pathway of the arachidonic acid metabolism, did not influence NO2- levels. Nordihydro-guaiaretic acid (50 microM), a preferential inhibitor of the lipoxygenase pathway, caused strong reduction of NO2- accumulation to 1.9 +/- 0.3 nmol/200 microliter. Simultaneous inhibition of cyclo- and lipoxygenase by BW755c resulted in an intermediate effect (7.3 +/- 1.1 nmol/200 microliter NO2-). These results show that the induction of NO production in activated macrophages is regulated by products of the lipoxygenase-pathway of the arachidonic acid metabolism.

The Fps/Fes kinase regulates the inflammatory response to endotoxin through down-regulation of TLR4, NF-kappaB activation, and TNF-alpha secretion in macrophages.

Science.gov (United States)

Parsons, Sean A; Greer, Peter A

2006-12-01

Fps/Fes and Fer are members of a distinct subfamily of cytoplasmic protein tyrosine kinases that have recently been implicated in the regulation of innate immunity. Previous studies showed that mice lacking Fps/Fes are hypersensitive to systemic LPS challenge, and Fer-deficient mice displayed enhanced recruitment of leukocytes in response to local LPS challenge. This study identifies physiological, cellular, and molecular defects that contribute to the hyperinflammatory phenotype in Fps/Fes null mice. Plasma TNF-alpha levels were elevated in LPS challenged Fps/Fes null mice as compared with wild-type mice and cultured Fps/Fes null peritoneal macrophages treated with LPS showed increased TNF-alpha production. Cultured Fps/Fes null macrophages also displayed prolonged LPS-induced degradation of IkappaB-alpha, increased phosphorylation of the p65 subunit of NF-kappaB, and defective TLR4 internalization, compared with wild-type macrophages. Together, these observations provide a likely mechanistic basis for elevated proinflammatory cytokine secretion by Fps/Fes null macrophages and the increased sensitivity of Fps/Fes null mice to endotoxin. We posit that Fps/Fes modulates the innate immune response of macrophages to LPS, in part, by regulating internalization and down-regulation of the TLR4 receptor complex.

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

Science.gov (United States)

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

2012-02-01

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

miR-181a Induces Macrophage Polarized to M2 Phenotype and Promotes M2 Macrophage-mediated Tumor Cell Metastasis by Targeting KLF6 and C/EBPα

Directory of Open Access Journals (Sweden)

Jia Bi

2016-01-01

Full Text Available Macrophages can acquire a variety of polarization status and functions: classically activated macrophages (M1 macrophages; alternatively activated macrophages (M2 macrophages. However, the molecular basis of the process is still unclear. Here, this study addresses that microRNA-181a (miR-181a is a key molecule controlling macrophage polarization. We found that miR-181a is overexpressed in M2 macrophages than in M1 macrophages. miR-181a expression was decreased when M2 phenotype converted to M1, whereas it increased when M1 phenotype converted to M2. Overexpression of miR-181a in M1 macrophages diminished M1 phenotype expression while promoting polarization to the M2 phenotype. In contrast, knockdown of miR-181a in M2 macrophages promoted M1 polarization and diminished M2 phenotype expression. Mechanistically, Bioinformatic analysis revealed that Kruppel-like factor 6 (KLF6 and CCAAT/enhancer binding protein-α (C/EBPα is a potential target of miR-181a and luciferase assay confirmed that KLF6 and C/EBPα translation is suppressed by miR-181a through interaction with the 3′UTR of KLF6 and C/EBPα mRNA. Further analysis showed that induction of miR-181a suppressed KLF6 and C/EBPα protein expression. Importantly, miR-181a also diminishes M2 macrophages-mediated migration and invasion capacity of tumor cells. Collectively, our results suggest that miR-181a plays a significant role in regulating macrophage polarization through directly target KLF6 and C/EBPα.

Macrophage/epithelium cross-talk regulates cell cycle progression and migration in pancreatic progenitors.

Directory of Open Access Journals (Sweden)

Kristin Mussar

Full Text Available Macrophages populate the mesenchymal compartment of all organs during embryogenesis and have been shown to support tissue organogenesis and regeneration by regulating remodeling of the extracellular microenvironment. Whether this mesenchymal component can also dictate select developmental decisions in epithelia is unknown. Here, using the embryonic pancreatic epithelium as model system, we show that macrophages drive the epithelium to execute two developmentally important choices, i.e. the exit from cell cycle and the acquisition of a migratory phenotype. We demonstrate that these developmental decisions are effectively imparted by macrophages activated toward an M2 fetal-like functional state, and involve modulation of the adhesion receptor NCAM and an uncommon "paired-less" isoform of the transcription factor PAX6 in the epithelium. Over-expression of this PAX6 variant in pancreatic epithelia controls both cell motility and cell cycle progression in a gene-dosage dependent fashion. Importantly, induction of these phenotypes in embryonic pancreatic transplants by M2 macrophages in vivo is associated with an increased frequency of endocrine-committed cells emerging from ductal progenitor pools. These results identify M2 macrophages as key effectors capable of coordinating epithelial cell cycle withdrawal and cell migration, two events critical to pancreatic progenitors' delamination and progression toward their differentiated fates.

Macrophage/epithelium cross-talk regulates cell cycle progression and migration in pancreatic progenitors.

Science.gov (United States)

Mussar, Kristin; Tucker, Andrew; McLennan, Linsey; Gearhart, Addie; Jimenez-Caliani, Antonio J; Cirulli, Vincenzo; Crisa, Laura

2014-01-01

Macrophages populate the mesenchymal compartment of all organs during embryogenesis and have been shown to support tissue organogenesis and regeneration by regulating remodeling of the extracellular microenvironment. Whether this mesenchymal component can also dictate select developmental decisions in epithelia is unknown. Here, using the embryonic pancreatic epithelium as model system, we show that macrophages drive the epithelium to execute two developmentally important choices, i.e. the exit from cell cycle and the acquisition of a migratory phenotype. We demonstrate that these developmental decisions are effectively imparted by macrophages activated toward an M2 fetal-like functional state, and involve modulation of the adhesion receptor NCAM and an uncommon "paired-less" isoform of the transcription factor PAX6 in the epithelium. Over-expression of this PAX6 variant in pancreatic epithelia controls both cell motility and cell cycle progression in a gene-dosage dependent fashion. Importantly, induction of these phenotypes in embryonic pancreatic transplants by M2 macrophages in vivo is associated with an increased frequency of endocrine-committed cells emerging from ductal progenitor pools. These results identify M2 macrophages as key effectors capable of coordinating epithelial cell cycle withdrawal and cell migration, two events critical to pancreatic progenitors' delamination and progression toward their differentiated fates.

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

Directory of Open Access Journals (Sweden)

Tapas K. Nayak

2017-01-01

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

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

Science.gov (United States)

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

2017-01-06

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

Post-transcriptional regulation of macrophage ABCA1, an early response gene to IFN-γ

International Nuclear Information System (INIS)

Alfaro Leon, Martha Leticia; Evans, Glenn F.; Farmen, Mark W.; Zuckerman, Steven H.

2005-01-01

Interferon-γ (IFN-γ) down-regulates receptors associated with reverse cholesterol transport including ABCA1. In the present study, the kinetics and mechanism of ABCA1 down-regulation were determined in mouse peritoneal macrophages. IFN-γ decreased ABCA1 mRNA 1 h following IFN-γ addition and was maximally reduced by 3 h. Down-regulation was protein synthesis dependent and involved post-transcriptional processes. ABCA1 message had a T 1/2 of 115 min in actinomycin treated cells that was reduced to a T 1/2 of 37 min by IFN-γ. The decrease in message stability was also associated with a rapid loss of ABCA1 protein, significant 3 h following IFN-γ addition. The kinetics of ABCA1 message and protein decrease was consistent with the early IFN-γ-induced changes in Stat1 phosphorylation and nuclear translocation observed in these cells. Therefore, ABCA1 can be considered as an early response gene to macrophage activation by IFN-γ with down-regulation occurring by message destabilization

BMP pathway regulation of and by macrophages.

Directory of Open Access Journals (Sweden)

Megha Talati

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

TRPV4 calcium-permeable channel is a novel regulator of oxidized LDL-induced macrophage foam cell formation.

Science.gov (United States)

Goswami, Rishov; Merth, Michael; Sharma, Shweta; Alharbi, Mazen O; Aranda-Espinoza, Helim; Zhu, Xiaoping; Rahaman, Shaik O

2017-09-01

Cardiovascular disease is the number one cause of death in United States, and atherosclerosis, a chronic inflammatory arterial disease, is the most dominant underlying pathology. Macrophages are thought to orchestrate atherosclerosis by generating lipid-laden foam cells and by secreting inflammatory mediators. Emerging data support a role for a mechanical factor, e.g., matrix stiffness, in regulation of macrophage function, vascular elasticity, and atherogenesis. However, the identity of the plasma membrane mechanosensor and the mechanisms by which pro-atherogenic signals are transduced/maintained are unknown. We have obtained evidence that TRPV4, an ion channel in the transient receptor potential vanilloid family and a known mechanosensor, is the likely mediator of oxidized low-density lipoprotein (oxLDL)-dependent macrophage foam cell formation, a critical process in atherogenesis. Specifically, we found that: i) genetic ablation of TRPV4 or pharmacologic inhibition of TRPV4 activity by a specific antagonist blocked oxLDL-induced macrophage foam cell formation, and ii) TRPV4 deficiency prevented pathophysiological range matrix stiffness or scratch-induced exacerbation of oxLDL-induced foam cell formation. Mechanistically, we found that: i) plasma membrane localization of TRPV4 was sensitized to the increasing level of matrix stiffness, ii) lack of foam cell formation in TRPV4 null cells was not due to lack of expression of CD36, a major receptor for oxLDL, and iii) TRPV4 channel activity regulated oxLDL uptake but not its binding on macrophages. Altogether, these findings identify a novel role for TRPV4 in regulating macrophage foam cell formation by modulating uptake of oxLDL. These findings suggest that therapeutic targeting of TRPV4 may provide a selective approach to the treatment of atherosclerosis. Copyright © 2017. Published by Elsevier Inc.

Increased Tim-3 expression alleviates liver injury by regulating macrophage activation in MCD-induced NASH mice.

Science.gov (United States)

Du, Xianhong; Wu, Zhuanchang; Xu, Yong; Liu, Yuan; Liu, Wen; Wang, Tixiao; Li, Chunyang; Zhang, Cuijuan; Yi, Fan; Gao, Lifen; Liang, Xiaohong; Ma, Chunhong

2018-05-07

As an immune checkpoint, Tim-3 plays roles in the regulation of both adaptive and innate immune cells including macrophages and is greatly involved in chronic liver diseases. However, the precise roles of Tim-3 in nonalcoholic steatohepatitis (NASH) remain unstated. In the current study, we analyzed Tim-3 expression on different subpopulations of liver macrophages and further investigated the potential roles of Tim-3 on hepatic macrophages in methionine and choline-deficient diet (MCD)-induced NASH mice. The results of flow cytometry demonstrated the significantly increased expression of Tim-3 on all detected liver macrophage subsets in MCD mice, including F4/80 + CD11b + , F4/80 + CD68 + , and F4/80 + CD169 + macrophages. Remarkably, Tim-3 knockout (KO) significantly accelerated MCD-induced liver steatosis, displaying higher serum ALT, larger hepatic vacuolation, more liver lipid deposition, and more severe liver fibrosis. Moreover, compared with wild-type C57BL/6 mice, Tim-3 KO MCD mice demonstrated an enhanced expression of NOX2, NLRP3, and caspase-1 p20 together with increased generation of IL-1β and IL-18 in livers. In vitro studies demonstrated that Tim-3 negatively regulated the production of reactive oxygen species (ROS) and related downstream pro-inflammatory cytokine secretion of IL-1β and IL-18 in macrophages. Exogenous administration of N-Acetyl-L-cysteine (NAC), a small molecular inhibitor of ROS, remarkably suppressed caspase-1 p20 expression and IL-1β and IL-18 production in livers of Tim-3 KO mice, thus significantly reducing the severity of steatohepatitis induced by MCD. In conclusion, Tim-3 is a promising protector in MCD-induced steatohepatitis by controlling ROS and the associated pro-inflammatory cytokine production in macrophages.

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

Directory of Open Access Journals (Sweden)

Chandra L Shrestha

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

The role of PTEN in regulation of hepatic macrophages activation and function in progression and reversal of liver fibrosis

Energy Technology Data Exchange (ETDEWEB)

Cheng, Yahui; Tian, Yuanyao; Xia, Jialu; Wu, Xiaoqin; Yang, Yang; Li, Xiaofeng; Huang, Cheng; Meng, Xiaoming; Ma, Taotao; Li, Jun, E-mail: lj@ahmu.edu.cn

2017-02-15

Activation of Kupffer cells (KCs) plays a pivotal role in the pathogenesis of liver fibrosis. The progression and reversal of CCl{sub 4}-induced mouse liver fibrosis showed a mixed induction of hepatic classical (M1) and alternative (M2) macrophage markers. Although the role of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in modulating myeloid cell activation has recently been identified, its function in macrophage activation during hepatic fibrosis remains to be fully appreciated. In our study, PTEN expression of KCs was remarkably decreased in CCl{sub 4}-induced mice but increased to a near-normal level in reversed mice. Moreover, PTEN was significantly decreased in IL4-induced RAW 264.7 cells in vitro and lower expression of PTEN was observed in M2 macrophages in vivo. In addition, loss- and gain-of-function studies suggested that PTEN regulates M2 macrophages polarization via activation of PI3K/Akt/STAT6 signaling, but had a limited effect on M1 macrophages polarization in vitro. Additionally, Ly294002, a chemical inhibitor of PI3K/Akt, could dramatically down-regulate the hallmarks of M2 macrophages. In conclusion, PTEN mediates macrophages activation by PI3K/Akt/STAT6 signaling pathway, which provides novel compelling evidences on the potential of PTEN in liver injury and opens new cellular target for the pharmacological therapy of liver fibrosis. - Highlights: • CCl{sub 4} treatment triggered a mixed M1/M2 macrophage phenotype in fibrosis. • Lower expression of PTEN in murine M2 macrophages in vivo and vitro. • PTEN modulates M2 macrophages activation via PI3K/Akt/STAT6 signaling. • Provide a new cellular target modulate macrophage mediated hepatic fibrosis.

Biology of Bony Fish Macrophages

Directory of Open Access Journals (Sweden)

Jordan W. Hodgkinson

2015-11-01

Full Text Available Macrophages are found across all vertebrate species, reside in virtually all animal tissues, and play critical roles in host protection and homeostasis. Various mechanisms determine and regulate the highly plastic functional phenotypes of macrophages, including antimicrobial host defenses (pro-inflammatory, M1-type, and resolution and repair functions (anti-inflammatory/regulatory, M2-type. The study of inflammatory macrophages in immune defense of teleosts has garnered much attention, and antimicrobial mechanisms of these cells have been extensively studied in various fish models. Intriguingly, both similarities and differences have been documented for the regulation of lower vertebrate macrophage antimicrobial defenses, as compared to what has been described in mammals. Advances in our understanding of the teleost macrophage M2 phenotypes likewise suggest functional conservation through similar and distinct regulatory strategies, compared to their mammalian counterparts. In this review, we discuss the current understanding of the molecular mechanisms governing teleost macrophage functional heterogeneity, including monopoetic development, classical macrophage inflammatory and antimicrobial responses as well as alternative macrophage polarization towards tissues repair and resolution of inflammation.

Biology of Bony Fish Macrophages.

Science.gov (United States)

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

2015-11-30

Macrophages are found across all vertebrate species, reside in virtually all animal tissues, and play critical roles in host protection and homeostasis. Various mechanisms determine and regulate the highly plastic functional phenotypes of macrophages, including antimicrobial host defenses (pro-inflammatory, M1-type), and resolution and repair functions (anti-inflammatory/regulatory, M2-type). The study of inflammatory macrophages in immune defense of teleosts has garnered much attention, and antimicrobial mechanisms of these cells have been extensively studied in various fish models. Intriguingly, both similarities and differences have been documented for the regulation of lower vertebrate macrophage antimicrobial defenses, as compared to what has been described in mammals. Advances in our understanding of the teleost macrophage M2 phenotypes likewise suggest functional conservation through similar and distinct regulatory strategies, compared to their mammalian counterparts. In this review, we discuss the current understanding of the molecular mechanisms governing teleost macrophage functional heterogeneity, including monopoetic development, classical macrophage inflammatory and antimicrobial responses as well as alternative macrophage polarization towards tissues repair and resolution of inflammation.

Tumor-associated macrophages as a paradigm of macrophage plasticity, diversity, and polarization: lessons and open questions.

Science.gov (United States)

Mantovani, Alberto; Locati, Massimo

2013-07-01

Macrophages are present in all body compartments, including cancerous tissues, and their functions are profoundly affected by signals from the microenvironment under homeostatic and pathological conditions. Tumor-associated macrophages are a major cellular component of cancer-related inflammation and have served as a paradigm for the plasticity and functional polarization of mononuclear phagocytes. Tumor-associated macrophages can exert dual influence of cancer depending on the activation state, with classically activated (M1) and alternatively activated (M2) cells generally exerting antitumoral and protumoral functions, respectively. These are extremes in a continuum of polarization states in a universe of diversity. Tumor-associated macrophages affect virtually all aspects of tumor tissues, including stem cells, metabolism, angiogenesis, invasion, and metastasis. Progress has been made in defining signaling molecules, transcription factors, epigenetic changes, and repertoire of microRNAs underlying macrophage polarization. Preclinical and early clinical data suggest that macrophages may serve as tools for the development of innovative diagnostic and therapeutic strategies in cancer and chronic nonresolving inflammatory diseases.

Immune invasion of the central nervous system parenchyma and experimental allergic encephalomyelitis, but not leukocyte extravasation from blood, are prevented in macrophage-depleted mice

DEFF Research Database (Denmark)

Tran, E H; Hoekstra, K; van Rooijen, N

1998-01-01

role of peripheral macrophages in experimental allergic encephalomyelitis (EAE), a Th1-mediated demyelinating disease that serves as a an animal model for multiple sclerosis (MS), by their depletion using mannosylated liposome-encapsulated dichloromethylene diphosphonate (Cl2MDP). Here we describe....../J mice was abrogated by Cl2MDP-mnL treatment. CD4+ T cell and MHC II+ B220+ B cell extravasation from blood vessels and Th1 cytokine production were not inhibited. However, invasion of the central nervous system intraparenchymal tissues by lymphocytes, F4/80+, Mac-1+, and MOMA-1+ macrophages was almost...

A methanolic extract of Trigonella foenum-graecum (fenugreek seeds regulates markers of macrophage polarization.

Directory of Open Access Journals (Sweden)

Nurudeen Hassan

2015-12-01

Full Text Available Background: Macrophages are key cellular mediators in diabetes-related inflammation. Molecular cues such as cytokines found in the tissue microenvironment regulates the polarization of macrophages into an M1 (pro-inflammatory or M2 (immunoregulatory phenotype. Recent evidence suggests that M1 macrophages in diabetic patients may contribute to the complications associated with the disease such as atherosclerosis. Trigonella foenum- graecum (Tfg: fenugreek seeds have been used in traditional medicine in Asia, Africa and the Middle-East for their alleged anti-diabetic properties. Objective: To identify the molecular mechanism(s through which Tfg seeds exert their effects, we investigated the role of a crude methanolic extract of Tfg (FME seeds on macrophage polarization in vitro. Method: THP-1 macrophages (Mϕ were treated with gBSA in the presence/absence of FME and the release and expression of M1 and M2 markers/cytokines were analysed. The role of FME on NF-κB activity was also explored using transfected HEK-293T cells. Results: This study found that the FME significantly (P<0.05 decreased gBSA-induced secretion of M1 cytokines (TNF-α, IL-1β, IL-6 and IL-8 in THP-1 Mϕ cells. In the presence of gBSA, FME also significantly increased the gene expression of the M2 marker Dectin-1, but had no effect on IL-10, IL-1Ra. FME also significantly decreased TNF-α induced NF-kB reporter activity. Conclusion: These results suggest that FME can regulate the expression of M1 and M2 markers in THP-1 Mϕ cells. This may be potentially through the modulation of NF-kB activity. Further work should be carried out to identify precise mechanism(s involved in the effects of FME and Tfg seeds.

Matrix metalloproteases as maestros for the dual role of LPS- and IL-10-stimulated macrophages in cancer cell behaviour

International Nuclear Information System (INIS)

Cardoso, Ana P.; Pinto, Marta L.; Pinto, Ana T.; Pinto, Marta T.; Monteiro, Cátia; Oliveira, Marta I.; Santos, Susana G.; Relvas, João B.; Seruca, Raquel; Mantovani, Alberto; Mareel, Marc; Barbosa, Mário A.; Oliveira, Maria J.

2015-01-01

The interactions established between macrophages and cancer cells are largely dependent on instructions from the tumour microenvironment. Macrophages may differentiate into populations with distinct inflammatory profiles, but knowledge on their role on cancer cell activities is still very scarce. In this work, we investigated the influence of pro-inflammatory (LPS-stimulated) and anti-inflammatory (IL-10-stimulated) macrophages on gastric and colorectal cancer cell invasion, motility/migration, angiogenesis and proteolysis, and the associated molecular mechanisms. Following exposure of gastric and colon cancer cell lines to LPS- and IL-10-stimulated human macrophages, either by indirect contact or conditioned media, we analyzed the effect of the different macrophage populations on cancer cell invasion, migration, motility and phosphorylation status of EGFR and several interacting partners. Cancer-cell induced angiogenesis upon the influence of conditioned media from both macrophage populations was assessed using the chick embryo chorioallantoic membrane assay. MMP activities were evaluated by gelatin zymograhy. Our results show that IL-10-stimulated macrophages are more efficient in promoting in vitro cancer cell invasion and migration. In addition, soluble factors produced by these macrophages enhanced in vivo cancer cell-induced angiogenesis, as opposed to their LPS-stimulated counterparts. We further demonstrate that differences in the ability of these macrophage populations to stimulate invasion or angiogenesis cannot be explained by the EGFR-mediated signalling, since both LPS- and IL-10-stimulated macrophages similarly induce the phosphorylation of cancer cell EGFR, c-Src, Akt, ERK1/2, and p38. Interestingly, both populations exert distinct proteolytic activities, being the IL-10-stimulated macrophages the most efficient in inducing matrix metalloprotease (MMP)-2 and MMP-9 activities. Using a broad-spectrum MMP inhibitor, we demonstrated that proteolysis was

miRNA let-7b modulates macrophage polarization and enhances tumor-associated macrophages to promote angiogenesis and mobility in prostate cancer.

Science.gov (United States)

Wang, Zhigang; Xu, Lu; Hu, Yinying; Huang, Yanqin; Zhang, Yujuan; Zheng, Xiufen; Wang, Shanshan; Wang, Yifan; Yu, Yanrong; Zhang, Meng; Yuan, Keng; Min, Weiping

2016-05-09

Macrophage polarization is a highly plastic physiological process that responds to a variety of environmental factors by changing macrophage phenotype and function. Tumor-associated macrophages (TAMs) are generally recognized as promoting tumor progression. As universal regulators, microRNAs (miRNAs) are functionally involved in numerous critical cellular processes including macrophage polarization. Let-7b, a miRNA, has differential expression patterns in inflamed tissues compared with healthy controls. However, whether and how miRNA let-7b regulates macrophage phenotype and function is unclear. In this report, we find that up-regulation of let-7b is characteristic of prostatic TAMs, and down-regulation of let-7b in TAMs leads to changes in expression profiles of inflammatory cytokines, such as IL-12, IL-23, IL-10 and TNF-α. As a result, TAMs treated with let-7b inhibitors reduce angiogenesis and prostate carcinoma (PCa) cell mobility. Let-7b may play a vital role in regulating macrophage polarization, thus modulating the prognosis of prostate cancer.

MAPK/p38 regulation of cytoskeleton rearrangement accelerates induction of macrophage activation by TLR4, but not TLR3.

Science.gov (United States)

Bian, Hongjun; Li, Feifei; Wang, Wenwen; Zhao, Qi; Gao, Shanshan; Ma, Jincai; Li, Xiao; Ren, Wanhua; Qin, Chengyong; Qi, Jianni

2017-11-01

Toll-like receptor 3 (TLR3) and TLR4 utilize adaptor proteins to activate mitogen‑activated protein kinase (MAPK), resulting in the acute but transient inflammatory response aimed at the clearance of pathogens. In the present study, it was demonstrated that macrophage activation by lipopolysaccharide (LPS) or poly(I:C), leading to changes in cell morphology, differed significantly between the mouse macrophage cell line RAW264.7 and mouse primary peritoneal macrophages. Moreover, the expression of α- and β-tubulin was markedly decreased following LPS stimulation. By contrast, α- and β-tubulin expression were only mildly increased following poly(I:C) treatment. However, the expression of β-actin and GAPDH was not significantly affected. Furthermore, it was verified that vincristine pretreatment abrogated the cytoskeleton rearrangement and decreased the synthesis and secretion of proinflammatory cytokines and migration of macrophages caused by LPS. Finally, it was observed that the MAPK/p38 signaling pathway regulating cytoskeleton rearrangement may participate in LPS‑induced macrophage cytokine production and migration. Overall, the findings of the present study indicated that MAPK/p38 regulation of the cytoskeleton, particularly tubulin proteins, plays an important role in LPS-induced inflammatory responses via alleviating the synthesis and secretion of proinflammatory cytokines and inhibiting the migration of macrophages.

Trichomonas vaginalis homolog of macrophage migration inhibitory factor induces prostate cell growth, invasiveness, and inflammatory responses.

Science.gov (United States)

Twu, Olivia; Dessí, Daniele; Vu, Anh; Mercer, Frances; Stevens, Grant C; de Miguel, Natalia; Rappelli, Paola; Cocco, Anna Rita; Clubb, Robert T; Fiori, Pier Luigi; Johnson, Patricia J

2014-06-03

The human-infective parasite Trichomonas vaginalis causes the most prevalent nonviral sexually transmitted infection worldwide. Infections in men may result in colonization of the prostate and are correlated with increased risk of aggressive prostate cancer. We have found that T. vaginalis secretes a protein, T. vaginalis macrophage migration inhibitory factor (TvMIF), that is 47% similar to human macrophage migration inhibitory factor (HuMIF), a proinflammatory cytokine. Because HuMIF is reported to be elevated in prostate cancer and inflammation plays an important role in the initiation and progression of cancers, we have explored a role for TvMIF in prostate cancer. Here, we show that TvMIF has tautomerase activity, inhibits macrophage migration, and is proinflammatory. We also demonstrate that TvMIF binds the human CD74 MIF receptor with high affinity, comparable to that of HuMIF, which triggers activation of ERK, Akt, and Bcl-2-associated death promoter phosphorylation at a physiologically relevant concentration (1 ng/mL, 80 pM). TvMIF increases the in vitro growth and invasion through Matrigel of benign and prostate cancer cells. Sera from patients infected with T. vaginalis are reactive to TvMIF, especially in males. The presence of anti-TvMIF antibodies indicates that TvMIF is released by the parasite and elicits host immune responses during infection. Together, these data indicate that chronic T. vaginalis infections may result in TvMIF-driven inflammation and cell proliferation, thus triggering pathways that contribute to the promotion and progression of prostate cancer.

Bioelectric modulation of macrophage polarization

Science.gov (United States)

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

2016-02-01

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

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

Directory of Open Access Journals (Sweden)

Katrin Paulsen

2015-01-01

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

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

Science.gov (United States)

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

2016-09-01

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

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

International Nuclear Information System (INIS)

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

2008-01-01

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

CK2 Secreted by Leishmania braziliensis Mediates Macrophage Association Invasion: A Comparative Study between Virulent and Avirulent Promastigotes

Directory of Open Access Journals (Sweden)

Ana Madeira Brito Zylbersztejn

2015-01-01

Full Text Available CK2 is a protein kinase distributed in different compartments of Leishmania braziliensis: an externally oriented ecto-CK2, an intracellular CK2, and a secreted CK2. This latter form is constitutively secreted from the parasite (CsCK2, but such secretion may be highly enhanced by the association of specific molecules, including enzyme substrates, which lead to a higher enzymatic activity, called inductively secreted CK2 (IsCK2. Here, we examined the influence of secreted CK2 (sCK2 activity on the infectivity of a virulent L. braziliensis strain. The virulent strain presented 121-fold higher total CK2 activity than those found in an avirulent strain. The use of specific CK2 inhibitors (TBB, DRB, or heparin inhibited virulent parasite growth, whereas no effect was observed in the avirulent parasites. When these inhibitors were added to the interaction assays between the virulent L. braziliensis strain and macrophages, association index was drastically inhibited. Polyamines enhanced sCK2 activity and increased the association index between parasites and macrophages. Finally, sCK2 and the supernatant of the virulent strain increased the association index between the avirulent strain and macrophages, which was inhibited by TBB. Thus, the kinase enzyme CK2 seems to be important to invasion mechanisms of L. braziliensis.

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

Science.gov (United States)

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

2017-01-01

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

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

Directory of Open Access Journals (Sweden)

Purcell Damian FJ

2008-02-01

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

Pathogenicity of Mycobacterium tuberculosis is expressed by regulating metabolic thresholds of the host macrophage.

Directory of Open Access Journals (Sweden)

Parul Mehrotra

2014-07-01

Full Text Available The success of Mycobacterium tuberculosis as a pathogen derives from its facile adaptation to the intracellular milieu of human macrophages. To explore this process, we asked whether adaptation also required interference with the metabolic machinery of the host cell. Temporal profiling of the metabolic flux, in cells infected with differently virulent mycobacterial strains, confirmed that this was indeed the case. Subsequent analysis identified the core subset of host reactions that were targeted. It also elucidated that the goal of regulation was to integrate pathways facilitating macrophage survival, with those promoting mycobacterial sustenance. Intriguingly, this synthesis then provided an axis where both host- and pathogen-derived factors converged to define determinants of pathogenicity. Consequently, whereas the requirement for macrophage survival sensitized TB susceptibility to the glycemic status of the individual, mediation by pathogen ensured that the virulence properties of the infecting strain also contributed towards the resulting pathology.

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

Science.gov (United States)

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

2016-02-01

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

miR-223 is upregulated in monocytes from patients with tuberculosis and regulates function of monocyte-derived macrophages.

Science.gov (United States)

Liu, Yanhua; Wang, Ruo; Jiang, Jing; Yang, Bingfen; Cao, Zhihong; Cheng, Xiaoxing

2015-10-01

Tuberculosis (TB) is a serious infectious disease that most commonly affects the lungs. Macrophages are among the first line defenders against establishment of Mycobacterium tuberculosis infection in the lungs. In this study, we found that activation and cytokine production in monocyte-derived macrophages (MDM) from patients with active TB was impaired. miR-223 expression was significantly elevated in monocytes and MDM from patients with TB compared with healthy controls. To determine the functional role of miR-223 in macrophages, stable miR-223-expressing and miR-223 antisense-expressing U937 cells were established. Compared with empty vector controls, expression of IL-1β, IL-6, TNF-α and IL-12p40 genes was significantly higher in miR-223 antisense-expressing U937 cells, but lower in miR-223-expressing U937 cells. miR-223 can negatively regulate activation of NF-κB by inhibition of p65 phosphorylation and nuclear translocation. It is concluded that miR-223 can regulate macrophage function by inhibition of cytokine production and NF-κB activation. Copyright © 2015 Elsevier Ltd. All rights reserved.

TPL-2 Regulates Macrophage Lipid Metabolism and M2 Differentiation to Control TH2-Mediated Immunopathology

Science.gov (United States)

Entwistle, Lewis J.; Khoury, Hania; Papoutsopoulou, Stamatia; Mahmood, Radma; Mansour, Nuha R.; Ching-Cheng Huang, Stanley; Pearce, Edward J.; Pedro S. de Carvalho, Luiz; Ley, Steven C.

2016-01-01

Persistent TH2 cytokine responses following chronic helminth infections can often lead to the development of tissue pathology and fibrotic scarring. Despite a good understanding of the cellular mechanisms involved in fibrogenesis, there are very few therapeutic options available, highlighting a significant medical need and gap in our understanding of the molecular mechanisms of TH2-mediated immunopathology. In this study, we found that the Map3 kinase, TPL-2 (Map3k8; Cot) regulated TH2-mediated intestinal, hepatic and pulmonary immunopathology following Schistosoma mansoni infection or S. mansoni egg injection. Elevated inflammation, TH2 cell responses and exacerbated fibrosis in Map3k8 –/–mice was observed in mice with myeloid cell-specific (LysM) deletion of Map3k8, but not CD4 cell-specific deletion of Map3k8, indicating that TPL-2 regulated myeloid cell function to limit TH2-mediated immunopathology. Transcriptional and metabolic assays of Map3k8 –/–M2 macrophages identified that TPL-2 was required for lipolysis, M2 macrophage activation and the expression of a variety of genes involved in immuno-regulatory and pro-fibrotic pathways. Taken together this study identified that TPL-2 regulated TH2-mediated inflammation by supporting lipolysis and M2 macrophage activation, preventing TH2 cell expansion and downstream immunopathology and fibrosis. PMID:27487182

Therapeutic potential of carbohydrates as regulators of macrophage activation.

Science.gov (United States)

Lundahl, Mimmi L E; Scanlan, Eoin M; Lavelle, Ed C

2017-12-15

It is well established for a broad range of disease states, including cancer and Mycobacterium tuberculosis infection, that pathogenesis is bolstered by polarisation of macrophages towards an anti-inflammatory phenotype, known as M2. As these innate immune cells are relatively long-lived, their re-polarisation to pro-inflammatory, phagocytic and bactericidal "classically activated" M1 macrophages is an attractive therapeutic approach. On the other hand, there are scenarios where the resolving inflammation, wound healing and tissue remodelling properties of M2 macrophages are beneficial - for example the successful introduction of biomedical implants. Although there are numerous endogenous and exogenous factors that have an impact on the macrophage polarisation spectrum, this review will focus specifically on prominent macrophage-modulating carbohydrate motifs with a view towards highlighting structure-function relationships and therapeutic potential. Copyright © 2017 Elsevier Inc. All rights reserved.

Nuclear Phosphatidylinositol-Phosphate Type I Kinase α-Coupled Star-PAP Polyadenylation Regulates Cell Invasion.

Science.gov (United States)

A P, Sudheesh; Laishram, Rakesh S

2018-03-01

Star-PAP, a nuclear phosphatidylinositol (PI) signal-regulated poly(A) polymerase (PAP), couples with type I PI phosphate kinase α (PIPKIα) and controls gene expression. We show that Star-PAP and PIPKIα together regulate 3'-end processing and expression of pre-mRNAs encoding key anti-invasive factors ( KISS1R , CDH1 , NME1 , CDH13 , FEZ1 , and WIF1 ) in breast cancer. Consistently, the endogenous Star-PAP level is negatively correlated with the cellular invasiveness of breast cancer cells. While silencing Star-PAP or PIPKIα increases cellular invasiveness in low-invasiveness MCF7 cells, Star-PAP overexpression decreases invasiveness in highly invasive MDA-MB-231 cells in a cellular Star-PAP level-dependent manner. However, expression of the PIPKIα-noninteracting Star-PAP mutant or the phosphodeficient Star-PAP (S6A mutant) has no effect on cellular invasiveness. These results strongly indicate that PIPKIα interaction and Star-PAP S6 phosphorylation are required for Star-PAP-mediated regulation of cancer cell invasion and give specificity to target anti-invasive gene expression. Our study establishes Star-PAP-PIPKIα-mediated 3'-end processing as a key anti-invasive mechanism in breast cancer. Copyright © 2018 A.P. and Laishram.

Cell Elasticity Determines Macrophage Function

Science.gov (United States)

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

2012-01-01

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

Cell elasticity determines macrophage function.

Directory of Open Access Journals (Sweden)

Naimish R Patel

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

Anti-Inflammatory Effects of Benfotiamine are Mediated Through the Regulation of Arachidonic Acid Pathway in Macrophages

Science.gov (United States)

Shoeb, Mohammad; Ramana, Kota V

2011-01-01

Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent anti-oxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study indicates a novel role of benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating the arachidonic acid (AA) pathway generated inflammatory lipid mediators in RAW 264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes (LTB4), prostaglandin E2 (PGE2), thromboxanes 2 (TXB2) and prostacyclin (PGI2) in macrophages. Further, LPS-induced expressions of AA metabolizing enzymes such as COX-2, LOX-5, TXB synthase and PGI2 synthase were significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-kB, and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adducts formation. Most importantly, as compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented the LPS-induced macrophage death and monocytes adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response. PMID:22067901

Regulation of alternative macrophage activation in the liver following acetaminophen intoxication by stem cell-derived tyrosine kinase

Energy Technology Data Exchange (ETDEWEB)

Gardner, Carol R., E-mail: cgardner@pharmacy.rutgers.edu [Department of Pharmacology and Toxicology, Rutgers University, Ernest Mario School of Pharmacy, Piscataway, NJ 08854 (United States); Hankey, Pamela [Department of Veterinary and Biomedical Science, Pennsylvania State University, University Park, PA 16802 (United States); Mishin, Vladimir; Francis, Mary [Department of Pharmacology and Toxicology, Rutgers University, Ernest Mario School of Pharmacy, Piscataway, NJ 08854 (United States); Yu, Shan [Department of Veterinary and Biomedical Science, Pennsylvania State University, University Park, PA 16802 (United States); Laskin, Jeffrey D. [Department of Environmental and Occupational Medicine, UMDNJ-Robert Wood Johnson Medical School, Piscataway, NJ 08854 (United States); Laskin, Debra L. [Department of Pharmacology and Toxicology, Rutgers University, Ernest Mario School of Pharmacy, Piscataway, NJ 08854 (United States)

2012-07-15

treated with acetaminophen. These data demonstrate that STK plays a role in regulating macrophage recruitment and activation in the liver following acetaminophen administration, and in hepatotoxicity. -- Highlights: ► STK regulates alternative macrophage activation after acetaminophen intoxication. ► Loss of STK results in increased sensitivity to acetaminophen. ► Increased toxicity involves oxidative stress and decreases in repair macrophages.

Gene expression in IFN-g-activated murine macrophages

Directory of Open Access Journals (Sweden)

Pereira C.A.

2004-01-01

Full Text Available Macrophages are critical for natural immunity and play a central role in specific acquired immunity. The IFN-gamma activation of macrophages derived from A/J or BALB/c mice yielded two different patterns of antiviral state in murine hepatitis virus 3 infection, which were related to a down-regulation of the main virus receptor. Using cDNA hybridization to evaluate mRNA accumulation in the cells, we were able to identify several genes that are differently up- or down-regulated by IFN-gamma in A/J (267 and 266 genes, respectively, up- and down-regulated or BALB/c (297 and 58 genes, respectively, up- and down-regulated mouse macrophages. Macrophages from mice with different genetic backgrounds behave differently at the molecular level and comparison of the patterns of non-activated and IFN-gamma-activated A/J or BALB/c mouse macrophages revealed, for instance, an up-regulation and a down-regulation of genes coding for biological functions such as enzymatic reactions, nucleic acid synthesis and transport, protein synthesis, transport and metabolism, cytoskeleton arrangement and extracellular matrix, phagocytosis, resistance and susceptibility to infection and tumors, inflammation, and cell differentiation or activation. The present data are reported in order to facilitate future correlation of proteomic/transcriptomic findings as well as of results obtained from a classical approach for the understanding of biological phenomena. The possible implication of the role of some of the gene products relevant to macrophage biology can now be further scrutinized. In this respect, a down-regulation of the main murine hepatitis virus 3 receptor gene was detected only in IFN-gamma-activated macrophages of resistant mice.

Expression analysis of G Protein-Coupled Receptors in mouse macrophages.

Science.gov (United States)

Lattin, Jane E; Schroder, Kate; Su, Andrew I; Walker, John R; Zhang, Jie; Wiltshire, Tim; Saijo, Kaoru; Glass, Christopher K; Hume, David A; Kellie, Stuart; Sweet, Matthew J

2008-04-29

Monocytes and macrophages express an extensive repertoire of G Protein-Coupled Receptors (GPCRs) that regulate inflammation and immunity. In this study we performed a systematic micro-array analysis of GPCR expression in primary mouse macrophages to identify family members that are either enriched in macrophages compared to a panel of other cell types, or are regulated by an inflammatory stimulus, the bacterial product lipopolysaccharide (LPS). Several members of the P2RY family had striking expression patterns in macrophages; P2ry6 mRNA was essentially expressed in a macrophage-specific fashion, whilst P2ry1 and P2ry5 mRNA levels were strongly down-regulated by LPS. Expression of several other GPCRs was either restricted to macrophages (e.g. Gpr84) or to both macrophages and neural tissues (e.g. P2ry12, Gpr85). The GPCR repertoire expressed by bone marrow-derived macrophages and thioglycollate-elicited peritoneal macrophages had some commonality, but there were also several GPCRs preferentially expressed by either cell population. The constitutive or regulated expression in macrophages of several GPCRs identified in this study has not previously been described. Future studies on such GPCRs and their agonists are likely to provide important insights into macrophage biology, as well as novel inflammatory pathways that could be future targets for drug discovery.

Up-regulation of TLR2 and TLR4 in high mobility group Box1-stimulated macrophages in pulpitis patients

Science.gov (United States)

Mahmoudi, Javad; Sabermarouf, Babak; Baradaran, Behzad; Sadat-Hatamnezhad, Leila; Shotorbani, Siamak Sandoghchian

2017-01-01

Objective(s): High Mobility Group Box1 (HMGB1) is a nonhistone, DNA-binding protein that serves a crucial role in regulating gene transcription and is involved in a variety of proinflammatory, extracellular activities. The aim of this study was to explore whether HMGB1 stimulation can up-regulate the expression of Toll-like Receptor 2 (TLR2) and Toll-like Receptor 4 (TLR4) on macrophages from pulpitis and to clarify the subsequent events involving Th17 cells and Th17 cell-associated cytokine changes. Materials and Methods: Having prepared dental pulp tissues of pulpitis and healthy controls, macrophage were isolated and cultured. Macrophages were thereafter stimulated by HMGB1 time course. RT-QPCR, flowcytometer, immunofluorescence, Western blotting, and ELISA techniques were used in the present research. Results: Our results showed that the expression of TLR2 and TLR4 on macrophages stimulated with HMGB1 increased in pulpitis compared with controls (macrophages without HMGB1 stimulation) with a statistical significance (Ppulpitis increased, and NF-kB, the downstream target of TLR2 and TLR4, also showed a marked elevation after macrophages’ stimulation by HMGB1. Conclusion: The evidence from the present study suggests that the enhanced TLR2 and TLR4 pathways and Th17 cell polarization may be due to HMGB1 stimulation in pulpitis. PMID:28293399

SHIP-1 Increases Early Oxidative Burst and Regulates Phagosome Maturation in Macrophages1

Science.gov (United States)

Kamen, Lynn A.; Levinsohn, Jonathan; Cadwallader, Amy; Tridandapani, Susheela; Swanson, Joel A.

2010-01-01

Although the inositol phosphatase SHIP-1 is generally thought to inhibit signaling for Fc receptor-mediated phagocytosis, the product of its activity, phosphatidylinositol 3,4 bisphosphate (PI(3,4)P2) has been implicated in activation of the NADPH oxidase. This suggests that SHIP-1 positively regulates generation of reactive oxygen species after phagocytosis. To examine how SHIP-1 activity contributes to Fc receptor-mediated phagocytosis, we measured and compared phospholipid dynamics, membrane trafficking and the oxidative burst in macrophages from SHIP-1-deficient and wild-type mice. SHIP-1-deficient macrophages showed significantly elevated ratios of PI(3,4,5) P3 to PI(3,4)P2 on phagosomal membranes. Imaging reactive oxygen intermediate activities in phagosomes revealed decreased early NADPH oxidase activity in SHIP-1-deficient macrophages. SHIP-1-deficiency also altered later stages of phagosome maturation, as indicated by the persistent elevation of PI(3)P and the early localization of Rab5a to phagosomes. These direct measurements of individual organelles indicate that phagosomal SHIP-1 enhances the early oxidative burst through localized alteration of the membrane 3′ phosphoinositide composition. PMID:18490750

CSF-1R as an inhibitor of apoptosis and promoter of proliferation, migration and invasion of canine mammary cancer cells

Science.gov (United States)

2013-01-01

Background Tumor-associated macrophages (TAMs) have high impact on the cancer development because they can facilitate matrix invasion, angiogenesis, and tumor cell motility. It gives cancer cells the capacity to invade normal tissues and metastasize. The signaling of colony-stimulating factor-1 receptor (CSF-1R) which is an important regulator of proliferation and differentiation of monocytes and macrophages regulates most of the tissue macrophages. However, CSF-1R is expressed also in breast epithelial tissue during some physiological stages i.g.: pregnancy and lactation. Its expression has been also detected in various cancers. Our previous study has showed the expression of CSF-1R in all examined canine mammary tumors. Moreover, it strongly correlated with grade of malignancy and ability to metastasis. This study was therefore designed to characterize the role of CSF-1R in canine mammary cancer cells proliferation, apoptosis, migration, and invasion. As far as we know, the study presented hereby is a pioneering experiment in this field of veterinary medicine. Results We showed that csf-1r silencing significantly increased apoptosis (Annexin V test), decreased proliferation (measured as Ki67 expression) and decreased migration (“wound healing” assay) of canine mammary cancer cells. Treatment of these cells with CSF-1 caused opposite effect. Moreover, csf-1r knock-down changed growth characteristics of highly invasive cell lines on Matrigel matrix, and significantly decreased the ability of these cells to invade matrix. CSF-1 treatment increased invasion of cancer cells. Conclusion The evidence of the expression and functional role of the CSF-1R in canine mammary cancer cells indicate that CSF-1R targeting may be a good therapeutic approach. PMID:23561040

Epigenetic Regulation of Inflammatory Gene Expression in Macrophages by Selenium

Science.gov (United States)

Narayan, Vivek; Ravindra, Kodihalli C.; Liao, Chang; Kaushal, Naveen; Carlson, Bradley A.; Prabhu, K. Sandeep

2014-01-01

Acetylation of histone and non-histone proteins by histone acetyltransferases plays a pivotal role in the expression of pro-inflammatory genes. Given the importance of dietary selenium in mitigating inflammation, we hypothesized that selenium supplementation may regulate inflammatory gene expression at the epigenetic level. The effect of selenium towards histone acetylation was examined in both in vitro and in vivo models of inflammation by chromatin immunoprecipitation (ChIP) assays and immunoblotting. Our results indicated that selenium supplementation, as selenite, decreased acetylation of histone H4 at K12 and K16 in COX-2 and TNF promoters, and of the p65 subunit of the redox sensitive transcription factor NFκB in primary and immortalized macrophages. On the other hand, selenomethionine had a much weaker effect. Selenite treatment of HIV-1 infected human monocytes also significantly decreased the acetylation of H4 at K12 and K16 on the HIV-1 promoter, supporting the downregulation of proviral expression by selenium. A similar decrease in histone acetylation was also seen in the colonic extracts of mice treated with dextran sodium sulfate that correlated well with the levels of selenium in the diet. Bone marrow-derived macrophages from Trspfl/flCreLysM mice that lack expression of selenoproteins in macrophages confirmed the important role of selenoproteins in the inhibition of histone H4 acetylation. Our studies suggest that the ability of selenoproteins to skew the metabolism of arachidonic acid to contribute, in part, to their ability to inhibit histone acetylation. In summary, our studies suggest a new role for selenoproteins in the epigenetic modulation of pro-inflammatory genes. PMID:25458528

Identification of NDRG1-regulated genes associated with invasive potential in cervical and ovarian cancer cells

International Nuclear Information System (INIS)

Zhao, Gang; Chen, Jiawei; Deng, Yanqiu; Gao, Feng; Zhu, Jiwei; Feng, Zhenzhong; Lv, Xiuhong; Zhao, Zheng

2011-01-01

Highlights: → NDRG1 was knockdown in cervical and ovarian cancer cell lines by shRNA technology. → NDRG1 knockdown resulted in increased cell invasion activities. → Ninety-six common deregulated genes in both cell lines were identified by cDNA microarray. → Eleven common NDRG1-regulated genes might enhance cell invasive activity. → Regulation of invasion by NDRG1 is an indirect and complicated process. -- Abstract: N-myc downstream regulated gene 1 (NDRG1) is an important gene regulating tumor invasion. In this study, shRNA technology was used to suppress NDRG1 expression in CaSki (a cervical cancer cell line) and HO-8910PM (an ovarian cancer cell line). In vitro assays showed that NDRG1 knockdown enhanced tumor cell adhesion, migration and invasion activities without affecting cell proliferation. cDNA microarray analysis revealed 96 deregulated genes with more than 2-fold changes in both cell lines after NDRG1 knockdown. Ten common upregulated genes (LPXN, DDR2, COL6A1, IL6, IL8, FYN, PTP4A3, PAPPA, ETV5 and CYGB) and one common downregulated gene (CLCA2) were considered to enhance tumor cell invasive activity. BisoGenet network analysis indicated that NDRG1 regulated these invasion effector genes/proteins in an indirect manner. Moreover, NDRG1 knockdown also reduced pro-invasion genes expression such as MMP7, TMPRSS4 and CTSK. These results suggest that regulation of invasion and metastasis by NDRG1 is a highly complicated process.

Biology of Bony Fish Macrophages

OpenAIRE

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

2015-01-01

Macrophages are found across all vertebrate species, reside in virtually all animal tissues, and play critical roles in host protection and homeostasis. Various mechanisms determine and regulate the highly plastic functional phenotypes of macrophages, including antimicrobial host defenses (pro-inflammatory, M1-type), and resolution and repair functions (anti-inflammatory/regulatory, M2-type). The study of inflammatory macrophages in immune defense of teleosts has garnered much attention, and ...

Wip1-dependent modulation of macrophage migration and phagocytosis

DEFF Research Database (Denmark)

Tang, Yiting; Pan, Bing; Zhou, Xin

2017-01-01

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

Macrophage polarization in nerve injury: do Schwann cells play a role?

Directory of Open Access Journals (Sweden)

Jo Anne Stratton

2016-01-01

Full Text Available In response to peripheral nerve injury, the inflammatory response is almost entirely comprised of infiltrating macrophages. Macrophages are a highly plastic, heterogenic immune cell, playing an indispensable role in peripheral nerve injury, clearing debris and regulating the microenvironment to allow for efficient regeneration. There are several cells within the microenvironment that likely interact with macrophages to support their function - most notably the Schwann cell, the glial cell of the peripheral nervous system. Schwann cells express several ligands that are known to interact with receptors expressed by macrophages, yet the effects of Schwann cells in regulating macrophage phenotype remains largely unexplored. This review discusses macrophages in peripheral nerve injury and how Schwann cells may regulate their behavior.

Divergence of macrophage phagocytic and antimicrobial programs in leprosy.

Science.gov (United States)

Montoya, Dennis; Cruz, Daniel; Teles, Rosane M B; Lee, Delphine J; Ochoa, Maria Teresa; Krutzik, Stephan R; Chun, Rene; Schenk, Mirjam; Zhang, Xiaoran; Ferguson, Benjamin G; Burdick, Anne E; Sarno, Euzenir N; Rea, Thomas H; Hewison, Martin; Adams, John S; Cheng, Genhong; Modlin, Robert L

2009-10-22

Effective innate immunity against many microbial pathogens requires macrophage programs that upregulate phagocytosis and direct antimicrobial pathways, two functions generally assumed to be coordinately regulated. We investigated the regulation of these key functions in human blood-derived macrophages. Interleukin-10 (IL-10) induced the phagocytic pathway, including the C-type lectin CD209 and scavenger receptors, resulting in phagocytosis of mycobacteria and oxidized low-density lipoprotein. IL-15 induced the vitamin D-dependent antimicrobial pathway and CD209, yet the cells were less phagocytic. The differential regulation of macrophage functional programs was confirmed by analysis of leprosy lesions: the macrophage phagocytosis pathway was prominent in the clinically progressive, multibacillary form of the disease, whereas the vitamin D-dependent antimicrobial pathway predominated in the self-limited form and in patients undergoing reversal reactions from the multibacillary to the self-limited form. These data indicate that macrophage programs for phagocytosis and antimicrobial responses are distinct and differentially regulated in innate immunity to bacterial infections.

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

Science.gov (United States)

Harwani, Sailesh C

2018-01-01

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

NDRG2 inhibits hepatocellular carcinoma adhesion, migration and invasion by regulating CD24 expression

International Nuclear Information System (INIS)

Zheng, Jin; Guo, Hang; Tao, Yurong; Xue, Yan; Jiang, Ning; Yao, Libo; Liu, Wenchao; Li, Yan; Yang, Jiandong; Liu, Qiang; Shi, Ming; Zhang, Rui; Shi, Hengjun; Ren, Qinyou; Ma, Ji

2011-01-01

The prognosis of most hepatocellular carcinoma (HCC) patients is poor due to the high metastatic rate of the disease. Understanding the molecular mechanisms underlying HCC metastasis is extremely urgent. The role of CD24 and NDRG2 (N-myc downstream-regulated gene 2), a candidate tumor suppressor gene, has not yet been explored in HCC. The mRNA and protein expression of CD24 and NDRG2 was analyzed in MHCC97H, Huh7 and L-02 cells. Changes in cell adhesion, migration and invasion were detected by up- or down-regulating NDRG2 by adenovirus or siRNA. The expression pattern of NDRG2 and CD24 in HCC tissues and the relationship between NDRG2 and HCC clinical features was analyzed by immunohistochemical and western blotting analysis. NDRG2 expression was negatively correlated with malignancy in HCC. NDRG2 exerted anti-tumor activity by regulating CD24, a molecule that mediates cell-cell interaction, tumor proliferation and adhesion. NDRG2 up-regulation decreased CD24 expression and cell adhesion, migration and invasion. By contrast, NDRG2 down-regulation enhanced CD24 expression and cell adhesion, migration and invasion. Immunohistochemical analysis of 50 human HCC clinical specimens showed a strong correlation between NDRG2 down-regulation and CD24 overexpression (P = 0.04). In addition, increased frequency of NDRG2 down-regulation was observed in patients with elevated AFP serum level (P = 0.006), late TNM stage (P = 0.009), poor differentiation grade (P = 0.002), tumor invasion (P = 0.004) and recurrence (P = 0.024). Our findings indicate that NDRG2 and CD24 regulate HCC adhesion, migration and invasion. The expression level of NDRG2 is closely related to the clinical features of HCC. Thus, NDRG2 plays an important physiological role in HCC metastasis

p62 regulates CD40-mediated NFκB activation in macrophages through interaction with TRAF6

Energy Technology Data Exchange (ETDEWEB)

Seibold, Kristina; Ehrenschwender, Martin, E-mail: martin.ehrenschwender@ukr.de

2015-08-14

CD40 is a member of the tumor necrosis factor (TNF) receptor family. Activation-induced recruitment of adapter proteins, so-called TNF-receptor-associated factors (TRAFs) to the cytoplasmic tail of CD40 triggers signaling cascades important in the immune system, but has also been associated with excessive inflammation in diseases such as atherosclerosis and rheumatoid arthritis. Especially, pro-inflammatory nuclear factor κB (NFκB) signaling emanating from CD40-associated TRAF6 appears to be a key pathogenic driving force. Consequently, targeting the CD40-TRAF6 interaction is emerging as a promising therapeutic strategy, but the underlying molecular machinery of this signaling axis is to date poorly understood. Here, we identified the multifunctional adaptor protein p62 as a critical regulator in CD40-mediated NFκB signaling via TRAF6. CD40 activation triggered formation of a TRAF6-p62 complex. Disturbing this interaction tremendously reduced CD40-mediated NFκB signaling in macrophages, while TRAF6-independent signaling pathways remained unaffected. This highlights p62 as a potential target in hyper-inflammatory, CD40-associated pathologies. - Highlights: • CD40 activation triggers interaction of the adapter protein TRAF6 with p62. • TRAF6-p62 interaction regulates CD40-mediated NFκB signaling in macrophages. • Defective TRAF6-p62 interaction reduces CD40-mediated NFκB activation in macrophages.

Anti-inflammatory effects of benfotiamine are mediated through the regulation of the arachidonic acid pathway in macrophages.

Science.gov (United States)

Shoeb, Mohammad; Ramana, Kota V

2012-01-01

Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent antioxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study (U.C. Yadav et al., Free Radic. Biol. Med. 48:1423-1434, 2010) indicates a novel role for benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating arachidonic acid (AA) pathway-generated inflammatory lipid mediators in RAW264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes, prostaglandin E2, thromboxane 2 (TXB2), and prostacyclin (PGI2) in macrophages. Further, LPS-induced expression of AA-metabolizing enzymes such as COX-2, LOX-5, TXB synthase, and PGI2 synthase was significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-κB and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adduct formation. Most importantly, compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented LPS-induced macrophage death and monocyte adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of the COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response. Copyright © 2011 Elsevier Inc. All rights reserved.

The cellular prion protein negatively regulates phagocytosis and cytokine expression in murine bone marrow-derived macrophages.

Directory of Open Access Journals (Sweden)

Min Wang

Full Text Available The cellular prion protein (PrP(C is a glycosylphosphatidylinositol (GPI-anchored glycoprotein on the cell surface. Previous studies have demonstrated contradictory roles for PrP(C in connection with the phagocytic ability of macrophages. In the present work, we investigated the function of PrP(C in phagocytosis and cytokine expression in bone marrow-derived macrophages infected with Escherichia coli. E. coli infection induced an increase in the PRNP mRNA level. Knockout of PrP(C promoted bacterial uptake; upregulated Rab5, Rab7, and Eea1 mRNA expression; and increased the recruitment of lysosomal-associated membrane protein-2 to phagosomes, suggesting enhanced microbicidal activity. Remarkably, knockout of PrP(C suppressed the proliferation of internalized bacteria and increased the expression of cytokines such as interleukin-1β. Collectively, our data reveal an important role of PrP(C as a negative regulator for phagocytosis, phagosome maturation, cytokine expression, and macrophage microbicidal activity.

miR-148a-3p Mediates Notch Signaling to Promote the Differentiation and M1 Activation of Macrophages

Directory of Open Access Journals (Sweden)

Fei Huang

2017-10-01

Full Text Available The Notch pathway plays critical roles in the differentiation and polarized activation of macrophages; however, the downstream molecular mechanisms underlying Notch activity in macrophages remain elusive. Our previous study has identified a group of microRNAs that mediate Notch signaling to regulate macrophage activation and tumor-associated macrophages (TAMs. In this study, we demonstrated that miR-148a-3p functions as a novel downstream molecule of Notch signaling to promote the differentiation of monocytes into macrophages in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF. Meanwhile, miR-148a-3p promoted M1 and inhibited M2 polarization of macrophages upon Notch activation. Macrophages overexpressing miR-148a-3p exhibited enhanced ability to engulf and kill bacteria, which was mediated by excessive production of reactive oxygen species (ROS. Further studies using reporter assay and Western blotting identified Pten as a direct target gene of miR-148a-3p in macrophages. Macrophages overexpressing miR-148a-3p increased their ROS production through the PTEN/AKT pathway, likely to defend against bacterial invasion. Moreover, miR-148a-3p also enhanced M1 macrophage polarization and pro-inflammatory responses through PTEN/AKT-mediated upregulation of NF-κB signaling. In summary, our data establish a novel molecular mechanism by which Notch signaling promotes monocyte differentiation and M1 macrophage activation through miR-148a-3p, and suggest that miR-148a-3p-modified monocytes or macrophages are potential new tools for the treatment of inflammation-related diseases.

Adipose Type One Innate Lymphoid Cells Regulate Macrophage Homeostasis through Targeted Cytotoxicity.

Science.gov (United States)

Boulenouar, Selma; Michelet, Xavier; Duquette, Danielle; Alvarez, David; Hogan, Andrew E; Dold, Christina; O'Connor, Donal; Stutte, Suzanne; Tavakkoli, Ali; Winters, Desmond; Exley, Mark A; O'Shea, Donal; Brenner, Michael B; von Andrian, Ulrich; Lynch, Lydia

2017-02-21

Adipose tissue has a dynamic immune system that adapts to changes in diet and maintains homeostatic tissue remodeling. Adipose type 1 innate lymphoid cells (AT1-ILCs) promote pro-inflammatory macrophages in obesity, but little is known about their functions at steady state. Here we found that human and murine adipose tissue harbor heterogeneous populations of AT1-ILCs. Experiments using parabiotic mice fed a high-fat diet (HFD) showed differential trafficking of AT1-ILCs, particularly in response to short- and long-term HFD and diet restriction. At steady state, AT1-ILCs displayed cytotoxic activity toward adipose tissue macrophages (ATMs). Depletion of AT1-ILCs and perforin deficiency resulted in alterations in the ratio of inflammatory to anti-inflammatory ATMs, and adoptive transfer of AT1-ILCs exacerbated metabolic disorder. Diet-induced obesity impaired AT1-ILC killing ability. Our findings reveal a role for AT1-ILCs in regulating ATM homeostasis through cytotoxicity and suggest that this function is relevant in both homeostasis and metabolic disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Epigenetic regulation of CpG promoter methylation in invasive prostate cancer cells

Directory of Open Access Journals (Sweden)

Farrar William L

2010-10-01

Full Text Available Abstract Background Recently, much attention has been focused on gaining a better understanding of the different populations of cells within a tumor and their contribution to cancer progression. One of the most commonly used methods to isolate a more aggressive sub-population of cells utilizes cell sorting based on expression of certain cell adhesion molecules. A recently established method we developed is to isolate these more aggressive cells based on their properties of increased invasive ability. These more invasive cells have been previously characterized as tumor initiating cells (TICs that have a stem-like genomic signature and express a number of stem cell genes including Oct3/4 and Nanog and are more tumorigenic compared to their 'non-invasive' counterpart. They also have a profile reminiscent of cells undergoing a classic pattern of epithelial to mesenchymal transition or EMT. Using this model of invasion, we sought to investigate which genes are under epigenetic control in this rare population of cells. Epigenetic modifications, specifically DNA methylation, are key events regulating the process of normal human development. To determine the specific methylation pattern in these invasive prostate cells, and if any developmental genes were being differentially regulated, we analyzed differences in global CpG promoter methylation. Results Differentially methylated genes were determined and select genes were chosen for additional analyses. The non-receptor tyrosine kinase BMX and transcription factor SOX1 were found to play a significant role in invasion. Ingenuity pathway analysis revealed the methylated gene list frequently displayed genes from the IL-6/STAT3 pathway. Cells which have decreased levels of the targets BMX and SOX1 also display loss of STAT3 activity. Finally, using Oncomine, it was determined that more aggressive metastatic prostate cancers in humans also have higher levels of both Stat3 and Sox1. Conclusions Using this

Macrophage immunoregulatory pathways in tuberculosis.

Science.gov (United States)

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

2014-12-01

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

Vinculin contributes to Cell Invasion by Regulating Contractile Activation

Science.gov (United States)

Mierke, Claudia Tanja

2008-07-01

Vinculin is a component of the focal adhesion complex and is described as a mechano-coupling protein connecting the integrin receptor and the actin cytoskeleton. Vinculin knock-out (k.o.) cells (vin-/-) displayed increased migration on a 2-D collagen- or fibronectin-coated substrate compared to wildtype cells, but the role of vinculin in cell migration through a 3-D connective tissue is unknown. We determined the invasiveness of established tumor cell lines using a 3-D collagen invasion assay. Gene expression analysis of 4 invasive and 4 non-invasive tumor cell lines revealed that vinculin expression was significantly increased in invasive tumor cell lines. To analyze the mechanisms by which vinculin increased cell invasion in a 3-D gel, we studied mouse embryonic fibroblasts wildtype and vin-/- cells. Wildtype cells were 3-fold more invasive compared vin-/- cells. We hypothesized that the ability to generate sufficient traction forces is a prerequisite for tumor cell migration in a 3-D connective tissue matrix. Using traction microscopy, we found that wildtype exerted 3-fold higher tractions on fibronectin-coated polyacrylamide gels compared to vin-/- cells. These results show that vinculin controls two fundamental functions that lead to opposite effects on cell migration in a 2-D vs. a 3-D environment: On the one hand, vinculin stabilizes the focal adhesions (mechano-coupling function) and thereby reduces motility in 2-D. On the other hand, vinculin is also a potent activator of traction generation (mechano-regulating function) that is important for cell invasion in a 3-D environment.

Biglycan- and Sphingosine Kinase-1 Signaling Crosstalk Regulates the Synthesis of Macrophage Chemoattractants

Directory of Open Access Journals (Sweden)

Louise Tzung-Harn Hsieh

2017-03-01

Full Text Available In its soluble form, the extracellular matrix proteoglycan biglycan triggers the synthesis of the macrophage chemoattractants, chemokine (C-C motif ligand CCL2 and CCL5 through selective utilization of Toll-like receptors (TLRs and their adaptor molecules. However, the respective downstream signaling events resulting in biglycan-induced CCL2 and CCL5 production have not yet been defined. Here, we show that biglycan stimulates the production and activation of sphingosine kinase 1 (SphK1 in a TLR4- and Toll/interleukin (IL-1R domain-containing adaptor inducing interferon (IFN-β (TRIF-dependent manner in murine primary macrophages. We provide genetic and pharmacological proof that SphK1 is a crucial downstream mediator of biglycan-triggered CCL2 and CCL5 mRNA and protein expression. This is selectively driven by biglycan/SphK1-dependent phosphorylation of the nuclear factor NF-κB p65 subunit, extracellular signal-regulated kinase (Erk1/2 and p38 mitogen-activated protein kinases. Importantly, in vivo overexpression of soluble biglycan causes Sphk1-dependent enhancement of renal CCL2 and CCL5 and macrophage recruitment into the kidney. Our findings describe the crosstalk between biglycan- and SphK1-driven extracellular matrix- and lipid-signaling. Thus, SphK1 may represent a new target for therapeutic intervention in biglycan-evoked inflammatory conditions.

Tumor-associated macrophage-derived IL-6 and IL-8 enhance invasive activity of LoVo cells induced by PRL-3 in a KCNN4 channel-dependent manner

International Nuclear Information System (INIS)

Xu, Heyang; Lai, Wei; Zhang, Yang; Liu, Lu; Luo, Xingxi; Zeng, Yujie; Wu, Heng; Lan, Qiusheng; Chu, Zhonghua

2014-01-01

Tumor-associated macrophages (TAMs) are known to promote cancer progression and metastasis through the release of a variety of cytokines. Phosphatase of regenerating liver (PRL-3) has been considered as a marker of colorectal cancer (CRC) liver metastasis. Our previous research suggests that PRL-3 can enhance the metastasis of CRC through the up-regulation of intermediate-conductance Ca 2+ -activated K + (KCNN4) channel, which is dependent on the autocrine secretion of tumor necrosis factor-alpha (TNF-α). However, whether TAMs participate in the progression and metastasis of CRC induced by PRL-3 remains unknown. We used flow cytometry, coculture, western blotting, invasion assays, real-time quantitative PCR, chromatin immunoprecipitation, luciferase reporter assays, and immunofluorescence staining to determine the effect of TAMs on the ability of PRL-3 to promote invasiveness of CRC cells. In this study, we found that TAMs facilitated the metastasis of CRC induced by PRL-3. When TAMs were cocultured with CRC cells, the expression of KCNN4 was increased in TAMs and the invasion of CRC cells was enhanced. Furthermore, cytokines that were secreted by TAMs, such as IL-6 and IL-8, were also significantly increased. This response was attenuated by treating TAMs with the KCNN4 channel-specific inhibitor, 1-[(2-chlorophenyl) diphenylmethyl]-1H-pyrazole (TRAM-34), which suggested that KCNN4 channels may be involved in inducing the secretion of IL-6 and IL-8 by TAMs and improving CRC cell invasiveness. Moreover, the expression of KCNN4 channels in TAMs was regulated through the NF-κB signal pathway, which is activated by TNF-α from CRC cells. Immunofluorescence analysis of colorectal specimens indicated that IL-6 and IL-8 double positive cells in the stroma showed positive staining for the TAM marker CD68, suggesting that TAMs produce IL-6 and IL-8. Increased numbers of these cells correlated with higher clinical stage. Our findings suggested that TAMs participate in the

Origins and Hallmarks of Macrophages: Development, Homeostasis, and Disease

Science.gov (United States)

Wynn, Thomas A.; Chawla, Ajay; Pollard, Jeffrey W.

2013-01-01

Preface Macrophages the most plastic cells of the hematopoietic system are found in all tissues and exhibit great functional diversity. They have roles in development, homeostasis, tissue repair, and immunity. While anatomically distinct, resident tissue macrophages exhibit different transcriptional profiles, and functional capabilities, they are all required for the maintenance of homeostasis. However, these reparative and homeostatic functions can be subverted by chronic insults, resulting in a causal association of macrophages with disease states. In this review, we discuss how macrophages regulate normal physiology and development and provide several examples of their pathophysiologic roles in disease. We define the “hallmarks” of macrophages performing particular functions, taking into account novel insights into the diversity of their lineages, identity, and regulation. This diversity is essential to understand because macrophages have emerged as important therapeutic targets in many important human diseases. PMID:23619691

IL-15 up-regulates the MMP-9 expression levels and induces inflammatory infiltration of macrophages in polymyositis through regulating the NF-kB pathway.

Science.gov (United States)

Yan, Wang; Fan, Weinv; Chen, Caijing; Wu, Yunqin; Fan, Zhenyi; Chen, Jiaqi; Chen, Zhaoying; Chen, Huimin

2016-10-10

This study was aimed to research the effects of IL-15 on inducing inflammatory infiltration of macrophages in polymyositis (PM) through the NF-kB pathway, and whether IL-15 was able to further regulate MMP-9 expression levels. Prepared PM cells, collected from the patients suffering from PM, were administered to SD rats. Also, a group of healthy SD rats was undergoing the same treatment as the control group. The test animals were treated with either anti-IL-15, IL-15, MMP-9 siRNA or ERK1/2 inhibitor. The blood toxicological parameters creatine kinase (CK) and CD163 were tested by using ELISA and immunohistochemistry assay. In addition, NF-kB expression in macrophages was measured by immunocytochemical assay. To measure the degree of cell infiltration the Transwell assay was performed. Lastly, western blot and zymography were carried out to compare MMP-9 and ERK expression levels between the two groups, both in vivo and in vitro. The results showed that S-CK, IL-15 and IL-15Rα levels increased rapidly after the conventional treatment was introduced to the PM infected SD rats. The PM model establishment and IL-15 treatment significantly increased the expressions of IL-15Rα, MMP-9, p-ERK and p-IKBα. However, the same effect can be suppressed by using anti-IL-15, MMP-9 siRNA or ERK1/2 inhibitor (P kB in the macrophages. IL-15 is able to significantly regulate the inflammatory infiltration of macrophages in PM patients through affecting the NF-kB pathway and MMP-9 expression levels. Copyright © 2016 Elsevier B.V. All rights reserved.

Cytosolic Pellino-1-Mediated K63-Linked Ubiquitination of IRF5 in M1 Macrophages Regulates Glucose Intolerance in Obesity

Directory of Open Access Journals (Sweden)

Donghyun Kim

2017-07-01

Full Text Available IRF5 is a signature transcription factor that induces M1 macrophage polarization. However, little is known regarding cytosolic proteins that induce IRF5 activation for M1 polarization. Here, we report the interaction between ubiquitin E3 ligase Pellino-1 and IRF5 in the cytoplasm, which increased nuclear translocation of IRF5 by K63-linked ubiquitination in human and mouse M1 macrophages. LPS and/or IFN-γ increased Pellino-1 expression, and M1 polarization was attenuated in Pellino-1-deficient macrophages in vitro and in vivo. Defective M1 polarization in Pellino-1-deficient macrophages improved glucose intolerance in mice fed a high-fat diet. Furthermore, macrophages in adipose tissues from obese humans exhibited increased Pellino-1 expression and IRF5 nuclear translocation compared with nonobese subjects, and these changes are associated with insulin resistance index. This study demonstrates that cytosolic Pellino-1-mediated K63-linked ubiquitination of IRF5 in M1 macrophages regulates glucose intolerance in obesity, suggesting a cytosolic mediator function of Pellino-1 in TLR4/IFN-γ receptor-IRF5 axis during M1 polarization.

Extratumoral Heme Oxygenase-1 (HO-1 Expressing Macrophages Likely Promote Primary and Metastatic Prostate Tumor Growth.

Directory of Open Access Journals (Sweden)

Sofia Halin Bergström

Full Text Available Aggressive tumors induce tumor-supporting changes in the benign parts of the prostate. One factor that has increased expression outside prostate tumors is hemoxygenase-1 (HO-1. To investigate HO-1 expression in more detail, we analyzed samples of tumor tissue and peritumoral normal prostate tissue from rats carrying cancers with different metastatic capacity, and human prostate cancer tissue samples from primary tumors and bone metastases. In rat prostate tumor samples, immunohistochemistry and quantitative RT-PCR showed that the main site of HO-1 synthesis was HO-1+ macrophages that accumulated in the tumor-bearing organ, and at the tumor-invasive front. Small metastatic tumors were considerably more effective in attracting HO-1+ macrophages than larger non-metastatic ones. In clinical samples, accumulation of HO-1+ macrophages was seen at the tumor invasive front, almost exclusively in high-grade tumors, and it correlated with the presence of bone metastases. HO-1+ macrophages, located at the tumor invasive front, were more abundant in bone metastases than in primary tumors. HO-1 expression in bone metastases was variable, and positively correlated with the expression of macrophage markers but negatively correlated with androgen receptor expression, suggesting that elevated HO-1 could be a marker for a subgroup of bone metastases. Together with another recent observation showing that selective knockout of HO-1 in macrophages reduced prostate tumor growth and metastatic capacity in animals, the results of this study suggest that extratumoral HO-1+ macrophages may have an important role in prostate cancer.

Nitric oxide–mediated regulation of ferroportin-1 controls macrophage iron homeostasis and immune function in Salmonella infection

Science.gov (United States)

Nairz, Manfred; Schleicher, Ulrike; Schroll, Andrea; Sonnweber, Thomas; Theurl, Igor; Ludwiczek, Susanne; Talasz, Heribert; Brandacher, Gerald; Moser, Patrizia L.; Muckenthaler, Martina U.; Fang, Ferric C.; Bogdan, Christian

2013-01-01

Nitric oxide (NO) generated by inducible NO synthase 2 (NOS2) affects cellular iron homeostasis, but the underlying molecular mechanisms and implications for NOS2-dependent pathogen control are incompletely understood. In this study, we found that NO up-regulated the expression of ferroportin-1 (Fpn1), the major cellular iron exporter, in mouse and human cells. Nos2−/− macrophages displayed increased iron content due to reduced Fpn1 expression and allowed for an enhanced iron acquisition by the intracellular bacterium Salmonella typhimurium. Nos2 gene disruption or inhibition of NOS2 activity led to an accumulation of iron in the spleen and splenic macrophages. Lack of NO formation resulted in impaired nuclear factor erythroid 2-related factor-2 (Nrf2) expression, resulting in reduced Fpn1 transcription and diminished cellular iron egress. After infection of Nos2−/− macrophages or mice with S. typhimurium, the increased iron accumulation was paralleled by a reduced cytokine (TNF, IL-12, and IFN-γ) expression and impaired pathogen control, all of which were restored upon administration of the iron chelator deferasirox or hyperexpression of Fpn1 or Nrf2. Thus, the accumulation of iron in Nos2−/− macrophages counteracts a proinflammatory host immune response, and the protective effect of NO appears to partially result from its ability to prevent iron overload in macrophages PMID:23630227

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

Science.gov (United States)

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

2010-08-01

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

Fractalkine Signaling Regulates Macrophage Recruitment into the Cochlea and Promotes the Survival of Spiral Ganglion Neurons after Selective Hair Cell Lesion.

Science.gov (United States)

Kaur, Tejbeer; Zamani, Darius; Tong, Ling; Rubel, Edwin W; Ohlemiller, Kevin K; Hirose, Keiko; Warchol, Mark E

2015-11-11

Macrophages are recruited into the cochlea in response to injury caused by acoustic trauma or ototoxicity, but the nature of the interaction between macrophages and the sensory structures of the inner ear remains unclear. The present study examined the role of fractalkine signaling in regulating the injury-evoked behavior of macrophages following the selective ablation of cochlear hair cells. We used a novel transgenic mouse model in which the human diphtheria toxin receptor (huDTR) is selectively expressed under the control of Pou4f3, a hair cell-specific transcription factor. Administration of diphtheria toxin (DT) to these mice resulted in nearly complete ablation of cochlear hair cells, with no evident pathology among supporting cells, spiral ganglion neurons, or cells of the cochlear lateral wall. Hair cell death led to an increase in macrophages associated with the sensory epithelium of the cochlea. Their numbers peaked at 14 days after DT and then declined at later survival times. Increased macrophages were also observed within the spiral ganglion, but their numbers remained elevated for (at least) 56 d after DT. To investigate the role of fractalkine signaling in macrophage recruitment, we crossed huDTR mice to a mouse line that lacks expression of the fractalkine receptor (CX3CR1). Disruption of fractalkine signaling reduced macrophage recruitment into both the sensory epithelium and spiral ganglion and also resulted in diminished survival of spiral ganglion neurons after hair cell death. Our results suggest a fractalkine-mediated interaction between macrophages and the neurons of the cochlea. It is known that damage to the inner ear leads to recruitment of inflammatory cells (macrophages), but the chemical signals that initiate this recruitment and the functions of macrophages in the damaged ear are unclear. Here we show that fractalkine signaling regulates macrophage recruitment into the cochlea and also promotes the survival of cochlear afferents after

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

Science.gov (United States)

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

2018-02-05

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

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

Directory of Open Access Journals (Sweden)

Toshihiko Suzuki

2007-08-01

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

Lack of RNase L attenuates macrophage functions.

Directory of Open Access Journals (Sweden)

Xin Yi

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

Consequences of gamma-irradiation on inflammatory cytokine regulation in human monocytes/macrophages; Consequences de l`irradiation gamma sur la regulation des cytokines de l`inflammation dans les monocytes/macrophages humains

Energy Technology Data Exchange (ETDEWEB)

Pons, I.; Gras, G.; Dormont, D.

1995-12-31

Inflammation is a frequent radiation-induced damage, especially after therapeutic irradiation. In this study, we have investigated, the inflammatory cytokine regulation after ionizing irradiation of monocytes/macrophages from four donors. Semi-quantitative RT-PCR revealed, after in vitro 24 h-differentiated monocytes irradiation between 5 to 40 Gy, no induction of interleukin-I{beta} (IL I{beta}), interleukin-6 (IL-6) and tumor necrosis factor-{alpha} (TNF-{alpha} mRNA) expression. Moreover, protein quantitation shows no significant increase of post-irradiation secretion. (author). 6 refs.

Insulin regulates multiple signaling pathways leading to monocyte/macrophage chemotaxis into the wound tissue

Directory of Open Access Journals (Sweden)

Yan Liu

2018-01-01

Full Text Available Wound healing is a complex process that involves sequential phases that overlap in time and space and affect each other dynamically at the gene and protein levels. We previously showed that insulin accelerates wound healing by stimulating faster and regenerative healing. One of the processes that insulin stimulates is an increase in monocyte/macrophage chemotaxis. In this study, we performed experiments in vivo and in vitro to elucidate the signaling transduction pathways that are involved in insulin-induced monocyte/macrophage chemotaxis. We found that insulin stimulates THP-1 cell chemotaxis in a dose-dependent and insulin receptor-dependent manner. We also show that the kinases PI3K-Akt, SPAK/JNK, and p38 MAPK are key molecules in the insulin-induced signaling pathways that lead to chemoattraction of the THP-1 cell. Furthermore, both PI3K-Akt and SPAK/JNK signaling involve Rac1 activation, an important molecule in regulating cell motility. Indeed, topical application of Rac1 inhibitor at an early stage during the healing process caused delayed and impaired healing even in the presence of insulin. These results delineate cell and molecular mechanisms involved in insulin-induced chemotaxis of monocyte/macrophage, cells that are critical for proper healing.

Macrophages in synovial inflammation

Directory of Open Access Journals (Sweden)

Aisling eKennedy

2011-10-01

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

ATM regulation of IL-8 links oxidative stress to cancer cell migration and invasion.

Science.gov (United States)

Chen, Wei-Ta; Ebelt, Nancy D; Stracker, Travis H; Xhemalce, Blerta; Van Den Berg, Carla L; Miller, Kyle M

2015-06-01

Ataxia-telangiectasia mutated (ATM) protein kinase regulates the DNA damage response (DDR) and is associated with cancer suppression. Here we report a cancer-promoting role for ATM. ATM depletion in metastatic cancer cells reduced cell migration and invasion. Transcription analyses identified a gene network, including the chemokine IL-8, regulated by ATM. IL-8 expression required ATM and was regulated by oxidative stress. IL-8 was validated as an ATM target by its ability to rescue cell migration and invasion defects in ATM-depleted cells. Finally, ATM-depletion in human breast cancer cells reduced lung tumors in a mouse xenograft model and clinical data validated IL-8 in lung metastasis. These findings provide insights into how ATM activation by oxidative stress regulates IL-8 to sustain cell migration and invasion in cancer cells to promote metastatic potential. Thus, in addition to well-established roles in tumor suppression, these findings identify a role for ATM in tumor progression.

Macrophage Functions in Early Dissemination and Dormancy of Breast Cancer

Science.gov (United States)

2016-09-01

mammary gland development 17,18, 69   arguing that normal mammary epithelial cells cooperate with these innate immune cells 70   for invasive... cells lacking 218     11   lymphoid and granulocytic markers (Supplementary Fig.3B). viSNE plots 30 of myelo-219   monocytic cells (Fig.5A...macrophages are actively recruited by pre-malignant ErbB2 overexpressing cancer cells and that these intra-epithelial macrophages then produce factors

Procyanidin dimer B2-mediated IRAK-M induction negatively regulates TLR4 signaling in macrophages

Energy Technology Data Exchange (ETDEWEB)

Sung, Nak-Yun [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Yang, Mi-So [Department of Microbiology, Infection Signaling Network Research Center, College of Medicine, Chungnam National University, Daejeon (Korea, Republic of); Song, Du-Sub [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); School of life sciences and Biotechnology, Korea University 5-ka, Anam-Dong, Sungbuk-ku, Seoul 136-701 (Korea, Republic of); Kim, Jae-Kyung; Park, Jong-Heum; Song, Beom-Seok; Park, Sang-Hyun; Lee, Ju-Woon [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Park, Hyun-Jin [School of life sciences and Biotechnology, Korea University 5-ka, Anam-Dong, Sungbuk-ku, Seoul 136-701 (Korea, Republic of); Kim, Jae-Hun [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Byun, Eui-Baek, E-mail: ebbyun80@kaeri.re.kr [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Byun, Eui-Hong, E-mail: ehbyun80@kongju.ac.k [Department of Food Science and Technology, Kongju National University, Yesan 340-800 (Korea, Republic of)

2013-08-16

Highlights: •Pro B2 elevated the expression of IRAK-M, a negative regulator of TLR signaling. •LPS-induced expression of cell surface molecules was inhibited by Pro B2. •LPS-induced production of pro-inflammatory cytokines was inhibited by Pro B2. •Pro B2 inhibited LPS-induced activation of MAPKs and NF-κB through IRAK-M. •Pro B2 inactivated naïve T cells by inhibiting LPS-induced cytokines via IRAK-M. -- Abstract: Polyphenolic compounds have been found to possess a wide range of physiological activities that may contribute to their beneficial effects against inflammation-related diseases; however, the molecular mechanisms underlying this anti-inflammatory activity are not completely characterized, and many features remain to be elucidated. In this study, we investigated the molecular basis for the down-regulation of toll-like receptor 4 (TLR4) signal transduction by procyanidin dimer B2 (Pro B2) in macrophages. Pro B2 markedly elevated the expression of the interleukin (IL)-1 receptor-associated kinase (IRAK)-M protein, a negative regulator of TLR signaling. Lipopolysaccharide (LPS)-induced expression of cell surface molecules (CD80, CD86, and MHC class I/II) and production of pro-inflammatory cytokines (tumor necrosis factor-α, IL-1β, IL-6, and IL-12p70) were inhibited by Pro B2, and this action was prevented by IRAK-M silencing. In addition, Pro B2-treated macrophages inhibited LPS-induced activation of mitogen-activated protein kinases such as extracellular signal-regulated kinase 1/2, p38, and c-Jun N-terminal kinase and the translocation of nuclear factor κB and p65 through IRAK-M. We also found that Pro B2-treated macrophages inactivated naïve T cells by inhibiting LPS-induced interferon-γ and IL-2 secretion through IRAK-M. These novel findings provide new insights into the understanding of negative regulatory mechanisms of the TLR4 signaling pathway and the immune-pharmacological role of Pro B2 in the immune response against the development

MiR-146a modulates macrophage polarization by inhibiting Notch1 pathway in RAW264.7 macrophages.

Science.gov (United States)

Huang, Cheng; Liu, Xue-Jiao; QunZhou; Xie, Juan; Ma, Tao-Tao; Meng, Xiao-Ming; Li, Jun

2016-03-01

Macrophages are heterogeneous and plastic cells which are able to undergo dynamic transition between M1 and M2 polarized phenotypes in response to the microenvironment signals. However, the underlying molecular mechanisms of macrophage polarization are still obscure. In the current study, it was revealed that miR-146a might play a pivotal role in macrophage polarization. As our results indicated, miR-146a was highly expressed in M2 macrophages rather than M1 macrophages. Over-expression of miR-146a resulted in significantly decreased production of pro-inflammatory cytokines including iNOS and TNF-α in M1 macrophages, while increased production of M2 marker genes such as Arg1 and CD206 in M2 macrophages. In contrast, knockdown of miR-146a promoted M1 macrophage polarization but diminished M2 macrophage polarization. Mechanistically, it was revealed that miR-146a modulated macrophage polarization by targeting Notch1. Of note, PPARγ was responsible as another target for miR-146a-mediated macrophage polarization. Taken together, it was suggested that miR-146a might serve as a molecular regulator in macrophage polarization and is a potential therapeutic target for inflammatory diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

Purinergic signaling to terminate TLR responses in macrophages

Directory of Open Access Journals (Sweden)

Kajal eHamidzadeh

2016-03-01

Full Text Available Macrophages undergo profound physiological alterations when they encounter pathogen associated molecular patterns (PAMPs. These alterations can result in the elaboration of cytokines and mediators that promote immune responses and contribute to the clearance of pathogens. These innate immune responses by myeloid cells are transient. The termination of these secretory responses is not due to the dilution of stimuli, but rather to the active down-regulation of innate responses induced by the very PAMPs that initiated them. Here we describe a purinergic autoregulatory program whereby TLR-stimulated macrophages control their activation state. In this program, TLR stimulated macrophages undergo metabolic alterations that result in the production of ATP and its release through membrane pannexin channels. This purine nucleotide is rapidly hydrolyzed to adenosine by ectoenzymes on the macrophage surface, CD39 and CD73. Adenosine then signals through the P1 class of seven transmembrane receptors to induce a regulatory state that is characterized by the down-regulation of inflammatory cytokines and the production of anti-inflammatory cytokines and growth factors. This purinergic autoregulatory system mitigates the collateral damage that would be caused by the prolonged activation of macrophages, and rather allows the macrophage to maintain homeostasis. The transient activation of macrophages can be prolonged by treating macrophages with IFN-γ. IFN-γ treated macrophages become less sensitive to the regulatory effects of adenosine, allowing them to sustain macrophage activation for the duration of an adaptive immune response.

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

Science.gov (United States)

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

2015-10-01

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

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

Directory of Open Access Journals (Sweden)

Clemencia Ovalle-Bracho

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

Biomimetic carbon monoxide delivery based on hemoglobin vesicles ameliorates acute pancreatitis in mice via the regulation of macrophage and neutrophil activity.

Science.gov (United States)

Taguchi, Kazuaki; Nagao, Saori; Maeda, Hitoshi; Yanagisawa, Hiroki; Sakai, Hiromi; Yamasaki, Keishi; Wakayama, Tomohiko; Watanabe, Hiroshi; Otagiri, Masaki; Maruyama, Toru

2018-11-01

Macrophages play a central role in various inflammatory disorders and are broadly divided into two subpopulations, M1 and M2 macrophage. In the healing process in acute inflammatory disorders, shifting the production of M1 macrophages to M2 macrophages is desirable, because M1 macrophages secrete pro-inflammatory cytokines, whilst the M2 variety secrete anti-inflammatory cytokines. Previous findings indicate that when macrophages are treated with carbon monoxide (CO), the secretion of anti-inflammatory cytokine is increased and the expression of pro-inflammatory cytokines is inhibited, indicating that CO may have a potential to modulate the production of macrophages toward the M2-like phenotype. In this study, we examined the issue of whether CO targeting macrophages using a nanotechnology-based CO donor, namely CO-bound hemoglobin vesicles (CO-HbV), modulates their polarization and show therapeutic effects against inflammatory disorders. The results showed that the CO-HbV treatment polarized a macrophage cell line toward an M2-like phenotype. Furthermore, in an in vivo study using acute pancreatitis model mice as a model of an inflammatory disease, a CO-HbV treatment also tended to polarize macrophages toward an M2-like phenotype and inhibited neutrophil infiltration in the pancreas, resulting in a significant inflammation. In addition to the suppression of acute pancreatitis, CO-HbV diminished a subsequent pancreatitis-associated acute lung injury. This could be due to the inhibition of the systemic inflammation, neutrophil infiltration in the lungs and the production of HMGB-1. These findings suggest that CO-HbV exerts superior anti-inflammatory effects against inflammatory disorders via the regulation of macrophage and neutrophil activity.

Alteration of Pituitary Tumor Transforming Gene-1 Regulates Trophoblast Invasion via the Integrin/Rho-Family Signaling Pathway.

Directory of Open Access Journals (Sweden)

Seung Mook Lim

Full Text Available Trophoblast invasion ability is an important factor in early implantation and placental development. Recently, pituitary tumor transforming gene 1 (PTTG1 was shown to be involved in invasion and proliferation of cancer. However, the role of PTTG1 in trophoblast invasion remains unknown. Thus, in this study we analyzed PTTG1 expression in trophoblasts and its effect on trophoblast invasion activity and determined the mechanism through which PTTG1 regulates trophoblast invasion. Trophoblast proliferation and invasion abilities, regardless of PTTG1 expression, were analyzed by quantitative real-time polymerase chain reaction, fluorescence-activated cell sorting analysis, invasion assay, western blot, and zymography after treatment with small interfering RNA against PTTG1 (siPTTG1. Additionally, integrin/Rho-family signaling in trophoblasts by PTTG1 alteration was analyzed. Furthermore, the effect of PTTG1 on trophoblast invasion was evaluated by microRNA (miRNA mimic and inhibitor treatment. Trophoblast invasion was significantly reduced through decreased matrix metalloproteinase (MMP-2 and MMP-9 expression when PTTG1 expression was inhibited by siPTTG1 (p < 0.05. Furthermore, knockdown of PTTG1 increased expression of integrin alpha 4 (ITGA4, ITGA5, and integrin beta 1 (ITGB1; otherwise, RhoA expression was significantly decreased (p < 0.05. Treatment of miRNA-186-5p mimic and inhibitor controlled trophoblast invasion ability by altering PTTG1 and MMP expression. PTTG1 can control trophoblast invasion ability via regulation of MMP expression through integrin/Rho-family signaling. In addition, PTTG1 expression and its function were regulated by miRNA-186-5p. These results help in understanding the mechanism through which PTTG1 regulates trophoblast invasion and thereby implantation and placental development.

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

Directory of Open Access Journals (Sweden)

Van Parys Alexander

2012-06-01

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

TAM receptor-dependent regulation of SOCS3 and MAPKs contributes to proinflammatory cytokine downregulation following chronic NOD2 stimulation of human macrophages.

Science.gov (United States)

Zheng, Shasha; Hedl, Matija; Abraham, Clara

2015-02-15

Microbial-induced cytokine regulation is critical to intestinal immune homeostasis. Acute stimulation of nucleotide-binding oligomerization domain 2 (NOD2), the Crohn's disease-associated sensor of bacterial peptidoglycan, induces cytokines. However, cytokines are attenuated after chronic NOD2 and pattern recognition receptor stimulation of macrophages; similar attenuation is observed in intestinal macrophages. The role of Tyro3, Axl, and Mer (TAM) receptors in regulating chronic pattern recognition receptor stimulation and NOD2-induced outcomes has not been examined. Moreover, TAM receptors have been relatively less investigated in human macrophages. Whereas TAM receptors did not downregulate acute NOD2-induced cytokines in primary human macrophages, they were essential for downregulating signaling and proinflammatory cytokine secretion after chronic NOD2 and TLR4 stimulation. Axl and Mer were similarly required in mice for cytokine downregulation after chronic NOD2 stimulation in vivo and in intestinal tissues. Consistently, TAM expression was increased in human intestinal myeloid-derived cells. Chronic NOD2 stimulation led to IL-10- and TGF-β-dependent TAM upregulation in human macrophages, which, in turn, upregulated suppressor of cytokine signaling 3 expression. Restoring suppressor of cytokine signaling 3 expression under TAM knockdown conditions restored chronic NOD2-mediated proinflammatory cytokine downregulation. In contrast to the upregulated proinflammatory cytokines, attenuated IL-10 secretion was maintained in TAM-deficient macrophages upon chronic NOD2 stimulation. The level of MAPK activation in TAM-deficient macrophages after chronic NOD2 stimulation was insufficient to upregulate IL-10 secretion; however, full restoration of MAPK activation under these conditions restored c-Fos, c-Jun, musculoaponeurotic fibrosarcoma oncogene homolog K, and PU.1 binding to the IL-10 promoter and IL-10 secretion. Therefore, TAM receptors are critical for

stg fimbrial operon from S. Typhi STH2370 contributes to association and cell disruption of epithelial and macrophage-like cells.

Science.gov (United States)

Berrocal, Liliana; Fuentes, Juan A; Trombert, A Nicole; Jofré, Matías R; Villagra, Nicolás A; Valenzuela, Luis M; Mora, Guido C

2015-07-07

Salmonella enterica serovar Typhi (S. Typhi) stg operon, encoding a chaperone/usher fimbria (CU), contributes to an increased adherence to human epithelial cells. However, one report suggests that the presence of the Stg fimbria impairs the monocyte--bacteria association, as deduced by the lower level of invasion to macrophage-like cells observed when the stg fimbrial cluster was overexpressed. Nevertheless, since other CU fimbrial structures increase the entry of S. Typhi into macrophages, and considering that transcriptomic analyses revealed that stg operon is indeed expressed in macrophages, we reassessed the role of the stg operon in the interaction between S. Typhi strain STH2370 and human cells, including macrophage-like cells and mononuclear cells directly taken from human peripheral blood. We compared S. Typhi STH2370 WT, a Chilean clinical strain, and the S. Typhi STH2370 Δstg mutant with respect to association and invasion using epithelial and macrophage-like cells. We observed that deletion of stg operon reduced the association and invasion of S. Typhi, in both cellular types. The presence of the cloned stg operon restored the WT phenotype in all the cases. Moreover, we compared Salmonella enterica sv. Typhimurium 14028s (S. Typhimurium, a serovar lacking stg operon) and S. Typhimurium heterologously expressing S. Typhi stg. We found that the latter presents an increased cell disruption of polarized epithelial cells and an increased association in both epithelial and macrophage-like cells. S. Typhi stg operon encodes a functional adhesin that participates in the interaction bacteria-eukaryotic cells, including epithelial cells and macrophages-like cells. The phenotypes associated to stg operon include increased association and consequent invasion in bacteria-eukaryotic cells, and cell disruption.

From mini-invasive to non-invasive treatment using monopolar radiofrequency: the next orthopaedic frontier.

Science.gov (United States)

Whipple, Terry L

2009-10-01

Tendinopathy arises from a failed tendon healing process. Current non-invasive therapeutic alternatives are anti-inflammatory in nature, and outcomes are unpredictable. The benefit of invasive alternatives resides in the induction of the healing response. A new technology that uses non-invasive monopolar capacitive coupled radiofrequency has demonstrated the ability to raise temperatures in tendons and ligaments above 50 degrees C, the threshold for collagen modulation, tissue shrinkage and recruitment of macrophages, fibroblasts, and heat shock protein factors, without damaging the overlying structures, resulting in activation of the wound healing response. Monopolar capacitive-coupled radiofrequency offers a new non-invasive choice for tendinopathies and sprained ligaments. It does not interfere with subsequent surgical procedures should they become necessary.

DMPD: Distinct functions of IRF-3 and IRF-7 in IFN-alpha gene regulation and controlof anti-tumor activity in primary macrophages. [Dynamic Macrophage Pathway CSML Database

Lifescience Database Archive (English)

Full Text Available 16846591 Distinct functions of IRF-3 and IRF-7 in IFN-alpha gene regulation and con...cott J. Biochem Pharmacol. 2006 Nov 30;72(11):1469-76. Epub 2006 Jul 17. (.png) (.svg) (.html) (.csml) Show Distinct function...anti-tumor activity in primary macrophages. PubmedID 16846591 Title Distinct functions of IRF-3 and IRF-7 in

Resveratrol Regulates Colorectal Cancer Cell Invasion by Modulation of Focal Adhesion Molecules.

Science.gov (United States)

Buhrmann, Constanze; Shayan, Parviz; Goel, Ajay; Shakibaei, Mehdi

2017-09-27

Resveratrol, a safe and multi-targeted agent, has been associated with suppression of survival, proliferation and metastasis of cancer, however, the underlying mechanisms for its anti-cancer activity, particularly on cellular signaling during cancer cell migration still remain poorly understood. We investigated the invasion response of two human colorectal cancer (CRC) cells (HCT116 and SW480) to resveratrol and studied the effect of specific pharmacological inhibitors, cytochalasin D (CytD) and focal adhesion kinase-inhibitor (FAK-I) on FAK, cell viability and migration in CRC. We found that resveratrol altered cell phenotype of both CRC cells, reduced cell viability and the results were comparable to FAK-I and CytD. These effects of resveratrol were associated with marked Sirt1 up-regulation, FAK down-regulation, inhibition of focal adhesion and potentiation of effects by combinatorial treatment of resveratrol and inhibitors. Interestingly, inhibition of FAK with FAK-I or treatment with CytD suppressed resveratrol-induced Sirt1 up-regulation and markedly down-regulated FAK expression. Resveratrol or combination treatment with inhibitors significantly activated caspase-3 and potentiated apoptosis. Moreover, resveratrol suppressed invasion and colony forming capacity, cell proliferation, β1-Integrin expression and activation of FAK of cells in alginate tumor microenvironment, similar to FAK-I or CytD. Finally, we demonstrated that resveratrol, FAK-I or CytD inhibited activation of NF-κB, suppressed NF-κB-dependent gene end-products involved in invasion, metastasis, and apoptosis; and these effects of resveratrol were potentiated by combination treatment with FAK-I or CytD. Our data illustrated that the anti-invasion effect of resveratrol by inhibition of FAK activity has a potential beneficial role in disease prevention and therapeutic management of CRC.

Transcriptional landscape of Mycobacterium tuberculosis infection in macrophages

KAUST Repository

Roy, Sugata

2018-04-24

Mycobacterium tuberculosis (Mtb) infection reveals complex and dynamic host-pathogen interactions, leading to host protection or pathogenesis. Using a unique transcriptome technology (CAGE), we investigated the promoter-based transcriptional landscape of IFNγ (M1) or IL-4/IL-13 (M2) stimulated macrophages during Mtb infection in a time-kinetic manner. Mtb infection widely and drastically altered macrophage-specific gene expression, which is far larger than that of M1 or M2 activations. Gene Ontology enrichment analysis for Mtb-induced differentially expressed genes revealed various terms, related to host-protection and inflammation, enriched in up-regulated genes. On the other hand, terms related to dis-regulation of cellular functions were enriched in down-regulated genes. Differential expression analysis revealed known as well as novel transcription factor genes in Mtb infection, many of them significantly down-regulated. IFNγ or IL-4/IL-13 pre-stimulation induce additional differentially expressed genes in Mtb-infected macrophages. Cluster analysis uncovered significant numbers, prolonging their expressional changes. Furthermore, Mtb infection augmented cytokine-mediated M1 and M2 pre-activations. In addition, we identified unique transcriptional features of Mtb-mediated differentially expressed lncRNAs. In summary we provide a comprehensive in depth gene expression/regulation profile in Mtb-infected macrophages, an important step forward for a better understanding of host-pathogen interaction dynamics in Mtb infection.

Rab39a interacts with phosphatidylinositol 3-kinase and negatively regulates autophagy induced by lipopolysaccharide stimulation in macrophages.

Directory of Open Access Journals (Sweden)

Shintaro Seto

Full Text Available Rab39a has pleiotropic functions in phagosome maturation, inflammatory activation and neuritogenesis. Here, we characterized Rab39a function in membrane trafficking of phagocytosis and autophagy induction in macrophages. Rab39a localized to the periphery of LAMP2-positive vesicles and showed the similar kinetics on the phagosome to that of LAMP1. The depletion of Rab39a did not influence the localization of LAMP2 to the phagosome, but it augments the autophagosome formation and LC3 processing by lipopolysaccharide (LPS stimulation. The augmentation of autophagosome formation in Rab39a-knockdown macrophages was suppressed by Atg5 depletion or an inhibitor for phosphatidylinostol 3-kinase (PI3K. Immunoprecipitation analysis revealed that Rab39a interacts with PI3K and that the amino acid residues from 34(th to 41(st in Rab39a were indispensable for this interaction. These results suggest that Rab39a negatively regulates the LPS-induced autophagy in macrophages.

Activation of PPARγ by a Natural Flavonoid Modulator, Apigenin Ameliorates Obesity-Related Inflammation Via Regulation of Macrophage Polarization

Directory of Open Access Journals (Sweden)

Xiujing Feng

2016-07-01

Full Text Available PPARγ has emerged as a master regulator of macrophage polarization and is the molecular target of the thiazolidinedione drugs. Here we show that apigenin binds and activates PPARγ by acting as a modulator. Activation of PPARγ by apigenin blocks p65 translocation into nuclei through inhibition of p65/PPARγ complex translocation into nuclei, thereby decreasing NF-κB activation and favoringM2 macrophage polarization. In HFD and ob/ob mice, apigenin significantly reverses M1 macrophage into M2 and reduces the infiltration of inflammatory cells in liver and adipose tissues, as well as decreases the levels of pro-inflammatory cytokines, thereby alleviating inflammation. Strikingly, apigenin reduces liver and muscular steatosis, decreases the levels of ALT, AST, TC and TG, improving glucose resistance obviously. Unlike rosiglitazone, apigenin does not cause significant weight gain, osteoporosis et al. Our findings identify apigenin as a modulator of PPARγ and a potential lead compound for treatment of metabolic disorders.

Effects of lipopolysaccharide on the catabolic activity of macrophages

International Nuclear Information System (INIS)

Cluff, C.; Ziegler, H.K.

1986-01-01

The ability of macrophages to degrade and catabolize antigens is of relevance both as a means to process complex antigens prior to presentation to T cells, as well as a way to down regulate immune responses by destroying the antigenicity of polypeptides. With these considerations, the authors have investigated the regulation of macrophage catabolic activity by lipopolysaccharide (LPS). Catabolic activity was quantitated by following the distribution and molecular form of 125 -I labelled surface components of heat-killed Listeria monocytogenes (HKLM) subsequent to their uptake by macrophages. They have compared the catabolic activity of macrophages from peritoneal exudates of mice injected i.p. with saline or LPS and have found that LPS-elicited macrophages display a greatly enhanced (3 fold) rate of catabolism. This increase in catabolic activity peaks 3 days after LPS injection and steadily declines thereafter, approaching a baseline level after 3 weeks. The enhancement of catabolic activity is under LPS gene control. LPS-elicited macrophages rapidly destroy the antigenicity of bacterial antigens and function poorly as antigen presenting cells in vitro. These results suggest that LPS elicits a macrophage population specialized for antigen degradation functions with negative regulatory effects on the induction of specific immune responses

Role of Macrophage-Induced Inflammation in Mesothelioma

Science.gov (United States)

2011-07-01

macrophages). Normal pleura becomes available intermittently , serving to slow the completion of this task. All is set in place for us to complete the...GFP) regulated by a Csf1r-promoter (Sasmono et al. 2003) show that macrophages travel up and down these fibers at a fast rate and also “jump” between...2010). Macrophages have also recently been shown to be important in adipogenesis at least during obesity , through their secretion of adipocyte growth

Biaryl amide compounds reduce the inflammatory response in macrophages by regulating Dectin-1.

Science.gov (United States)

Hyung, Kyeong Eun; Lee, Mi Ji; Lee, Yun-Jung; Lee, Do Ik; Min, Hye Young; Park, So-Young; Min, Kyung Hoon; Hwang, Kwang Woo

2016-03-01

Macrophages are archetypal innate immune cells that play crucial roles in the recognition and phagocytosis of invading pathogens, which they identify using pattern recognition receptors (PRRs). Dectin-1 is essential for antifungal immune responses, recognizing the fungal cellular component β-glucan, and its role as a PRR has been of increasing interest. Previously, we discovered and characterized a novel biaryl amide compound, MPS 03, capable of inhibiting macrophage phagocytosis of zymosan. Therefore, in this study we aimed to identify other biaryl amide compounds with greater effectiveness than MPS 03, and elucidate their cellular mechanisms. Several MPS 03 derivatives were screened, four of which reduced zymosan phagocytosis in a similar manner to MPS 03. To establish whether such phagocytosis inhibition influenced the production of inflammatory mediators, pro-inflammatory cytokine and nitric oxide (NO) levels were measured. The production of TNF-α, IL-6, IL-12, and NO was significantly reduced in a dose-dependent manner. Moreover, the inflammation-associated MAPK signaling pathway was also affected by biaryl amide compounds. To investigate the underlying cellular mechanism, PRR expression was measured. MPS 03 and its derivatives were found to inhibit zymosan phagocytosis by decreasing Dectin-1 expression. Furthermore, when macrophages were stimulated by zymosan after pretreatment with biaryl amide compounds, downstream transcription factors such as NFAT, AP-1, and NF-κB were downregulated. In conclusion, biaryl amide compounds reduce zymosan-induced inflammatory responses by downregulating Dectin-1 expression. Therefore, such compounds could be used to inhibit Dectin-1 in immunological experiments and possibly regulate excessive inflammatory responses. Copyright © 2016. Published by Elsevier B.V.

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

Directory of Open Access Journals (Sweden)

Mário Henrique M Barros

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

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

Science.gov (United States)

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

2013-01-01

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

Preservation Analysis of Macrophage Gene Coexpression Between Human and Mouse Identifies PARK2 as a Genetically Controlled Master Regulator of Oxidative Phosphorylation in Humans

Directory of Open Access Journals (Sweden)

Veronica Codoni

2016-10-01

Full Text Available Macrophages are key players involved in numerous pathophysiological pathways and an in-depth characterization of their gene regulatory networks can help in better understanding how their dysfunction may impact on human diseases. We here conducted a cross-species network analysis of macrophage gene expression data between human and mouse to identify conserved networks across both species, and assessed whether such networks could reveal new disease-associated regulatory mechanisms. From a sample of 684 individuals processed for genome-wide macrophage gene expression profiling, we identified 27 groups of coexpressed genes (modules. Six modules were found preserved (P < 10−4 in macrophages from 86 mice of the Hybrid Mouse Diversity Panel. One of these modules was significantly [false discovery rate (FDR = 8.9 × 10−11] enriched for genes belonging to the oxidative phosphorylation (OXPHOS pathway. This pathway was also found significantly (FDR < 10−4 enriched in susceptibility genes for Alzheimer, Parkinson, and Huntington diseases. We further conducted an expression quantitative trait loci analysis to identify SNP that could regulate macrophage OXPHOS gene expression in humans. This analysis identified the PARK2 rs192804963 as a trans-acting variant influencing (minimal P-value = 4.3 × 10−8 the expression of most OXPHOS genes in humans. Further experimental work demonstrated that PARK2 knockdown expression was associated with increased OXPHOS gene expression in THP1 human macrophages. This work provided strong new evidence that PARK2 participates to the regulatory networks associated with oxidative phosphorylation and suggested that PARK2 genetic variations could act as a trans regulator of OXPHOS gene macrophage expression in humans.

Macrophage markers in serum and tumor have prognostic impact in American Joint Committee on Cancer stage I/II melanoma

DEFF Research Database (Denmark)

Jensen, Trine O.; Schmidt, Henrik; Møller, Holger John

2009-01-01

PURPOSE: To evaluate the prognostic role of soluble CD163 (sCD163) in serum and macrophage infiltration in primary melanomas from patients with American Joint Committee on Cancer (AJCC) stage I/II melanoma. The scavenger receptor CD163 is associated with anti-inflammatory macrophages...... melanomas from 190 patients were available for immunohistochemical analyzes of CD163(+) and CD68(+) macrophage infiltration. They were estimated semiquantitatively in three different tumor compartments: tumor nests, tumor stroma, and at the invasive front of the tumor. RESULTS: Serum sCD163 treated......, HR = 1.4; 95% CI, 1.1 to 1.8; P = .003). Melanomas with dense CD163(+) macrophage infiltration in tumor stroma and CD68(+) macrophage infiltration at the invasive front were associated with poor overall survival (CD163, HR = 2.7; 95% CI, 0.8 to 9.3; P = .11; and CD68, HR = 2.8; 95% CI, 1.2 to 6.8; P...

Myelin down-regulates myelin phagocytosis by microglia and macrophages through interactions between CD47 on myelin and SIRPα (signal regulatory protein-α on phagocytes

Directory of Open Access Journals (Sweden)

Reichert Fanny

2011-03-01

Full Text Available Abstract Background Traumatic injury to axons produces breakdown of axons and myelin at the site of the lesion and then further distal to this where Wallerian degeneration develops. The rapid removal of degenerated myelin by phagocytosis is advantageous for repair since molecules in myelin impede regeneration of severed axons. Thus, revealing mechanisms that regulate myelin phagocytosis by macrophages and microglia is important. We hypothesize that myelin regulates its own phagocytosis by simultaneous activation and down-regulation of microglial and macrophage responses. Activation follows myelin binding to receptors that mediate its phagocytosis (e.g. complement receptor-3, which has been previously studied. Down-regulation, which we test here, follows binding of myelin CD47 to the immune inhibitory receptor SIRPα (signal regulatory protein-α on macrophages and microglia. Methods CD47 and SIRPα expression was studied by confocal immunofluorescence microscopy, and myelin phagocytosis by ELISA. Results We first document that myelin, oligodendrocytes and Schwann cells express CD47 without SIRPα and further confirm that microglia and macrophages express both CD47 and SIRPα. Thus, CD47 on myelin can bind to and subsequently activate SIRPα on phagocytes, a prerequisite for CD47/SIRPα-dependent down-regulation of CD47+/+ myelin phagocytosis by itself. We then demonstrate that phagocytosis of CD47+/+ myelin is augmented when binding between myelin CD47 and SIRPα on phagocytes is blocked by mAbs against CD47 and SIRPα, indicating that down-regulation of phagocytosis indeed depends on CD47-SIRPα binding. Further, phagocytosis in serum-free medium of CD47+/+ myelin is augmented after knocking down SIRPα levels (SIRPα-KD in phagocytes by lentiviral infection with SIRPα-shRNA, whereas phagocytosis of myelin that lacks CD47 (CD47-/- is not. Thus, myelin CD47 produces SIRPα-dependent down-regulation of CD47+/+ myelin phagocytosis in phagocytes

Activating transcription factor 6 mediates oxidized LDL-induced cholesterol accumulation and apoptosis in macrophages by up-regulating CHOP expression.

Science.gov (United States)

Yao, Shutong; Zong, Chuanlong; Zhang, Ying; Sang, Hui; Yang, Mingfeng; Jiao, Peng; Fang, Yongqi; Yang, Nana; Song, Guohua; Qin, Shucun

2013-01-01

This study was to explore whether activating transcription factor 6 (ATF6), an important sensor to endoplasmic reticulum (ER) stress, would mediate oxidized low-density lipoprotein (ox-LDL)- induced cholesterol accumulation and apoptosis in cultured macrophages and the underlying molecular mechanisms. Intracellular lipid droplets and total cholesterol levels were assayed by oil red O staining and enzymatic colorimetry, respectively. Cell viability and apoptosis were determined using MTT assay and AnnexinV-FITC apoptosis detection kit, respectively. The nuclear translocation of ATF6 in cells was detected by immunofluorescence analysis. Protein and mRNA levels were examined by Western blot analysis and real time-PCR, respectively. ATF6 siRNA was transfected to RAW264.7 cells by lipofectamin. Exposure of cells to ox-LDL induced glucose-regulated protein 78 (GRP78). C/EBP homologous protein (CHOP), a key-signaling component of ER stress-induced apoptosis, was up-regulated in ox-LDL-treated cells. ATF6, a factor that positively regulates CHOP expression, was activated by ox-LDL in a concentration- and time- dependent manner. The role of the ATF6-mediated ER stress pathway was further confirmed through the siRNA-mediated knockdown of ATF6, which attenuated ox-LDL-induced upregulation of CHOP, cholesterol accumulation and apoptosis in macrophages. In addition, the phosphorylation of double-stranded RNA-activated protein kinase-like endoplasmic reticulum kinase (PERK), another factor that positively regulates CHOP expression, was induced in the presence of ox-LDL, and PERK-specific siRNA also inhibited the ox-LDL-induced upregulation of CHOP and apoptosis in RAW264.7 cells. These results demonstrate that ER stress-related proteins, particularly ATF6 and its downstream molecule CHOP, are involved in ox-LDL-induced cholesterol accumulation and apoptosis in macrophages.

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

Directory of Open Access Journals (Sweden)

Qiwei Wang

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

Novel Role of Endogenous Catalase in Macrophage Polarization in Adipose Tissue.

Science.gov (United States)

Park, Ye Seul; Uddin, Md Jamal; Piao, Lingjuan; Hwang, Inah; Lee, Jung Hwa; Ha, Hunjoo

2016-01-01

Macrophages are important components of adipose tissue inflammation, which results in metabolic diseases such as insulin resistance. Notably, obesity induces a proinflammatory phenotypic switch in adipose tissue macrophages, and oxidative stress facilitates this switch. Thus, we examined the role of endogenous catalase, a key regulator of oxidative stress, in the activity of adipose tissue macrophages in obese mice. Catalase knockout (CKO) exacerbated insulin resistance, amplified oxidative stress, and accelerated macrophage infiltration into epididymal white adipose tissue in mice on normal or high-fat diet. Interestingly, catalase deficiency also enhanced classical macrophage activation (M1) and inflammation but suppressed alternative activation (M2) regardless of diet. Similarly, pharmacological inhibition of catalase activity using 3-aminotriazole induced the same phenotypic switch and inflammatory response in RAW264.7 macrophages. Finally, the same phenotypic switch and inflammatory responses were observed in primary bone marrow-derived macrophages from CKO mice. Taken together, the data indicate that endogenous catalase regulates the polarization of adipose tissue macrophages and thereby inhibits inflammation and insulin resistance.

Immunoregulation of bovine macrophages by factors in the salivary glands of Rhipicephalus microplus

Directory of Open Access Journals (Sweden)

Brake Danett K

2012-02-01

Full Text Available Abstract Background Alternative strategies are required to control the southern cattle tick, Rhipicephalus microplus, due to evolving resistance to commercially available acaricides. This invasive ectoparasite is a vector of economically important diseases of cattle such as bovine babesiosis and anaplasmosis. An understanding of the biological intricacies underlying vector-host-pathogen interactions is required to innovate sustainable tick management strategies that can ultimately mitigate the impact of animal and zoonotic tick-borne diseases. Tick saliva contains molecules evolved to impair host innate and adaptive immune responses, which facilitates blood feeding and pathogen transmission. Antigen presenting cells are central to the development of robust T cell responses including Th1 and Th2 determination. In this study we examined changes in co-stimulatory molecule expression and cytokine response of bovine macrophages exposed to salivary gland extracts (SGE obtained from 2-3 day fed, pathogen-free adult R. microplus. Methods Peripheral blood-derived macrophages were treated for 1 hr with 1, 5, or 10 μg/mL of SGE followed by 1, 6, 24 hr of 1 μg/mL of lipopolysaccharide (LPS. Real-time PCR and cytokine ELISA were used to measure changes in co-stimulatory molecule expression and cytokine response. Results Changes were observed in co-stimulatory molecule expression of bovine macrophages in response to R. microplus SGE exposure. After 6 hrs, CD86, but not CD80, was preferentially up-regulated on bovine macrophages when treated with 1 μg/ml SGE and then LPS, but not SGE alone. At 24 hrs CD80, CD86, and CD69 expression was increased with LPS, but was inhibited by the addition of SGE. SGE also inhibited LPS induced upregulation of TNFα, IFNγ and IL-12 cytokines, but did not alter IL-4 or CD40 mRNA expression. Conclusions Molecules from the salivary glands of adult R. microplus showed bimodal concentration-, and time-dependent effects on

Fibroblast growth factor receptor 1 activation in mammary tumor cells promotes macrophage recruitment in a CX3CL1-dependent manner.

Directory of Open Access Journals (Sweden)

Johanna R Reed

Full Text Available Tumor formation is an extensive process requiring complex interactions that involve both tumor cell-intrinsic pathways and soluble mediators within the microenvironment. Tumor cells exploit the intrinsic functions of many soluble molecules, including chemokines and their receptors, to regulate pro-tumorigenic phenotypes that are required for growth and progression of the primary tumor. Previous studies have shown that activation of inducible FGFR1 (iFGFR1 in mammary epithelial cells resulted in increased proliferation, migration, and invasion in vitro and tumor formation in vivo. These studies also demonstrated that iFGFR1 activation stimulated recruitment of macrophages to the epithelium where macrophages contributed to iFGFR1-mediated epithelial cell proliferation and angiogenesis. The studies presented here further utilize this model to identify the mechanisms that regulate FGFR1-induced macrophage recruitment. Results from this study elucidate a novel role for the inflammatory chemokine CX3CL1 in FGFR1-induced macrophage migration. Specifically, we illustrate that activation of both the inducible FGFR1 construct in mouse mammary epithelial cells and endogenous FGFR in the triple negative breast cancer cell line, HS578T, leads to expression of the chemokine CX3CL1. Furthermore, we demonstrate that FGFR-induced CX3CL1 is sufficient to recruit CX3CR1-expressing macrophages in vitro. Finally, blocking CX3CR1 in vivo leads to decreased iFGFR1-induced macrophage recruitment, which correlates with decreased angiogenesis. While CX3CL1 is a known target of FGF signaling in the wound healing environment, these studies demonstrate that FGFR activation also leads to induction of CX3CL1 in a tumor setting. Furthermore, these results define a novel role for CX3CL1 in promoting macrophage recruitment during mammary tumor formation, suggesting that the CX3CL1/CX3CR1 axis may represent a potential therapeutic approach for targeting breast cancers associated

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

Science.gov (United States)

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

2015-11-01

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

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

Science.gov (United States)

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

2015-06-01

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

Tumor-associated macrophages in glioblastoma multiforme-a suitable target for somatostatin receptor-based imaging and therapy?

Directory of Open Access Journals (Sweden)

Constantin Lapa

Full Text Available Glioblastoma multiforme (GBM is the most common primary brain tumor in adults. Tumor-associated macrophages (TAM have been shown to promote malignant growth and to correlate with poor prognosis. [1,4,7,10-tetraazacyclododecane-NN',N″,N'″-tetraacetic acid]-d-Phe1,Tyr3-octreotate (DOTATATE labeled with Gallium-68 selectively binds to somatostatin receptor 2A (SSTR2A which is specifically expressed and up-regulated in activated macrophages. On the other hand, the role of SSTR2A expression on the cell surface of glioma cells has not been fully elucidated yet. The aim of this study was to non-invasively assess SSTR2A expression of both glioma cells as well as macrophages in GBM.15 samples of patient-derived GBM were stained immunohistochemically for macrophage infiltration (CD68, proliferative activity (Ki67 as well as expression of SSTR2A. Anti-CD45 staining was performed to distinguish between resident microglia and tumor-infiltrating macrophages. In a subcohort, positron emission tomography (PET imaging using 68Ga-DOTATATE was performed and the semiquantitatively evaluated tracer uptake was compared to the results of immunohistochemistry.The amount of microglia/macrophages ranged from 50% in the tumor samples with the vast majority being resident microglial cells. A strong SSTR2A immunostaining was observed in endothelial cells of proliferating vessels, in neurons and neuropile. Only faint immunostaining was identified on isolated microglial and tumor cells. Somatostatin receptor imaging revealed areas of increased tracer accumulation in every patient. However, retention of the tracer did not correlate with immunohistochemical staining patterns.SSTR2A seems not to be overexpressed in GBM samples tested, neither on the cell surface of resident microglia or infiltrating macrophages, nor on the surface of tumor cells. These data suggest that somatostatin receptor directed imaging and treatment strategies are less promising in GBM.

Expression and regulation of HIF-1 alpha in macrophages under inflammatory conditions; significant reduction of VEGF by CaMKII inhibitor

NARCIS (Netherlands)

Westra, Johanna; Brouwer, Elisabeth; van Roosmalen, Ingrid A. M.; Doornbos-van der Meer, Berber; van Leeuwen, Miek A.; Posthumus, Marcel D.; Kallenberg, Cees G. M.

2010-01-01

Background: Macrophages expressing the pro-angiogenic transcription factor hypoxia-inducible factor (HIF)-1alpha have been demonstrated in rheumatoid arthritis (RA) in the synovial tissue. Aim of the present study was to investigate intracellular signal transduction regulation of pro-inflammatory

Phosphorylation of the adaptor protein SH2B1β regulates its ability to enhance growth hormone-dependent macrophage motility

OpenAIRE

Su, Hsiao-Wen; Lanning, Nathan J.; Morris, David L.; Argetsinger, Lawrence S.; Lumeng, Carey N.; Carter-Su, Christin

2013-01-01

Previous studies have shown that growth hormone (GH) recruits the adapter protein SH2B1β to the GH-activated, GH receptor-associated tyrosine kinase JAK2, implicating SH2B1β in GH-dependent actin cytoskeleton remodeling, and suggesting that phosphorylation at serines 161 and 165 in SH2B1β releases SH2B1β from the plasma membrane. Here, we examined the role of SH2B1β in GH regulation of macrophage migration. We show that GH stimulates migration of cultured RAW264.7 macrophages, and primary cul...

M1 Macrophages but Not M2 Macrophages Are Characterized by Upregulation of CRP Expression via Activation of NFκB: a Possible Role for Ox-LDL in Macrophage Polarization.

Science.gov (United States)

Kaplan, Marielle; Shur, Anna; Tendler, Yvgeny

2018-04-23

Arterial macrophages comprise a heterogeneous population: pro-inflammatory (M1) and anti-inflammatory (M2). Since C-reactive protein (CRP) is produced by macrophages in atherosclerotic lesions, understanding of CRP regulation in macrophages could be crucial to decipher inflammatory patterns in atherogenesis. We aimed to analyze CRP expression in M1/M2 macrophages and to question whether it involves NFκB signaling pathway. Furthermore, we questioned whether oxidative stress affect macrophage phenotype and modulate macrophage CRP expression. M1/M2 macrophage polarization was validated using THP-1 macrophages. CRP mRNA and protein expression were determined using real-time PCR and immunohistochemistry. Involvement of NFκB was determined by nuclear translocation of p50 subunit and the use of NFκB inhibitor. Involvement of oxidative stress in macrophage phenotypes induction was studied using oxidized-LDL (Ox-LDL) and antioxidants. M1 macrophages were characterized by elevated CRP mRNA expression (by 67%), CRP protein levels (by 108%), and upregulation of NFκB activation compared to control, but these features were not shared by M2 macrophages. Macrophages incubation with Ox-LDL led to a moderate M1 phenotype combined with a M2 phenotype, correlated with increased CRP mRNA expression. Antioxidants inhibited by up to 86% IL6 expression but did not significantly affect IL10 secretion. Antioxidants significantly inhibited CRP expression in M1 macrophages, but not in M2 macrophages. Elevated expression of CRP was characteristic of M1 macrophages rather than M2 through NFκB activation. Oxidative stress could be one of the endogenous triggers for macrophage activation to a mixed M1 and M2 phenotype, in association with increased expression of CRP.

miR-29b and miR-125a Regulate Podoplanin and Suppress Invasion in Glioblastoma

Science.gov (United States)

Cortez, Maria Angelica; Nicoloso, Milena Sabrina; Shimizu, Masayoshi; Rossi, Simona; Gopisetty, Gopal; Molina, Jennifer R.; Carlotti, Carlos; Tirapelli, Daniela; Neder, Luciano; Brassesco, Maria Sol; Scrideli, Carlos Alberto; Tone, Luiz Gonzaga; Georgescu, Maria-Magdalena; Zhang, Wei; Puduvalli, Vinay; Calin, George Adrian

2017-01-01

Glioblastoma is the most frequent and malignant brain tumor, characterized by an elevated capacity for cellular proliferation and invasion. Recently, it was demonstrated that podoplanin membrane sialo-glycoprotein encoded by PDPN gene is over-expressed and related to cellular invasion in astrocytic tumors; however the mechanisms of regulation are still unknown. MicroRNAs are noncoding RNAs that regulate gene expression and several biological processes and diseases, including cancer. Nevertheless, their roles in invasion, proliferation, and apoptosis of glioblastoma are not completely understood. In this study, we focused on miR-29b and miR-125a, which were predicted to regulate PDPN, and demonstrated that these microRNAs directly target the 3′ untranslated region of PDPN and inhibit invasion, apoptosis, and proliferation of glioblastomas. Furthermore, we report that miR-29b and miR-125a are downregulated in glioblastomas and also in CD133-positive cells. Taken together, these results suggest that miR-29b and miR-125a represent potential therapeutic targets in glioblastoma. PMID:20665731

Contribution of Hfe expression in macrophages to the regulation of hepatic hepcidin levels and iron loading

OpenAIRE

Makui, Hortence; Soares, Ricardo J.; Jiang, Wenlei; Constante, Marco; Santos, Manuela M.

2005-01-01

Hereditary hemochromatosis (HH), an iron overload disease associated with mutations in the HFE gene, is characterized by increased intestinal iron absorption and consequent deposition of excess iron, primarily in the liver. Patients with HH and Hfe-deficient (Hfe−/−) mice manifest inappropriate expression of the iron absorption regulator hepcidin, a peptide hormone produced by the liver in response to iron loading. In this study, we investigated the contribution of Hfe expression in macrophag...

In vitro modification of Candida albicans invasiveness.

Science.gov (United States)

Fontenla de Petrino, S E; de Jorrat, M E; Sirena, A; Valdez, J C; Mesón, O

1986-05-01

Candida albicans produces germ-tubes (GT) when it is incubated in animal or human serum. This dimorphism is responsible for its invasive ability. The purpose of the present paper is (1) to evaluate the ability of rat peritoneal macrophages to inhibit GT production of ingested Candida albicans, obtained from immunized rats and then activated in vitro with Candida-induced lymphokines; (2) to determinate any possible alteration of phagocytic and candidacidal activities. The phagocytes were obtained from rats immunized with viable C. albicans. Some of them were exposed to Candida-induced lymphokines in order to activate the macrophages in vitro. The monolayers of activated, immune and normal macrophages were infected with a C. albicans suspension during 4 hr. Activated macrophages presented not only the highest phagocytic and candidacidal activities but a noticeable inhibition of GT formation and incremented candidacidal activity.

Macrophages Contribute to the Spermatogonial Niche in the Adult Testis

Directory of Open Access Journals (Sweden)

Tony DeFalco

2015-08-01

Full Text Available The testis produces sperm throughout the male reproductive lifespan by balancing self-renewal and differentiation of spermatogonial stem cells (SSCs. Part of the SSC niche is thought to lie outside the seminiferous tubules of the testis; however, specific interstitial components of the niche that regulate spermatogonial divisions and differentiation remain undefined. We identified distinct populations of testicular macrophages, one of which lies on the surface of seminiferous tubules, in close apposition to areas of tubules enriched for undifferentiated spermatogonia. These macrophages express spermatogonial proliferation- and differentiation-inducing factors, such as colony-stimulating factor 1 (CSF1 and enzymes involved in retinoic acid (RA biosynthesis. We show that transient depletion of macrophages leads to a disruption in spermatogonial differentiation. These findings reveal an unexpected role for macrophages in the spermatogonial niche in the testis and raise the possibility that macrophages play previously unappreciated roles in stem/progenitor cell regulation in other tissues.

An mDia2/ROCK signaling axis regulates invasive egress from epithelial ovarian cancer spheroids.

Science.gov (United States)

Pettee, Krista M; Dvorak, Kaitlyn M; Nestor-Kalinoski, Andrea L; Eisenmann, Kathryn M

2014-01-01

Multi-cellular spheroids are enriched in ascites of epithelial ovarian cancer (OvCa) patients. They represent an invasive and chemoresistant cellular population fundamental to metastatic dissemination. The molecular mechanisms triggering single cell invasive egress from spheroids remain enigmatic. mDia formins are Rho GTPase effectors that are key regulators of F-actin cytoskeletal dynamics. We hypothesized that mDia2-driven F-actin dynamics promote single cell invasive transitions in clinically relevant three-dimensional (3D) OvCa spheroids. The current study is a dissection of the contribution of the F-actin assembly factor mDia2 formin in invasive transitions and using a clinically relevant ovarian cancer spheroid model. We show that RhoA-directed mDia2 activity is required for tight spheroid organization, and enrichment of mDia2 in the invasive cellular protrusions of collagen-embedded OVCA429 spheroids. Depleting mDia2 in ES-2 spheroids enhanced invasive dissemination of single amoeboid-shaped cells. This contrasts with spheroids treated with control siRNA, where a mesenchymal invasion program predominated. Inhibition of another RhoA effector, ROCK, had no impact on ES-2 spheroid formation but dramatically inhibited spheroid invasion through induction of a highly elongated morphology. Concurrent inhibition of ROCK and mDia2 blocked single cell invasion from ES-2 spheroids more effectively than inhibition of either protein alone, indicating that invasive egress of amoeboid cells from mDia2-depleted spheroids is ROCK-dependent. Our findings indicate that multiple GTPase effectors must be suppressed in order to fully block invasive egress from ovarian cancer spheroids. Furthermore, tightly regulated interplay between ROCK and mDia2 signaling pathways dictates the invasive capacities and the type of invasion program utilized by motile spheroid-derived ovarian cancer cells. As loss of the gene encoding mDia2, DRF3, has been linked to cancer progression and

An mDia2/ROCK signaling axis regulates invasive egress from epithelial ovarian cancer spheroids.

Directory of Open Access Journals (Sweden)

Krista M Pettee

Full Text Available Multi-cellular spheroids are enriched in ascites of epithelial ovarian cancer (OvCa patients. They represent an invasive and chemoresistant cellular population fundamental to metastatic dissemination. The molecular mechanisms triggering single cell invasive egress from spheroids remain enigmatic. mDia formins are Rho GTPase effectors that are key regulators of F-actin cytoskeletal dynamics. We hypothesized that mDia2-driven F-actin dynamics promote single cell invasive transitions in clinically relevant three-dimensional (3D OvCa spheroids. The current study is a dissection of the contribution of the F-actin assembly factor mDia2 formin in invasive transitions and using a clinically relevant ovarian cancer spheroid model. We show that RhoA-directed mDia2 activity is required for tight spheroid organization, and enrichment of mDia2 in the invasive cellular protrusions of collagen-embedded OVCA429 spheroids. Depleting mDia2 in ES-2 spheroids enhanced invasive dissemination of single amoeboid-shaped cells. This contrasts with spheroids treated with control siRNA, where a mesenchymal invasion program predominated. Inhibition of another RhoA effector, ROCK, had no impact on ES-2 spheroid formation but dramatically inhibited spheroid invasion through induction of a highly elongated morphology. Concurrent inhibition of ROCK and mDia2 blocked single cell invasion from ES-2 spheroids more effectively than inhibition of either protein alone, indicating that invasive egress of amoeboid cells from mDia2-depleted spheroids is ROCK-dependent. Our findings indicate that multiple GTPase effectors must be suppressed in order to fully block invasive egress from ovarian cancer spheroids. Furthermore, tightly regulated interplay between ROCK and mDia2 signaling pathways dictates the invasive capacities and the type of invasion program utilized by motile spheroid-derived ovarian cancer cells. As loss of the gene encoding mDia2, DRF3, has been linked to cancer

miR-367 promotes proliferation and invasion of hepatocellular carcinoma cells by negatively regulating PTEN

Energy Technology Data Exchange (ETDEWEB)

Meng, Xiangrui, E-mail: mengxiangruibb2008@163.com [Oncology Department, The First Affiliated Hospital of Zhengzhou University, Zhengzhou (China); Lu, Peng [Gastrointestinal Surgery Department, People' s Hospital of Zhengzhou, Zhengzhou (China); Fan, Qingxia [Oncology Department, The First Affiliated Hospital of Zhengzhou University, Zhengzhou (China)

2016-01-29

MicroRNAs play important roles in the carcinogenesis of many types of cancers by inhibiting gene expression at posttranscriptional level. However, the roles of microRNAs in hepatocellular carcinoma, are still unclear. Here, we identified that miR-367 promotes hepatocellular carcinoma (HCC) cell proliferation by negatively regulates its target gene PTEN. The expression of miR-367 and PTEN are significantly inverse correlated in 35 HCC patients. In HCC cell line, CCK-8 proliferation assay indicated that the cell proliferation was promoted by miR-367, while miR-367 inhibitor significantly inhibited the cell proliferation. Transwell assay showed that miR-367 mimics significantly promoted the migration and invasion of HCC cells, whereas miR-367 inhibitors significantly reduced cell migration and invasion. Luciferase assays confirmed that miR-367 directly bound to the 3'untranslated region of PTEN, and western blotting showed that miR-367 suppressed the expression of PTEN at the protein levels. This study indicated that miR-367 negatively regulates PTEN and promotes proliferation and invasion of HCC cells. Thus, miR-367 may represent a potential therapeutic target for HCC intervention. - Highlights: • miR-367 mimics promote the proliferation and invasion of HCC cells. • miR-367 inhibitors inhibit the proliferation and invasion of HCC cells. • miR-367 targets 3′UTR of PTEN in HCC cells. • miR-367 negatively regulates PTEN in HCC cells.

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

Science.gov (United States)

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.

LDL Receptor-Related Protein-1 (LRP1 Regulates Cholesterol Accumulation in Macrophages.

Directory of Open Access Journals (Sweden)

Anna P Lillis

Full Text Available Within the circulation, cholesterol is transported by lipoprotein particles and is taken up by cells when these particles associate with cellular receptors. In macrophages, excessive lipoprotein particle uptake leads to foam cell formation, which is an early event in the development of atherosclerosis. Currently, mechanisms responsible for foam cell formation are incompletely understood. To date, several macrophage receptors have been identified that contribute to the uptake of modified forms of lipoproteins leading to foam cell formation, but the in vivo contribution of the LDL receptor-related protein 1 (LRP1 to this process is not known [corrected]. To investigate the role of LRP1 in cholesterol accumulation in macrophages, we generated mice with a selective deletion of LRP1 in macrophages on an LDL receptor (LDLR-deficient background (macLRP1-/-. After feeding mice a high fat diet for 11 weeks, peritoneal macrophages isolated from Lrp+/+ mice contained significantly higher levels of total cholesterol than those from macLRP1-/- mice. Further analysis revealed that this was due to increased levels of cholesterol esters. Interestingly, macLRP1-/- mice displayed elevated plasma cholesterol and triglyceride levels resulting from accumulation of large, triglyceride-rich lipoprotein particles in the circulation. This increase did not result from an increase in hepatic VLDL biosynthesis, but rather results from a defect in catabolism of triglyceride-rich lipoprotein particles in macLRP1-/- mice. These studies reveal an important in vivo contribution of macrophage LRP1 to cholesterol homeostasis.

Radiation Therapy Induces Macrophages to Suppress Immune Responses Against Pancreatic Tumors in Mice

Science.gov (United States)

Seifert, Lena; Werba, Gregor; Tiwari, Shaun; Ly, Nancy Ngoc Giao; Nguy, Susanna; Alothman, Sara; Alqunaibit, Dalia; Avanzi, Antonina; Daley, Donnele; Barilla, Rocky; Tippens, Daniel; Torres-Hernandez, Alejandro; Hundeyin, Mautin; Mani, Vishnu R.; Hajdu, Cristina; Pellicciotta, Ilenia; Oh, Philmo; Du, Kevin; Miller, George

2016-01-01

Background & Aims The role of radiation therapy in the treatment of patients with pancreatic ductal adenocarcinoma (PDA) is controversial. Randomized controlled trials investigating the efficacy of radiation therapy in patients with locally advanced unresectable PDA have reported mixed results, with effects ranging from modest benefit to worse outcome, compared with control therapies. We investigated whether radiation causes inflammatory cells to acquire an immune-suppressive phenotype that limits the therapeutic effects of radiation on invasive PDAs and accelerates progression of pre-invasive foci. Methods We investigated the effects of radiation in p48Cre;LSL-KrasG12D (KC) and p48Cre;LSLKrasG12D;LSL-Trp53R172H (KPC) mice, as well as in C57BL/6 mice with orthotopic tumors grown from FC1242 cells derived from KPC mice. Some mice were given neutralizing antibodies against macrophage colony stimulating factor 1 (CSF1 or MCSF) or F4/80. Pancreata were exposed to doses of radiation ranging from 2–12 Gy and analyzed by flow cytometry. Results Pancreata of KC mice exposed to radiation had a higher frequency of advanced pancreatic intraepithelial lesions and more foci of invasive cancer than pancreata of unexposed mice (controls); radiation reduced survival time by more than 6 months. A greater proportion of macrophages from invasive and pre-invasive pancreatic tumors had an immune-suppressive, M2-like phenotype, compared with control mice. Pancreata from mice exposed to radiation had fewer CD8+ T cells than controls and greater numbers of CD4+ T cells of T-helper 2 and T-regulatory cell phenotypes. Adoptive transfer of T cells from irradiated PDA to tumors of control mice accelerated tumor growth. Radiation induced production of MCSF by PDA cells. An antibody against MCSF prevented radiation from altering the phenotype of macrophages in tumors, increasing the anti-tumor T-cell response and slowing tumor growth. Conclusions Radiation exposure causes macrophages in PDAs

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

International Nuclear Information System (INIS)

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

2013-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

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

2013-11-15

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

Selective phosphorylation during early macrophage differentiation

KAUST Repository

Zhang, Huoming

2015-08-26

The differentiation of macrophages from monocytes is a tightly controlled and complex biological process. Although numerous studies have been conducted using biochemical approaches or global gene/gene profiling, the mechanisms of the early stages of differentiation remain unclear. Here we used SILAC-based quantitative proteomics approach to perform temporal phosphoproteome profiling of early macrophage differentiation. We identified a large set of phosphoproteins and grouped them as PMA-regulated and non-regulated phosphoproteins in the early stages of differentiation. Further analysis of the PMA-regulated phosphoproteins revealed that transcriptional suppression, cytoskeletal reorganization and cell adhesion were among the most significantly activated pathways. Some key involved regulators of these pathways are mTOR, MYB, STAT1 and CTNNB. Moreover, we were able to classify the roles and activities of several transcriptional factors during different differentiation stages and found that E2F is likely to be an important regulator during the relatively late stages of differentiation. This study provides the first comprehensive picture of the dynamic phosphoproteome during myeloid cells differentiation, and identifies potential molecular targets in leukemic cells.

Activating transcription factor 4 underlies the pathogenesis of arsenic trioxide-mediated impairment of macrophage innate immune functions

Energy Technology Data Exchange (ETDEWEB)

Srivastava, Ritesh K.; Li, Changzhao [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Wang, Yong [Department of Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Weng, Zhiping; Elmets, Craig A. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Harrod, Kevin S. [Department of Anesthesiology and Perioperative Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Deshane, Jessy S., E-mail: treena@uab.edu [Department of Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Athar, Mohammad, E-mail: mathar@uab.edu [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States)

2016-10-01

Chronic arsenic exposure to humans is considered immunosuppressive with augmented susceptibility to several infectious diseases. The exact molecular mechanisms, however, remain unknown. Earlier, we showed the involvement of unfolded protein response (UPR) signaling in arsenic-mediated impairment of macrophage functions. Here, we show that activating transcription factor 4 (ATF4), a UPR transcription factor, regulates arsenic trioxide (ATO)-mediated dysregulation of macrophage functions. In ATO-treated ATF4{sup +/+} wild-type mice, a significant down-regulation of CD11b expression was associated with the reduced phagocytic functions of peritoneal and lung macrophages. This severe immuno-toxicity phenotype was not observed in ATO-treated ATF4{sup +/−} heterozygous mice. To confirm these observations, we demonstrated in Raw 264.7 cells that ATF4 knock-down rescues ATO-mediated impairment of macrophage functions including cytokine production, bacterial engulfment and clearance of engulfed bacteria. Sustained activation of ATF4 by ATO in macrophages induces apoptosis, while diminution of ATF4 expression protects against ATO-induced apoptotic cell death. Raw 264.7 cells treated with ATO also manifest dysregulated Ca{sup ++} homeostasis. ATO induces Ca{sup ++}-dependent calpain-1 and caspase-12 expression which together regulated macrophage apoptosis. Additionally, apoptosis was also induced by mitochondria-regulated pathway. Restoring ATO-impaired Ca{sup ++} homeostasis in ER/mitochondria by treatments with the inhibitors of inositol 1,4,5-trisphosphate receptor (IP3R) and voltage-dependent anion channel (VDAC) attenuate innate immune functions of macrophages. These studies identify a novel role for ATF4 in underlying pathogenesis of macrophage dysregulation and immuno-toxicity of arsenic. - Highlights: • ATF4 regulates arsenic-mediated impairment in macrophage functions. • Arsenic-mediated alterations in pulmonary macrophage are diminished in ATF4{sup +/−} mice

Activating transcription factor 4 underlies the pathogenesis of arsenic trioxide-mediated impairment of macrophage innate immune functions

International Nuclear Information System (INIS)

Srivastava, Ritesh K.; Li, Changzhao; Wang, Yong; Weng, Zhiping; Elmets, Craig A.; Harrod, Kevin S.; Deshane, Jessy S.; Athar, Mohammad

2016-01-01

Chronic arsenic exposure to humans is considered immunosuppressive with augmented susceptibility to several infectious diseases. The exact molecular mechanisms, however, remain unknown. Earlier, we showed the involvement of unfolded protein response (UPR) signaling in arsenic-mediated impairment of macrophage functions. Here, we show that activating transcription factor 4 (ATF4), a UPR transcription factor, regulates arsenic trioxide (ATO)-mediated dysregulation of macrophage functions. In ATO-treated ATF4 +/+ wild-type mice, a significant down-regulation of CD11b expression was associated with the reduced phagocytic functions of peritoneal and lung macrophages. This severe immuno-toxicity phenotype was not observed in ATO-treated ATF4 +/− heterozygous mice. To confirm these observations, we demonstrated in Raw 264.7 cells that ATF4 knock-down rescues ATO-mediated impairment of macrophage functions including cytokine production, bacterial engulfment and clearance of engulfed bacteria. Sustained activation of ATF4 by ATO in macrophages induces apoptosis, while diminution of ATF4 expression protects against ATO-induced apoptotic cell death. Raw 264.7 cells treated with ATO also manifest dysregulated Ca ++ homeostasis. ATO induces Ca ++ -dependent calpain-1 and caspase-12 expression which together regulated macrophage apoptosis. Additionally, apoptosis was also induced by mitochondria-regulated pathway. Restoring ATO-impaired Ca ++ homeostasis in ER/mitochondria by treatments with the inhibitors of inositol 1,4,5-trisphosphate receptor (IP3R) and voltage-dependent anion channel (VDAC) attenuate innate immune functions of macrophages. These studies identify a novel role for ATF4 in underlying pathogenesis of macrophage dysregulation and immuno-toxicity of arsenic. - Highlights: • ATF4 regulates arsenic-mediated impairment in macrophage functions. • Arsenic-mediated alterations in pulmonary macrophage are diminished in ATF4 +/− mice. • Changes in macrophage

Dysregulation of Macrophage Activation Profiles by Engineered Nanoparticles

Energy Technology Data Exchange (ETDEWEB)

Kodali, Vamsi; Littke, Matthew H.; Tilton, Susan C.; Teeguarden, Justin G.; Shi, Liang; Frevert, Charles W.; Wang, Wei; Pounds, Joel G.; Thrall, Brian D.

2013-08-27

Although the potential human health impacts from exposure to engineered nanoparticles (ENPs) are uncertain, past epidemiological studies have established correlations between exposure to ambient air pollution particulates and the incidence of pneumonia and lung infections. Using amorphous silica and superparamagnetic iron oxide (SPIO) as model high production volume ENPs, we examined how macrophage activation by bacterial lipopolysaccharide (LPS) or the lung pathogen Streptococcus pneumoniae is altered by ENP pretreatment. Neither silica nor SPIO treatment elicited direct cytotoxic or pro-inflammatory effects in bone marrow-derived macrophages. However, pretreatment of macrophages with SPIO caused extensive reprogramming of nearly 500 genes regulated in response to LPS challenge, hallmarked by exaggerated activation of oxidative stress response pathways and suppressed activation of both pro- and anti-inflammatory pathways. Silica pretreatment altered regulation of only 67 genes, but there was strong correlation with gene sets affected by SPIO. Macrophages exposed to SPIO displayed a phenotype suggesting an impaired ability to transition from an M1 to M2-like activation state, characterized by suppressed IL-10 induction, enhanced TNFα production, and diminished phagocytic activity toward S. pneumoniae. Studies in macrophages deficient in scavenger receptor A (SR-A) showed SR-A participates in cell uptake of both the ENPs and S. pneumonia and co-regulates the anti-inflammatory IL-10 pathway. Thus, mechanisms for dysregulation of innate immunity exist by virtue that common receptor recognition pathways are used by some ENPs and pathogenic bacteria, although the extent of transcriptional reprogramming of macrophage function depends on the physicochemical properties of the ENP after internalization. Our results also illustrate that biological effects of ENPs may be indirectly manifested only after challenging normal cell function. Finally, nanotoxicology screening

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

International Nuclear Information System (INIS)

Kaye, P.M.; Feldmann, M.

1986-01-01

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

Engineering mechanical microenvironment of macrophage and its biomedical applications.

Science.gov (United States)

Li, Jing; Li, Yuhui; Gao, Bin; Qin, Chuanguang; He, Yining; Xu, Feng; Yang, Hui; Lin, Min

2018-03-01

Macrophages are the most plastic cells in the hematopoietic system and can be widely found in almost all tissues. Recently studies have shown that mechanical cues (e.g., matrix stiffness and stress/strain) can significantly affect macrophage behaviors. Although existing reviews on the physical and mechanical cues that regulate the macrophage's phenotype are available, engineering mechanical microenvironment of macrophages in vitro as well as a comprehensive overview and prospects for their biomedical applications (e.g., tissue engineering and immunotherapy) has yet to be summarized. Thus, this review provides an overview on the existing methods for engineering mechanical microenvironment of macrophages in vitro and then a section on their biomedical applications and further perspectives are presented.

Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

International Nuclear Information System (INIS)

Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C.; Ballestas, Mary E.; Elmets, Craig A.; Robbins, David J.; Matalon, Sadis; Deshane, Jessy S.; Afaq, Farrukh; Bickers, David R.; Athar, Mohammad

2013-01-01

Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions

Unfolded protein response (UPR) signaling regulates arsenic trioxide-mediated macrophage innate immune function disruption

Energy Technology Data Exchange (ETDEWEB)

Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Ballestas, Mary E. [Department of Pediatrics Infectious Disease, Children' s of Alabama, School of Medicine, University of Alabama at Birmingham, AL (United States); Elmets, Craig A. [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Robbins, David J. [Department of Surgery, Molecular Oncology Program, Miller School of Medicine, University of Miami, Miami (United States); Matalon, Sadis [Department of Anesthesiology, University of Alabama at Birmingham, Birmingham, AL (United States); Deshane, Jessy S. [Department of Medicine, Division of Pulmonary, Allergy and Critical Care Medicine, University of Alabama at Birmingham, Birmingham, AL (United States); Afaq, Farrukh [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States); Bickers, David R. [Department of Dermatology, Columbia University Medical Center, New York (United States); Athar, Mohammad, E-mail: mathar@uab.edu [Department of Dermatology and Skin Diseases Research Center, University of Alabama at Birmingham, Birmingham, AL (United States)

2013-11-01

Arsenic exposure is known to disrupt innate immune functions in humans and in experimental animals. In this study, we provide a mechanism by which arsenic trioxide (ATO) disrupts macrophage functions. ATO treatment of murine macrophage cells diminished internalization of FITC-labeled latex beads, impaired clearance of phagocytosed fluorescent bacteria and reduced secretion of pro-inflammatory cytokines. These impairments in macrophage functions are associated with ATO-induced unfolded protein response (UPR) signaling pathway characterized by the enhancement in proteins such as GRP78, p-PERK, p-eIF2α, ATF4 and CHOP. The expression of these proteins is altered both at transcriptional and translational levels. Pretreatment with chemical chaperon, 4-phenylbutyric acid (PBA) attenuated the ATO-induced activation in UPR signaling and afforded protection against ATO-induced disruption of macrophage functions. This treatment also reduced ATO-mediated reactive oxygen species (ROS) generation. Interestingly, treatment with antioxidant N-acetylcysteine (NAC) prior to ATO exposure, not only reduced ROS production and UPR signaling but also improved macrophage functions. These data demonstrate that UPR signaling and ROS generation are interdependent and are involved in the arsenic-induced pathobiology of macrophage. These data also provide a novel strategy to block the ATO-dependent impairment in innate immune responses. - Highlights: • Inorganic arsenic to humans and experimental animals disrupt innate immune responses. • The mechanism underlying arsenic impaired macrophage functions involves UPR signaling. • Chemical chaperon attenuates arsenic-mediated macrophage function impairment. • Antioxidant, NAC blocks impairment in arsenic-treated macrophage functions.

L-Plastin promotes podosome longevity and supports macrophage motility

Science.gov (United States)

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

2016-01-01

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

Cell cycle-dependent Rho GTPase activity dynamically regulates cancer cell motility and invasion in vivo.

Science.gov (United States)

Kagawa, Yoshinori; Matsumoto, Shinji; Kamioka, Yuji; Mimori, Koshi; Naito, Yoko; Ishii, Taeko; Okuzaki, Daisuke; Nishida, Naohiro; Maeda, Sakae; Naito, Atsushi; Kikuta, Junichi; Nishikawa, Keizo; Nishimura, Junichi; Haraguchi, Naotsugu; Takemasa, Ichiro; Mizushima, Tsunekazu; Ikeda, Masataka; Yamamoto, Hirofumi; Sekimoto, Mitsugu; Ishii, Hideshi; Doki, Yuichiro; Matsuda, Michiyuki; Kikuchi, Akira; Mori, Masaki; Ishii, Masaru

2013-01-01

The mechanism behind the spatiotemporal control of cancer cell dynamics and its possible association with cell proliferation has not been well established. By exploiting the intravital imaging technique, we found that cancer cell motility and invasive properties were closely associated with the cell cycle. In vivo inoculation of human colon cancer cells bearing fluorescence ubiquitination-based cell cycle indicator (Fucci) demonstrated an unexpected phenomenon: S/G2/M cells were more motile and invasive than G1 cells. Microarray analyses showed that Arhgap11a, an uncharacterized Rho GTPase-activating protein (RhoGAP), was expressed in a cell-cycle-dependent fashion. Expression of ARHGAP11A in cancer cells suppressed RhoA-dependent mechanisms, such as stress fiber formation and focal adhesion, which made the cells more prone to migrate. We also demonstrated that RhoA suppression by ARHGAP11A induced augmentation of relative Rac1 activity, leading to an increase in the invasive properties. RNAi-based inhibition of Arhgap11a reduced the invasion and in vivo expansion of cancers. Additionally, analysis of human specimens showed the significant up-regulation of Arhgap11a in colon cancers, which was correlated with clinical invasion status. The present study suggests that ARHGAP11A, a cell cycle-dependent RhoGAP, is a critical regulator of cancer cell mobility and is thus a promising therapeutic target in invasive cancers.

Cell cycle-dependent Rho GTPase activity dynamically regulates cancer cell motility and invasion in vivo.

Directory of Open Access Journals (Sweden)

Yoshinori Kagawa

Full Text Available The mechanism behind the spatiotemporal control of cancer cell dynamics and its possible association with cell proliferation has not been well established. By exploiting the intravital imaging technique, we found that cancer cell motility and invasive properties were closely associated with the cell cycle. In vivo inoculation of human colon cancer cells bearing fluorescence ubiquitination-based cell cycle indicator (Fucci demonstrated an unexpected phenomenon: S/G2/M cells were more motile and invasive than G1 cells. Microarray analyses showed that Arhgap11a, an uncharacterized Rho GTPase-activating protein (RhoGAP, was expressed in a cell-cycle-dependent fashion. Expression of ARHGAP11A in cancer cells suppressed RhoA-dependent mechanisms, such as stress fiber formation and focal adhesion, which made the cells more prone to migrate. We also demonstrated that RhoA suppression by ARHGAP11A induced augmentation of relative Rac1 activity, leading to an increase in the invasive properties. RNAi-based inhibition of Arhgap11a reduced the invasion and in vivo expansion of cancers. Additionally, analysis of human specimens showed the significant up-regulation of Arhgap11a in colon cancers, which was correlated with clinical invasion status. The present study suggests that ARHGAP11A, a cell cycle-dependent RhoGAP, is a critical regulator of cancer cell mobility and is thus a promising therapeutic target in invasive cancers.

Punicalagin, a PTP1B inhibitor, induces M2c phenotype polarization via up-regulation of HO-1 in murine macrophages.

Science.gov (United States)

Xu, Xiaolong; Guo, Yuhong; Zhao, Jingxia; He, Shasha; Wang, Yan; Lin, Yan; Wang, Ning; Liu, Qingquan

2017-09-01

Current data have shown that punicalagin (PUN), an ellagitannin isolated from pomegranate, possesses anti-inflammatory and anti-oxidant properties; however, its direct targets have not yet been reported. This is the first report that PTP1B serves as a direct target of PUN, with IC 50 value of 1.04μM. Results from NPOI further showed that the K on and K off of PUN-PTP1B complex were 3.38e2M -1 s -1 and 4.13e-3s -1 , respectively. The active site Arg24 of PTP1B was identified as a key binding site of PUN by computation simulation and point mutation. Moreover, inhibition of PTP1B by PUN promoted an M2c-like macrophage polarization and enhanced anti-inflammatory cytokines expression, including IL-10 and M-CSF. Based on gene expression profile, we elucidated that PUN treatment significantly up-regulated 275 genes and down-regulated 1059 genes. M1-like macrophage marker genes, such as Tlr4, Irf1/2, Hmgb1, and Stat1 were down-regulated, while M2 marker genes, including Tmem171, Gpr35, Csf1, Il1rn, Cebpb, Fos, Vegfα, Slc11a1, and Bhlhe40 were up-regulated in PUN-treated macrophages. Hmox-1, a gene encoding HO-1 protein, was preferentially expressed with 16-fold change. Inhibition of HO-1 obviously restored PUN-induced M2 polarization and IL-10 secretion. In addition, phosphorylation of both Akt and STAT3 contributed to PUN-induced HO-1 expression. This study provided new insights into the mechanisms of PUN-mediated anti-inflammatory and anti-oxidant activities and provided new therapeutic strategies for inflammatory diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Co-ordinate regulation of Salmonella typhimurium invasion genes by environmental and regulatory factors is mediated by control of hilA expression.

Science.gov (United States)

Bajaj, V; Lucas, R L; Hwang, C; Lee, C A

1996-11-01

During infection of their hosts, salmonellae enter intestinal epithelial cells. It has been proposed that when Salmonella typhimurium is present in the intestinal lumen, several environmental and regulatory conditions modulate the expression of invasion factors required for bacterial entry into host cells. We report here that the expression of six different S. typhimurium invasion genes encoded on SPI1 (Salmonella pathogenicity island 1) is co-ordinately regulated by oxygen, osmolarity, pH, PhoPQ, and HilA. HilA is a transcriptional activator of the OmpR/ToxR family that is also encoded on SPI1. We have found that HilA plays a central role in the co-ordinated regulation of invasion genes by environmental and regulatory conditions. HilA can activate the expression of two invasion gene-lacZY fusions on reporter plasmids in Escherichia coll, suggesting that HilA acts directly at invasion-gene promoters in S. typhimurium. We have found that the regulation of invasion genes by oxygen, osmolarity, pH, and PhoPQ is indirect and is mediated by regulation of hilA expression by these environmental and regulatory factors. We hypothesize that the complex and co-ordinate regulation of Invasion genes by HilA is an important feature of salmonella pathogenesis and allows salmonellae to enter intestinal epithelial cells.

Neuron-macrophage crosstalk in the intestine: a ‘microglia’ perspective

Directory of Open Access Journals (Sweden)

Simon eVerheijden

2015-10-01

Full Text Available Intestinal macrophages are strategically located in different layers of the intestine, including the mucosa, submucosa and muscularis externa, where they perform complex tasks to maintain intestinal homeostasis. As the gastrointestinal tract is continuously challenged by foreign antigens, macrophage activation should be tightly controlled to prevent chronic inflammation and tissue damage. Unraveling the precise cellular and molecular mechanisms underlying the tissue-specific control of macrophage activation is crucial to get more insight into intestinal immune regulation. Two recent reports provide unanticipated evidence that the enteric nervous system acts as a critical regulator of macrophage function in the myenteric plexus. Both studies clearly illustrate that enteric neurons reciprocally interact with intestinal macrophages and are actively involved in shaping their phenotype. This concept has striking parallels with the central nervous system (CNS, where neuronal signals maintain microglia, the resident macrophages of the CNS, in a quiescent, anti-inflammatory state. This inevitably evokes the perception that the ENS and CNS share mechanisms of neuroimmune interaction. In line, intestinal macrophages, both in the muscularis externa and (submucosa, express high levels of CX3CR1, a feature that was once believed to be unique for microglia. CX3CR1 is the sole receptor of fractalkine (CX3CL1, a factor mainly produced by neurons in the CNS to facilitate neuron-microglia communication. The striking parallels between resident macrophages of the brain and intestine might provide a promising new line of thought to get more insight into cellular and molecular mechanisms controlling macrophage activation in the gut.

Regulation and control of nitric oxide (NO) in macrophages

DEFF Research Database (Denmark)

Kovacevic, Zaklina; Sahni, Sumit; Lok, K.H.

2017-01-01

We recently demonstrated that a novel storage and transport mechanism for nitric oxide (NO) mediated by glutathione-S-transferase P1 (GSTP1) and multidrug resistance protein 1 (MRP1/ABCC1), protects M1-macrophage (M1-MØ) models from large quantities of endogenous NO. This system stores and transp......We recently demonstrated that a novel storage and transport mechanism for nitric oxide (NO) mediated by glutathione-S-transferase P1 (GSTP1) and multidrug resistance protein 1 (MRP1/ABCC1), protects M1-macrophage (M1-MØ) models from large quantities of endogenous NO. This system stores...... be responsible for delivering cytotoxic NO as DNICs via MRP1 from M1-MØs, to tumor cell targets....

Interaction with extracellular matrix proteins influences Lsh/Ity/Bcg (candidate Nramp) gene regulation of macrophage priming/activation for tumour necrosis factor-alpha and nitrite release.

Science.gov (United States)

Formica, S; Roach, T I; Blackwell, J M

1994-05-01

The murine resistance gene Lsh/Ity/Bcg regulates activation of macrophages for tumour necrosis factor-alpha (TNF-alpha)-dependent production of nitric oxide mediating antimicrobial activity against Leishmania, Salmonella and Mycobacterium. As Lsh is differentially expressed in macrophages from different tissue sites, experiments were performed to determine whether interaction with extracellular matrix (ECM) proteins would influence the macrophage TNF-alpha response. Plating of bone marrow-derived macrophages onto purified fibrinogen or fibronectin-rich L929 cell-derived matrices, but not onto mannan, was itself sufficient to stimulate TNF-alpha release, with significantly higher levels released from congenic B10.L-Lshr compared to C57BL/10ScSn (Lshs) macrophages. Only macrophages plated onto fibrinogen also released measurable levels of nitrites, again higher in Lshr compared to Lshs macrophages. Addition of interferon-gamma (IFN-gamma), but not bacterial lipopolysaccharide or mycobacterial lipoarabinomannan, as a second signal enhanced the TNF-alpha and nitrite responses of macrophages plated onto fibrinogen, particularly in the Lshr macrophages. Interaction with fibrinogen and fibronectin also primed macrophages for an enhanced TNF-alpha response to leishmanial parasites, but this was only translated into enhanced nitrite responses in the presence of IFN-gamma. In these experiments, Lshr macrophages remained superior in their TNF-alpha responses throughout, but to a degree which reflected the magnitude of the difference observed on ECM alone. Hence, the specificity for the enhanced TNF-alpha responses of Lshr macrophages lay in their interaction with fibrinogen and fibronectin ECM, while a differential nitrite response was only observed with fibrinogen and/or IFN-gamma. The results are discussed in relation to the possible function of the recently cloned candidate gene Nramp, which has structural identity to eukaryote transporters and an N-terminal cytoplasmic

Shigella mediated depletion of macrophages in a murine breast cancer model is associated with tumor regression.

Directory of Open Access Journals (Sweden)

Katharina Galmbacher

Full Text Available A tumor promoting role of macrophages has been described for a transgenic murine breast cancer model. In this model tumor-associated macrophages (TAMs represent a major component of the leukocytic infiltrate and are associated with tumor progression. Shigella flexneri is a bacterial pathogen known to specificly induce apotosis in macrophages. To evaluate whether Shigella-induced removal of macrophages may be sufficient for achieving tumor regression we have developed an attenuated strain of S. flexneri (M90TDeltaaroA and infected tumor bearing mice. Two mouse models were employed, xenotransplantation of a murine breast cancer cell line and spontanous breast cancer development in MMTV-HER2 transgenic mice. Quantitative analysis of bacterial tumor targeting demonstrated that attenuated, invasive Shigella flexneri primarily infected TAMs after systemic administration. A single i.v. injection of invasive M90TDeltaaroA resulted in caspase-1 dependent apoptosis of TAMs followed by a 74% reduction in tumors of transgenic MMTV-HER-2 mice 7 days post infection. TAM depletion was sustained and associated with complete tumor regression.These data support TAMs as useful targets for antitumor therapy and highlight attenuated bacterial pathogens as potential tools.

Alternative activation of macrophages and pulmonary fibrosis are modulated by scavenger receptor, macrophage receptor with collagenous structure.

Science.gov (United States)

Murthy, Shubha; Larson-Casey, Jennifer L; Ryan, Alan J; He, Chao; Kobzik, Lester; Carter, A Brent

2015-08-01

Alternative activation of alveolar macrophages is linked to fibrosis following exposure to asbestos. The scavenger receptor, macrophage receptor with collagenous structure (MARCO), provides innate immune defense against inhaled particles and pathogens; however, a receptor for asbestos has not been identified. We hypothesized that MARCO acts as an initial signaling receptor for asbestos, polarizes macrophages to a profibrotic M2 phenotype, and is required for the development of asbestos-induced fibrosis. Compared with normal subjects, alveolar macrophages isolated from patients with asbestosis express higher amounts of MARCO and have greater profibrotic polarization. Arginase 1 (40-fold) and IL-10 (265-fold) were higher in patients. In vivo, the genetic deletion of MARCO attenuated the profibrotic environment and pulmonary fibrosis in mice exposed to chrysotile. Moreover, alveolar macrophages from MARCO(-/-) mice polarize to an M1 phenotype, whereas wild-type mice have higher Ym1 (>3.0-fold) and nearly 7-fold more active TGF-β1 in bronchoalveolar lavage (BAL) fluid (BALF). Arg(432) and Arg(434) in domain V of MARCO are required for the polarization of macrophages to a profibrotic phenotype as mutation of these residues reduced FIZZ1 expression (17-fold) compared with cells expressing MARCO. These observations demonstrate that a macrophage membrane protein regulates the fibrotic response to lung injury and suggest a novel target for therapeutic intervention. © FASEB.

Transcriptional regulator GntR of Brucella abortus regulates cytotoxicity, induces the secretion of inflammatory cytokines and affects expression of the type IV secretion system and quorum sensing system in macrophages.

Science.gov (United States)

Li, Zhiqiang; Wang, Shuli; Zhang, Hui; Zhang, Jinliang; Xi, Li; Zhang, Junbo; Chen, Chuangfu

2017-03-01

The pathogenic mechanisms of Brucella are still poorly understood. GntR is a transcriptional regulator and plays an important role in the intracellular survival of Brucella. To investigate whether GntR is involved in the cytotoxicity of Brucella abortus (B. abortus), we created a 2308ΔgntR mutant of B. abortus 2308 (S2308). Lactate dehydrogenase (LDH) cytotoxicity assays using a murine macrophage cell line (RAW 264.7) show that high-dose infection with the parental strain produces a high level of cytotoxicity to macrophages, but the 2308ΔgntR mutant exhibits a very low level of cytotoxicity, indicating that mutation of GntR impairs the cytotoxicity of B. abortus to macrophages. After the macrophages are infected with 2308ΔgntR, the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-8 (IL-8) increase and are slightly higher than that for the S2308 infected group, indicating that the 2308ΔgntR mutant could induce the secretion of inflammatory cytokines. The virulence factor detection experiments indicate that genes involved in the type IV secretion system (T4SS) and quorum sensing system (QSS) are down-regulated in 2308ΔgntR. The lower levels of survival of 2308ΔgntR under various stress conditions and the increased sensitivity of 2308ΔgntR to polymyxin B suggest that GntR is a virulence factor and that deletion of gntR reduces of B. abortus to stress conditions. Taken together, our results demonstrate that GntR is involved in the cytotoxicity, virulence and intracellular survival of B. abortus during its infection.

Spliced XBP1 promotes macrophage survival and autophagy by interacting with Beclin-1

Energy Technology Data Exchange (ETDEWEB)

Tian, Ping-Ge [Southern Medical University, Guangzhou, Guangdong 510515 (China); Jiang, Zhi-Xin [Centre Laboratory, The 305th Hospital of the People' s Liberation Army, Beijing 100017 (China); Li, Jian-Hua [Department of Geriatric Cardiology, Chinese PLA General Hosptial, Beijing 100853 (China); Zhou, Zhe, E-mail: zhouzhe76@126.com [Laboratory of Biotechnology, Beijing Institute of Radiation Medicine, Beijing 100850 (China); Zhang, Qing-Hua, E-mail: 1056055170@qq.com [Department of Cardiology, The 305th Hospital of the People' s Liberation Army, Beijing 100017 (China)

2015-08-07

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 the 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.

Current Concept and Update of the Macrophage Plasticity Concept: Intracellular Mechanisms of Reprogramming and M3 Macrophage “Switch” Phenotype

Science.gov (United States)

Malyshev, Igor; Malyshev, Yuri

2015-01-01

Macrophages play a key role in immunity. In this review, we consider the traditional notion of macrophage plasticity, data that do not fit into existing concepts, and a hypothesis for existence of a new switch macrophage phenotype. Depending on the microenvironment, macrophages can reprogram their phenotype toward the proinflammatory M1 phenotype or toward the anti-inflammatory M2 phenotype. Macrophage reprogramming involves well-coordinated changes in activities of signalling and posttranslational mechanisms. Macrophage reprogramming is provided by JNK-, PI3K/Akt-, Notch-, JAK/STAT-, TGF-β-, TLR/NF-κB-, and hypoxia-dependent pathways. Posttranscriptional regulation is based on micro-mRNA. We have hypothesized that, in addition to the M1 and M2 phenotypes, an M3 switch phenotype exists. This switch phenotype responds to proinflammatory stimuli with reprogramming towards the anti-inflammatory M2 phenotype or, contrarily, it responds to anti-inflammatory stimuli with reprogramming towards the proinflammatory M1 phenotype. We have found signs of such a switch phenotype in lung diseases. Understanding the mechanisms of macrophage reprogramming will assist in the selection of new therapeutic targets for correction of impaired immunity. PMID:26366410

Essential Role of DAP12 Signaling in Macrophage Programming into a Fusion-Competent State

Science.gov (United States)

Helming, Laura; Tomasello, Elena; Kyriakides, Themis R.; Martinez, Fernando O.; Takai, Toshiyuki; Gordon, Siamon; Vivier, Eric

2009-01-01

Multinucleated giant cells, formed by fusion of macrophages, are a hallmark of granulomatous inflammation. With a genetic approach, we show that signaling through the adaptor protein DAP12 (DNAX activating protein of 12 kD), its associated receptor triggering receptor expressed by myeloid cells 2 (TREM-2), and the downstream protein tyrosine kinase Syk is required for the cytokine-induced formation of giant cells and that overexpression of DAP12 potentiates macrophage fusion. We also present evidence that DAP12 is a general macrophage fusion regulator and is involved in modulating the expression of several macrophage-associated genes, including those encoding known mediators of macrophage fusion, such as DC-STAMP and Cadherin 1. Thus, DAP12 is involved in programming of macrophages through the regulation of gene and protein expression to induce a fusion-competent state. PMID:18957693

Spiromastixones Inhibit Foam Cell Formation via Regulation of Cholesterol Efflux and Uptake in RAW264.7 Macrophages

Directory of Open Access Journals (Sweden)

Chongming Wu

2015-10-01

Full Text Available Bioassay-guided evaluation shows that a deep sea-derived fungus, Spiromastix sp. MCCC 3A00308, possesses lipid-lowering activity. Chromatographic separation of a culture broth resulted in the isolation of 15 known depsidone-based analogues, labeled spiromastixones A–O (1–15. Each of these compounds was tested for its ability to inhibit oxidized low-density lipoprotein (oxLDL-induced foam cell formation in RAW264.7 macrophages. Spiromastixones 6–8 and 12–14 significantly decreased oxLDL-induced lipid over-accumulation, reduced cell surface area, and reduced intracellular cholesterol concentration. Of these compounds, spiromastixones 6 and 14 exerted the strongest inhibitory effects. Spiromastixones 6 and 14 dramatically inhibited cholesterol uptake and stimulated cholesterol efflux to apolipoprotein A1 (ApoA1 and high-density lipoprotein (HDL in RAW264.7 macrophages. Mechanistic investigation indicated that spiromastixones 6, 7, 12 and 14 significantly up-regulated the mRNA levels of ATP-binding cassette sub-family A1 (ABCA1 and down-regulated those of scavenger receptor CD36, while the transcription of ATP-binding cassette sub-family A1 (ABCG1 and proliferator-activated receptor gamma (PPARγ were selectively up-regulated by 6 and 14. A transactivation reporter assay revealed that spiromastixones 6 and 14 remarkably enhanced the transcriptional activity of PPARγ. These results suggest that spiromastixones inhibit foam cell formation through upregulation of PPARγ and ABCA1/G1 and downregulation of CD36, indicating that spiromastixones 6 and 14 are promising lead compounds for further development as anti-atherogenic agents.

Burkholderia pseudomallei transcriptional adaptation in macrophages

Directory of Open Access Journals (Sweden)

Chieng Sylvia

2012-07-01

Full Text Available Abstract Background Burkholderia pseudomallei is a facultative intracellular pathogen of phagocytic and non-phagocytic cells. How the bacterium interacts with host macrophage cells is still not well understood and is critical to appreciate the strategies used by this bacterium to survive and how intracellular survival leads to disease manifestation. Results Here we report the expression profile of intracellular B. pseudomallei following infection of human macrophage-like U937 cells. During intracellular growth over the 6 h infection period, approximately 22 % of the B. pseudomallei genome showed significant transcriptional adaptation. B. pseudomallei adapted rapidly to the intracellular environment by down-regulating numerous genes involved in metabolism, cell envelope, motility, replication, amino acid and ion transport system and regulatory function pathways. Reduced expression in catabolic and housekeeping genes suggested lower energy requirement and growth arrest during macrophage infection, while expression of genes encoding anaerobic metabolism functions were up regulated. However, whilst the type VI secretion system was up regulated, expression of many known virulence factors was not significantly modulated over the 6hours of infection. Conclusions The transcriptome profile described here provides the first comprehensive view of how B. pseudomallei survives within host cells and will help identify potential virulence factors and proteins that are important for the survival and growth of B. pseudomallei within human cells.

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

Science.gov (United States)

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

2013-02-01

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

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

Directory of Open Access Journals (Sweden)

Silvia Y Bando

2010-09-01

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

Selective Inhibitors of Kv11.1 Regulate IL-6 Expression by Macrophages in Response to TLR/IL-1R Ligands

Directory of Open Access Journals (Sweden)

Cheryl Hunter

2010-01-01

Full Text Available The mechanism by which the platelet-endothelial cell adhesion molecule PECAM-1 regulates leukodiapedesis, vascular endothelial integrity, and proinflammatory cytokine expression in vivo is not known. We recently identified PECAM-1 as a negative regulator of Kv11.1, a specific voltage-gated potassium channel that functioned in human macrophages to reset a resting membrane potential following depolarization. We demonstrate here that dofetilide (DOF, a selective inhibitor of the Kv11.1 current, had a profound inhibitory effect on neutrophil recruitment in mice following TLR/IL-1R–elicited peritonitis or intrascrotal injection of IL-1β, but had no effect on responses seen with TNFα. Furthermore, inhibitors of Kv11.1 (DOF, E4031, and astemizole, but not Kv1.3 (margatoxin, suppressed the expression of IL-6 and MCP-1 cytokines by murine resident peritoneal macrophages, while again having no effect on TNFα. In contrast, IL-6 expression by peritoneal mesothelial cells was unaffected. Using murine P388 cells, which lack endogenous C/EBPβexpression and are unresponsive to LPS for the expression of both IL-6 and MCP-1, we observed that DOF inhibited LPS-induced expression of IL-6 mRNA following ectopic expression of wild-type C/EBPβ, but not a serine-64 point mutant. Finally, DOF inhibited the constitutive activation of cdk2 in murine peritoneal macrophages; cdk2 is known to phosphorylate C/EBPβ at serine-64. Taken together, our results implicate a potential role for Kv11.1 in regulating cdk2 and C/EBPβ activity, where robust transactivation of both IL-6 and MCP-1 transcription is known to be dependent on serine-64 of C/EBPβ. Our data might also explain the altered phenotypes displayed by PECAM-1 knockout mice in several disease models.

[Role of Rac1 signaling pathway of azathioprine and peptidoglycan in the regulation of monocyte-macrophage apoptosis in Crohn's disease].

Science.gov (United States)

Zhou, Z; Jing, Y; Ran, Y; Zhao, J; Zhou, L; Wang, B M

2018-04-01

Objective: To evaluate the changes of macrophages and expression of Rac1 in the inflammatory site of Crohn's disease, and to investigate the effects of 6-thioguanine (6-TG) and peptidoglycan on apoptosis of human peripheral blood monocyte-macrophage by regulating Rac1 signaling pathway. Methods: Ten patients with Crohn's disease and eight healthy controls diagnosed were enrolled at Department of Gastroenterology and Hepatology, Tianjin Medical University General Hospital from January 2013 to January 2014. The number of macrophages, apoptosis and expression of Rac1 in the inflammation sites and non-inflammation sites of intestinal mucosa were detected in both patients and controls. Peripheral blood mononuclear cells (PBMCs) were sorted by CD 14 immunomagnetic beads. The apoptosis of monocytes, expression of Rac1 and related apoptosis signaling molecules were detected in patients treated with peptidoglycan, 6-TG and Rac1 inhibitor NSC23766 and another 15 healthy donors. Results: The number of macrophages and apoptotic cells significantly increased in the inflammatory group of Crohn's disease patients compared with the non-inflammatory group. The expression of PAK1, downstream molecular of Rac1 signaling pathway of macrophages was also significantly higher in the inflammatory group of Crohn's disease patients than that in healthy controls and non-inflammatory group. Compared with control group, anti-apoptotic signals (NF-κB, Bcl-xL and STAT-3) in PBMCs increased in the peptidoglycan group, while slightly decreased in 6-TG group. 6-TG and NSC23766 significantly promoted peptidoglycan-related anti-apoptosis [peptidoglycan group (8.6±3.7)%, peptidoglycan+ 6-TG group (42.0±2.7)%, peptidoglycan+ NSC23766 group (58.5±6.9)%, PRac1 signaling pathway leading to macrophage apoptosis.

Decreased inducibility of TNF expression in lipid-loaded macrophages

Directory of Open Access Journals (Sweden)

Kallin Bengt

2002-10-01

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

Tumor-Associated Macrophages in Oncolytic Virotherapy: Friend or Foe?

Directory of Open Access Journals (Sweden)

Nicholas L. Denton

2016-07-01

Full Text Available Cancer therapy remains a challenge due to toxicity limitations of chemotherapy and radiation therapy. Oncolytic viruses that selectively replicate and destroy cancer cells are of increasing interest. In addition to direct cell lysis, these vectors stimulate an anti-tumor immune response. A key regulator of tumor immunity is the tumor-associated macrophage population. Macrophages can either support oncolytic virus therapy through pro-inflammatory stimulation of the anti-tumor response at the cost of hindering direct oncolysis or through immunosuppressive protection of virus replication at the cost of hindering the anti-tumor immune response. Despite similarities in macrophage interaction between adult and pediatric tumors and the abundance of research supporting macrophage modulation in adult tumors, there are few studies investigating macrophage modulation in pediatric cancers or modulation of immunotherapy. We review the current state of knowledge regarding macrophages in cancers and their influence on oncolytic virotherapy.

MicroRNA-24 Modulates Staphylococcus aureus-Induced Macrophage Polarization by Suppressing CHI3L1.

Science.gov (United States)

Jingjing, Zhang; Nan, Zhang; Wei, Wu; Qinghe, Guo; Weijuan, Wang; Peng, Wang; Xiangpeng, Wang

2017-06-01

Macrophages play a crucial role in host innate anti-Staphylococcus aureus defense, which is tightly regulated by multiple factors, including microRNAs. A recent study showed that miR-24 plays an important role in macrophage polarization. Here, we investigated the biological function of miR-24 in S. aureus-stimulated macrophages. The results revealed that miR-24 expression was significantly decreased in both human and mouse macrophage cell lines with S. aureus stimulation in a time-dependent manner. Moreover, miR-24 overexpression significantly decreased the production of M1 phenotype markers, such as IL-6, iNOS, TNF-α, CD86, and CD80, whereas it increased the production of M2 markers, such as Arg1, CCL17, CCL22, CD163, and CD206, in S. aureus-stimulated macrophages. Conversely, knockdown of miR-24 promoted M1 macrophage polarization but diminished M2 macrophage polarization in S. aureus-stimulated macrophages. Furthermore, CHI3L1 was predicted as a target gene of miR-24 using bioinformatics software and identified by luciferase reporter assay. Additionally, miR-24 overexpression inhibited CHI3L1 expression and downregulated the downstream MAPK pathway in S. aureus-stimulated macrophages. Finally, CHI3L1 overexpression rescued macrophage polarization and MAPK pathway inhibition induced by miR-24 mimic transfection in S. aureus-stimulated macrophages. In conclusion, the data suggest that miR-24 serves as a molecular regulator in S. aureus-induced macrophage polarization through targeting of CHI3L1 and regulation of the MAPK pathway, which may provide a promising therapeutic target for S. aureus-related infections and inflammatory diseases.

The Role of Macrophage Lipophagy in Reverse Cholesterol Transport

Directory of Open Access Journals (Sweden)

Se-Jin Jeong

2017-03-01

Full Text Available Macrophage cholesterol efflux is a central step in reverse cholesterol transport, which helps to maintain cholesterol homeostasis and to reduce atherosclerosis. Lipophagy has recently been identified as a new step in cholesterol ester hydrolysis that regulates cholesterol efflux, since it mobilizes cholesterol from lipid droplets of macrophages via autophagy and lysosomes. In this review, we briefly discuss recent advances regarding the mechanisms of the cholesterol efflux pathway in macrophage foam cells, and present lipophagy as a therapeutic target in the treatment of atherosclerosis.

Revisiting mouse peritoneal macrophages: heterogeneity, development and function

Directory of Open Access Journals (Sweden)

Alexandra Dos Anjos Cassado

2015-05-01

Full Text Available Tissue macrophages play a crucial role in the maintenance of tissue homeostasis and also contribute to inflammatory and reparatory responses during pathogenic infection and tissue injury. The high heterogeneity of these macrophages is consistent with their adaptation to distinct tissue environments and specialization to develop niche-specific functions. Although peritoneal macrophages are one of best-studied macrophage populations, only recently it was demonstrated the co-existence of two subsets in mouse PerC, which exhibit distinct phenotypes, functions and origins. These macrophage subsets have been classified according to their morphology as LPMs (large peritoneal macrophages and SPMs (small peritoneal macrophages. LPMs, the most abundant subset under steady-state conditions, express high levels of F4/80 and low levels of class II molecules of the major histocompatibility complex (MHC. LPMs appear to be originated from embriogenic precursors, and their maintenance in PerC is regulated by expression of specific transcription factors and tissue-derived signals. Conversely, SPMs, a minor subset in unstimulated PerC, have a F4/80lowMHC-IIhigh phenotype and are generated from bone-marrow-derived myeloid precursors. In response to infectious or inflammatory stimuli, the cellular composition of PerC is dramatically altered, where LPMs disappear and SPMs become the prevalent population together with their precursor, the inflammatory monocyte. SPMs appear to be the major source of inflammatory mediators in PerC during infection whereas LPMs contribute for gut-associated lymphoid tissue (GALT-independent and retinoic acid-dependent IgA production by peritoneal B-1 cells. In the last years, considerable efforts have been made to broaden our understanding of LPM and SPM origin, transcriptional regulation and functional profile. This review addresses these issues, focusing on the impact of tissue-derived signals and external stimulation in the complex

Unique proteomic signatures distinguish macrophages and dendritic cells.

Directory of Open Access Journals (Sweden)

Lev Becker

Full Text Available Monocytes differentiate into heterogeneous populations of tissue macrophages and dendritic cells (DCs that regulate inflammation and immunity. Identifying specific populations of myeloid cells in vivo is problematic, however, because only a limited number of proteins have been used to assign cellular phenotype. Using mass spectrometry and bone marrow-derived cells, we provided a global view of the proteomes of M-CSF-derived macrophages, classically and alternatively activated macrophages, and GM-CSF-derived DCs. Remarkably, the expression levels of half the plasma membrane proteins differed significantly in the various populations of cells derived in vitro. Moreover, the membrane proteomes of macrophages and DCs were more distinct than those of classically and alternatively activated macrophages. Hierarchical cluster and dual statistical analyses demonstrated that each cell type exhibited a robust proteomic signature that was unique. To interrogate the phenotype of myeloid cells in vivo, we subjected elicited peritoneal macrophages harvested from wild-type and GM-CSF-deficient mice to mass spectrometric and functional analysis. Unexpectedly, we found that peritoneal macrophages exhibited many features of the DCs generated in vitro. These findings demonstrate that global analysis of the membrane proteome can help define immune cell phenotypes in vivo.

Growth hormone (GH) differentially regulates NF-kB activity in preadipocytes and macrophages: implications for GH's role in adipose tissue homeostasis in obesity.

Science.gov (United States)

Kumar, P Anil; Chitra, P Swathi; Lu, Chunxia; Sobhanaditya, J; Menon, Ram

2014-06-01

Adipose tissue remodeling in obesity involves macrophage infiltration and chronic inflammation. NF-kB-mediated chronic inflammation of the adipose tissue is directly implicated in obesity-associated insulin resistance. We have investigated the effect of growth hormone (GH) on NF-kB activity in preadipocytes (3T3-F442A) and macrophages (J774A.1). Our studies indicate that whereas GH increases NF-kB activity in preadipocytes, it decreases NF-kB activity in macrophages. This differential response of NF-kB activity to GH correlates with the GH-dependent expression of a cadre of NF-kB-activated cytokines in these two cell types. Activation of NF-kB by GH in preadipocytes heightens inflammatory response by stimulating production of multiple cytokines including TNF-α, IL-6, and MCP-1, the mediators of both local and systemic insulin resistance and chemokines that recruit macrophages. Our studies also suggest differential regulation of miR132 and SIRT1 expression as a mechanism underlying the observed variance in GH-dependent NF-kB activity and altered cytokine profile in preadipocytes and macrophages. These findings further our understanding of the complex actions of GH on adipocytes and insulin sensitivity.

Localization and counting of CD68-labelled macrophages in placentas of normal and preeclamptic women

Science.gov (United States)

Al-khafaji, Lina Ali; Al-Yawer, Malak A.

2017-09-01

In the human placenta, there are two types of placental macrophages Hofbauer cells of fetal villi and decidual macrophages of maternal decidua basalis. Placental macrophages adopt a specialized phenotype that may hold a key role in synthesis of vital mediators involved in the establishment and maintenance of pregnancy, parturition and maternal-fetal tolerance. Aberrant behavior of these macrophages can affect trophoblast functions and placental development and potentially can lead to a spectrum of adverse pregnancy outcomes. Yet, the populations and functions of placental macrophages in women with different parity and women with preeclampsia remain ill-defined and subject of controversy. Immuno-histochemical study using CD68 primary antibody revealed a significant increase in number of CD68 positive fetal and decidual macrophages in preeclamptic subgroups as compared to controls. Fetal macrophages were seen to be localized near fetal vessel wall and near syncytium which were significantly increased in primiparous preeclamptic subgroup. Our study assumed that there may be intermingling of signals between macrophages and trophoblast cells resulting in impairment of trophoblast invasion and spiral artery remodeling which is the primary placental defect in pregnancies complicated by preeclampsia.

Endoglin-mediated suppression of prostate cancer invasion is regulated by activin and bone morphogenetic protein type II receptors.

Directory of Open Access Journals (Sweden)

Michael J Breen

Full Text Available Mortality from prostate cancer (PCa is due to the formation of metastatic disease. Understanding how that process is regulated is therefore critical. We previously demonstrated that endoglin, a type III transforming growth factor β (TGFβ superfamily receptor, suppresses human PCa cell invasion and metastasis. Endoglin-mediated suppression of invasion was also shown by us to be dependent upon the type I TGFβ receptor, activin receptor-like kinase 2 (ALK2, and the downstream effector, Smad1. In this study we demonstrate for the first time that two type II TGFβ receptors are required for endoglin-mediated suppression of invasion: activin A receptor type IIA (ActRIIA and bone morphogenetic protein receptor type II (BMPRII. Downstream signaling through these receptors is predominantly mediated by Smad1. ActRIIA stimulates Smad1 activation in a kinase-dependent manner, and this is required for suppression of invasion. In contrast BMPRII regulates Smad1 in a biphasic manner, promoting Smad1 signaling through its kinase domain but suppressing it through its cytoplasmic tail. BMPRII's Smad1-regulatory effects are dependent upon its expression level. Further, its ability to suppress invasion is independent of either kinase function or tail domain. We demonstrate that ActRIIA and BMPRII physically interact, and that each also interacts with endoglin. The current findings demonstrate that both BMPRII and ActRIIA are necessary for endoglin-mediated suppression of human PCa cell invasion, that they have differential effects on Smad1 signaling, that they make separate contributions to regulation of invasion, and that they functionally and physically interact.

Granulocyte-macrophage colony stimulatory factor enhances the pro-inflammatory response of interferon-γ-treated macrophages to Pseudomonas aeruginosa infection.

Directory of Open Access Journals (Sweden)

Sonali Singh

Full Text Available Pseudomonas aeruginosa is an opportunistic pathogen that can cause severe infections at compromised epithelial surfaces, such those found in burns, wounds, and in lungs damaged by mechanical ventilation or recurrent infections, particularly in cystic fibrosis (CF patients. CF patients have been proposed to have a Th2 and Th17-biased immune response suggesting that the lack of Th1 and/or over exuberant Th17 responses could contribute to the establishment of chronic P. aeruginosa infection and deterioration of lung function. Accordingly, we have observed that interferon (IFN-γ production by peripheral blood mononuclear cells from CF patients positively correlated with lung function, particularly in patients chronically infected with P. aeruginosa. In contrast, IL-17A levels tended to correlate negatively with lung function with this trend becoming significant in patients chronically infected with P. aeruginosa. These results are in agreement with IFN-γ and IL-17A playing protective and detrimental roles, respectively, in CF. In order to explore the protective effect of IFN-γ in CF, the effect of IFN-γ alone or in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF, on the ability of human macrophages to control P. aeruginosa growth, resist the cytotoxicity induced by this bacterium or promote inflammation was investigated. Treatment of macrophages with IFN-γ, in the presence and absence of GM-CSF, failed to alter bacterial growth or macrophage survival upon P. aeruginosa infection, but changed the inflammatory potential of macrophages. IFN-γ caused up-regulation of monocyte chemoattractant protein-1 (MCP-1 and TNF-α and down-regulation of IL-10 expression by infected macrophages. GM-CSF in combination with IFN-γ promoted IL-6 production and further reduction of IL-10 synthesis. Comparison of TNF-α vs. IL-10 and IL-6 vs. IL-10 ratios revealed the following hierarchy in regard to the pro-inflammatory potential of human

Granulocyte-macrophage colony stimulatory factor enhances the pro-inflammatory response of interferon-γ-treated macrophages to Pseudomonas aeruginosa infection.

Science.gov (United States)

Singh, Sonali; Barr, Helen; Liu, Yi-Chia; Robins, Adrian; Heeb, Stephan; Williams, Paul; Fogarty, Andrew; Cámara, Miguel; Martínez-Pomares, Luisa

2015-01-01

Pseudomonas aeruginosa is an opportunistic pathogen that can cause severe infections at compromised epithelial surfaces, such those found in burns, wounds, and in lungs damaged by mechanical ventilation or recurrent infections, particularly in cystic fibrosis (CF) patients. CF patients have been proposed to have a Th2 and Th17-biased immune response suggesting that the lack of Th1 and/or over exuberant Th17 responses could contribute to the establishment of chronic P. aeruginosa infection and deterioration of lung function. Accordingly, we have observed that interferon (IFN)-γ production by peripheral blood mononuclear cells from CF patients positively correlated with lung function, particularly in patients chronically infected with P. aeruginosa. In contrast, IL-17A levels tended to correlate negatively with lung function with this trend becoming significant in patients chronically infected with P. aeruginosa. These results are in agreement with IFN-γ and IL-17A playing protective and detrimental roles, respectively, in CF. In order to explore the protective effect of IFN-γ in CF, the effect of IFN-γ alone or in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF), on the ability of human macrophages to control P. aeruginosa growth, resist the cytotoxicity induced by this bacterium or promote inflammation was investigated. Treatment of macrophages with IFN-γ, in the presence and absence of GM-CSF, failed to alter bacterial growth or macrophage survival upon P. aeruginosa infection, but changed the inflammatory potential of macrophages. IFN-γ caused up-regulation of monocyte chemoattractant protein-1 (MCP-1) and TNF-α and down-regulation of IL-10 expression by infected macrophages. GM-CSF in combination with IFN-γ promoted IL-6 production and further reduction of IL-10 synthesis. Comparison of TNF-α vs. IL-10 and IL-6 vs. IL-10 ratios revealed the following hierarchy in regard to the pro-inflammatory potential of human macrophages

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

Science.gov (United States)

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

2014-01-01

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

Macrophage Interaction with Paracoccidioides brasiliensis Yeast Cells Modulates Fungal Metabolism and Generates a Response to Oxidative Stress.

Directory of Open Access Journals (Sweden)

Juliana Alves Parente-Rocha

Full Text Available Macrophages are key players during Paracoccidioides brasiliensis infection. However, the relative contribution of the fungal response to counteracting macrophage activity remains poorly understood. In this work, we evaluated the P. brasiliensis proteomic response to macrophage internalization. A total of 308 differentially expressed proteins were detected in P. brasiliensis during infection. The positively regulated proteins included those involved in alternative carbon metabolism, such as enzymes involved in gluconeogenesis, beta-oxidation of fatty acids and amino acids catabolism. The down-regulated proteins during P. brasiliensis internalization in macrophages included those related to glycolysis and protein synthesis. Proteins involved in the oxidative stress response in P. brasiliensis yeast cells were also up-regulated during macrophage infection, including superoxide dismutases (SOD, thioredoxins (THX and cytochrome c peroxidase (CCP. Antisense knockdown mutants evaluated the importance of CCP during macrophage infection. The results suggested that CCP is involved in a complex system of protection against oxidative stress and that gene silencing of this component of the antioxidant system diminished the survival of P. brasiliensis in macrophages and in a murine model of infection.

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

Science.gov (United States)

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

2012-01-01

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

Rainbow trout CK9, a CCL25-like ancient chemokine that attracts and regulates B cells and macrophages, the main antigen presenting cells in fish.

Science.gov (United States)

Aquilino, Carolina; Granja, Aitor G; Castro, Rosario; Wang, Tiehui; Abos, Beatriz; Parra, David; Secombes, Christopher J; Tafalla, Carolina

2016-04-05

CK9 is a rainbow trout (Oncorhynchus mykiss) CC chemokine phylogenetically related to mammalian CCL25. Although CK9 is known to be transcriptionally regulated in response to inflammation particularly in mucosal tissues, its functionality has never been revealed. In the current work, we have demonstrated that CK9 is chemoattractant for antigen presenting cells (APCs) expressing major histocompatibility complex class II (MHC II) on the cell surface. Among these APCs, CK9 has a strong chemotactic capacity for both B cells (IgM+ and IgT+) and macrophages. Along with its chemotactic capacities, CK9 modulated the MHC II turnover of B lymphocytes and up-regulated the phagocytic capacity of both IgM+ cells and macrophages. Although CK9 had no lymphoproliferative effects, it increased the survival of IgT+ lymphocytes. Furthermore, we have established that the chemoattractant capacity of CK9 is strongly increased after pre-incubation of leukocytes with a T-independent antigen, whereas B cell receptor (BCR) cross-linking strongly abrogated their capacity to migrate to CK9, indicating that CK9 preferentially attracts B cells at the steady state or under BCR-independent stimulation. These results point to CK9 being a key regulator of B lymphocyte trafficking in rainbow trout, able to modulate innate functions of teleost B lymphocytes and macrophages.

Of macrophages and red blood cells; a complex love story.

Science.gov (United States)

de Back, Djuna Z; Kostova, Elena B; van Kraaij, Marian; van den Berg, Timo K; van Bruggen, Robin

2014-01-01

Macrophages tightly control the production and clearance of red blood cells (RBC). During steady state hematopoiesis, approximately 10(10) RBC are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

Immunomodulatory activity of andrographolide on macrophage activation and specific antibody response

Science.gov (United States)

Wang, Wei; Wang, Jing; Dong, Sheng-fu; Liu, Chun-hong; Italiani, Paola; Sun, Shu-hui; Xu, Jing; Boraschi, Diana; Ma, Shi-ping; Qu, Di

2010-01-01

Aim: To investigate the immunomodulatory effects of andrographolide on both innate and adaptive immune responses. Methods: Andrographolide (10 μg/mL in vitro or 1 mg/kg in vivo) was used to modulate LPS-induced classical activated (M1) or IL-4-induced alternative activated (M2) macrophages in vitro and humor immune response to HBsAg in vivo. Cytokine gene expression profile (M1 vs M2) was measured by real-time PCR, IL-12/IL-10 level was detected by ELISA, and surface antigen expression was evaluated by flow cytometry, whereas phosphorylation level of ERK 1/2 and AKT was determined by Western blot. The level of anti-HBs antibodies in HBsAg immunized mice was detected by ELISA, and the number of HBsAg specific IL-4-producing splenocyte was enumerated by ELISPOT. Results: Andrographolide treatment in vitro attenuated either LPS or IL-4 induced macrophage activation, inhibited both M1 and M2 cytokines expression and decreased IL-12/IL-10 ratio (the ratio of M1/M2 polarization). Andrographolide down-regulated the expression of mannose receptor (CD206) in IL-4 induced macrophages and major histocompability complex/costimulatory molecules (MHC I, CD40, CD80, CD86) in LPS-induced macrophages. Correspondingly, anti-HBs antibody production and the number of IL-4-producing splenocytes were reduced by in vivo administration of andrographolide. Reduced phosphorylation levels of ERK1/2 and AKT were observed in macrophages treated with andrographolide. Conclusion: Andrographolide can modulate the innate and adaptive immune responses by regulating macrophage phenotypic polarization and Ag-specific antibody production. MAPK and PI3K signaling pathways may participate in the mechanisms of andrographolide regulating macrophage activation and polarization. PMID:20139902

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

International Nuclear Information System (INIS)

Okamoto, Mika; Wang, Xin; Baba, Masanori

2005-01-01

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

Cigarette smoking decreases global microRNA expression in human alveolar macrophages.

Directory of Open Access Journals (Sweden)

Joel W Graff

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

Nicotinamide: a vitamin able to shift macrophage differentiation toward macrophages with restricted inflammatory features.

Science.gov (United States)

Weiss, Ronald; Schilling, Erik; Grahnert, Anja; Kölling, Valeen; Dorow, Juliane; Ceglarek, Uta; Sack, Ulrich; Hauschildt, Sunna

2015-11-01

The differentiation of human monocytes into macrophages is influenced by environmental signals. Here we asked in how far nicotinamide (NAM), a vitamin B3 derivative known to play a major role in nicotinamide adenine dinucleotide (NAD)-mediated signaling events, is able to modulate monocyte differentiation into macrophages developed in the presence of granulocyte macrophage colony-stimulating factor (GM-MØ) or macrophage colony-stimulating factor (M-MØ). We found that GM-MØ undergo biochemical, morphological and functional modifications in response to NAM, whereas M-MØ were hardly affected. GM-MØ exposed to NAM acquired an M-MØ-like structure while the LPS-induced production of pro-inflammatory cytokines and COX-derived eicosanoids were down-regulated. In contrast, NAM had no effect on the production of IL-10 or the cytochrome P450-derived eicosanoids. Administration of NAM enhanced intracellular NAD concentrations; however, it did not prevent the LPS-mediated drain on NAD pools. In search of intracellular molecular targets of NAM known to be involved in LPS-induced cytokine and eicosanoid synthesis, we found NF-κB activity to be diminished. In conclusion, our data show that vitamin B3, when present during the differentiation of monocytes into GM-MØ, interferes with biochemical pathways resulting in strongly reduced pro-inflammatory features. © The Author(s) 2015.

The role of the tissue microenvironment in the regulation of cancer cell motility and invasion

Directory of Open Access Journals (Sweden)

Brábek Jan

2010-09-01

Full Text Available Abstract During malignant neoplastic progression the cells undergo genetic and epigenetic cancer-specific alterations that finally lead to a loss of tissue homeostasis and restructuring of the microenvironment. The invasion of cancer cells through connective tissue is a crucial prerequisite for metastasis formation. Although cell invasion is foremost a mechanical process, cancer research has focused largely on gene regulation and signaling that underlie uncontrolled cell growth. More recently, the genes and signals involved in the invasion and transendothelial migration of cancer cells, such as the role of adhesion molecules and matrix degrading enzymes, have become the focus of research. In this review we discuss how the structural and biomechanical properties of extracellular matrix and surrounding cells such as endothelial cells influence cancer cell motility and invasion. We conclude that the microenvironment is a critical determinant of the migration strategy and the efficiency of cancer cell invasion.

Functional Roles of p38 Mitogen-Activated Protein Kinase in Macrophage-Mediated Inflammatory Responses

Directory of Open Access Journals (Sweden)

Yanyan Yang

2014-01-01

Full Text Available Inflammation is a natural host defensive process that is largely regulated by macrophages during the innate immune response. Mitogen-activated protein kinases (MAPKs are proline-directed serine and threonine protein kinases that regulate many physiological and pathophysiological cell responses. p38 MAPKs are key MAPKs involved in the production of inflammatory mediators, including tumor necrosis factor-α (TNF-α and cyclooxygenase-2 (COX-2. p38 MAPK signaling plays an essential role in regulating cellular processes, especially inflammation. In this paper, we summarize the characteristics of p38 signaling in macrophage-mediated inflammation. In addition, we discuss the potential of using inhibitors targeting p38 expression in macrophages to treat inflammatory diseases.

The Current State of Nanoparticle-Induced Macrophage Polarization and Reprogramming Research

Directory of Open Access Journals (Sweden)

Xiaoyuan Miao

2017-02-01

Full Text Available Macrophages are vital regulators of the host defense in organisms. In response to different local microenvironments, resting macrophages (M0 can be polarized into different phenotypes, pro-inflammatory (M1 or anti-inflammatory (M2, and perform different roles in different physiological or pathological conditions. Polarized macrophages can also be further reprogrammed by reversing their phenotype according to the changed milieu. Macrophage polarization and reprogramming play essential roles in maintaining the steady state of the immune system and are involved in the processes of many diseases. As foreign substances, nanoparticles (NPs mainly target macrophages after entering the body. NPs can perturb the polarization and reprogramming of macrophages, affect their immunological function and, therefore, affect the pathological process of disease. Optimally-designed NPs for the modulation of macrophage polarization and reprogramming might provide new solutions for treating diseases. Systematically investigating how NPs affect macrophage polarization is crucial for understanding the regulatory effects of NPs on immune cells in vivo. In this review, macrophage polarization by NPs is summarized and discussed.

Metabolic-epigenetic crosstalk in macrophage activation

NARCIS (Netherlands)

Baardman, Jeroen; Licht, Iris; de Winther, Menno P. J.; van den Bossche, Jan

2015-01-01

Epigenetic enzymes are emerging as crucial controllers of macrophages, innate immune cells that determine the outcome of many inflammatory diseases. Recent studies demonstrate that the activity of particular chromatin-modifying enzymes is regulated by the availability of specific metabolites like

Small cell lung cancer: Recruitment of macrophages by circulating tumor cells.

Science.gov (United States)

Hamilton, Gerhard; Rath, Barbara; Klameth, Lukas; Hochmair, Maximilan J

2016-03-01

Tumor-associated macrophages (TAMs) play an important role in tumor progression, suppression of antitumor immunity and dissemination. Blood monocytes infiltrate the tumor region and are primed by local microenvironmental conditions to promote tumor growth and invasion. Although many of the interacting cytokines and factors are known for the tumor-macrophage interactions, the putative contribution of circulating tumor cells (CTCs) is not known so far. These specialized cells are characterized by increased mobility, ability to degrade the extracellular matrix (ECM) and to enter the blood stream and generate secondary lesions which is a leading cause of death for the majority of tumor patients. The first establishment of two permanent CTC lines, namely BHGc7 and 10, from blood samples of advanced stage small cell lung cancer (SCLC) patients allowed us to investigate the CTC-immune cell interaction. Cocultures of peripheral blood mononuclear cells (PBMNCs) with CTCs or addition of CTC-conditioned medium (CTC-CM) in vitro resulted in monocyte-macrophage differentiation and appearance of CD14 + , CD163 weak and CD68 + macrophages expressing markers of TAMs. Furthermore, we screened the supernatants of CTC-primed macrophages for presence of approximately 100 cytokines and compared the expression with those induced by the local metastatic SCLC26A cell line. Macrophages recruited by SCLC26A-CM showed expression of osteopontin (OPN), monocyte chemoattractant protein-1 (MCP-1), IL-8, chitinase3-like 1 (CHI3L1), platelet factor (Pf4), IL-1ra and matrix metalloproteinase-9 (MMP-9) among other minor cytokines/chemokines. In contrast, BHGc7-CM induced marked overexpression of complement factor D (CFD)/adipsin and vitamin D-BP (VDBP), as well as increased secretion of OPN, lipocalin-2 (LCN2), CHI3L1, uPAR, MIP-1 and GDF-15/MIC-1. BHGc10, derived independently from relapsed SCLC, revealed an almost identical pattern with added expression of ENA-78/CXCL5. CMs of the non-tumor HEK293

Strain specific transcriptional response in Mycobacterium tuberculosis infected macrophages

Directory of Open Access Journals (Sweden)

Koo Mi-Sun

2012-01-01

Full Text Available Abstract Background Tuberculosis (TB, a bacterial infection caused by Mycobacterium tuberculosis (Mtb remains a significant health problem worldwide with a third of the world population infected and nearly nine million new cases claiming 1.1 million deaths every year. The outcome following infection by Mtb is determined by a complex and dynamic host-pathogen interaction in which the phenotype of the pathogen and the immune status of the host play a role. However, the molecular mechanism by which Mtb strains induce different responses during intracellular infection of the host macrophage is not fully understood. To explore the early molecular events triggered upon Mtb infection of macrophages, we studied the transcriptional responses of murine bone marrow-derived macrophages (BMM to infection with two clinical Mtb strains, CDC1551 and HN878. These strains have previously been shown to differ in their virulence/immunogenicity in the mouse and rabbit models of pulmonary TB. Results In spite of similar intracellular growth rates, we observed that compared to HN878, infection by CDC1551 of BMM was associated with an increased global transcriptome, up-regulation of a specific early (6 hours immune response network and significantly elevated nitric oxide production. In contrast, at 24 hours post-infection of BMM by HN878, more host genes involved in lipid metabolism, including cholesterol metabolism and prostaglandin synthesis were up-regulated, compared to infection with CDC1551. In association with the differences in the macrophage responses to infection with the 2 Mtb strains, intracellular CDC1551 expressed higher levels of stress response genes than did HN878. Conclusions In association with the early and more robust macrophage activation, intracellular CDC1551 cells were exposed to a higher level of stress leading to increased up-regulation of the bacterial stress response genes. In contrast, sub-optimal activation of macrophages and induction of

Identification of a Novel Pathway of Transforming Growth Factor-β1 Regulation by Extracellular NAD+ in Mouse Macrophages

Science.gov (United States)

Zamora, Ruben; Azhar, Nabil; Namas, Rajaie; Metukuri, Mallikarjuna R.; Clermont, Thierry; Gladstone, Chase; Namas, Rami A.; Hermus, Linda; Megas, Cristina; Constantine, Gregory; Billiar, Timothy R.; Fink, Mitchell P.; Vodovotz, Yoram

2012-01-01

Extracellular β-nicotinamide adenine dinucleotide (NAD+) is anti-inflammatory. We hypothesized that NAD+ would modulate the anti-inflammatory cytokine Transforming Growth Factor (TGF)-β1. Indeed, NAD+ led to increases in both active and latent cell-associated TGF-β1 in RAW 264.7 mouse macrophages as well as in primary peritoneal macrophages isolated from both C3H/HeJ (TLR4-mutant) and C3H/HeOuJ (wild-type controls for C3H/HeJ) mice. NAD+ acts partially via cyclic ADP-ribose (cADPR) and subsequent release of Ca2+. Treatment of macrophages with the cADPR analog 3-deaza-cADPR or Ca2+ ionophores recapitulated the effects of NAD+ on TGF-β1, whereas the cADPR antagonist 8-Br-cADPR, Ca2+ chelation, and antagonism of L-type Ca2+ channels suppressed these effects. The time and dose effects of NAD+ on TGF-β1 were complex and could be modeled both statistically and mathematically. Model-predicted levels of TGF-β1 protein and mRNA were largely confirmed experimentally but also suggested the presence of other mechanisms of regulation of TGF-β1 by NAD+. Thus, in vitro and in silico evidence points to NAD+ as a novel modulator of TGF-β1. PMID:22829588

Identification of BCAP-{sub L} as a negative regulator of the TLR signaling-induced production of IL-6 and IL-10 in macrophages by tyrosine phosphoproteomics

Energy Technology Data Exchange (ETDEWEB)

Matsumura, Takayuki [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Oyama, Masaaki; Kozuka-Hata, Hiroko [Medical Proteomics Laboratory, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Ishikawa, Kosuke; Inoue, Takafumi [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Muta, Tatsushi [Laboratory of Cell Recognition and Response, Graduate School of Life Sciences, Tohoku University, Sendai 980-8578 (Japan); Semba, Kentaro, E-mail: ksemba@waseda.jp [Consolidated Research Institute for Advanced Science and Medical Care, Waseda University, Shinjuku-ku, Tokyo 162-0041 (Japan); Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480 (Japan); Inoue, Jun-ichiro, E-mail: jun-i@ims.u-tokyo.ac.jp [Medical Proteomics Laboratory, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan); Division of Cellular and Molecular Biology, Institute of Medical Science, University of Tokyo, Minato-ku, Tokyo 108-8639 (Japan)

2010-09-17

Research highlights: {yields} Twenty five tyrosine-phosphorylated proteins in LPS-stimulated macrophages were determined. {yields} BCAP is a novel tyrosine-phosphorylated protein in LPS-stimulated macrophages. {yields} BCAP-{sub L} inhibits IL-6 and IL-10 production in LPS-stimulated macrophages. -- Abstract: Toll-like receptor (TLR) signaling in macrophages is essential for anti-pathogen responses such as cytokine production and antigen presentation. Although numerous reports suggest that protein tyrosine kinases (PTKs) are involved in cytokine induction in response to lipopolysaccharides (LPS; TLR4 ligand) in macrophages, the PTK-mediated signal transduction pathway has yet to be analyzed in detail. Here, we carried out a comprehensive and quantitative dynamic tyrosine phosphoproteomic analysis on the TLR4-mediated host defense system in RAW264.7 macrophages using stable isotope labeling by amino acids in cell culture (SILAC). We determined the temporal profiles of 25 proteins based on SILAC-encoded peptide(s). Of these, we focused on the tyrosine phosphorylation of B-cell adaptor for phosphatidylinositol 3-kinase (BCAP) because the function of BCAP remains unknown in TLR signaling in macrophages. Furthermore, Bcap has two distinct transcripts, a full-length (Bcap-{sub L}) and an alternatively initiated or spliced (Bcap-{sub S}) mRNA, and little is known about the differential functions of the BCAP-{sub L} and BCAP-{sub S} proteins. Our study showed, for the first time, that RNAi-mediated selective depletion of BCAP-{sub L} enhanced IL-6 and IL-10 production but not TNF-{alpha} production in TLR ligand-stimulated macrophages. We propose that BCAP-{sub L} (but not BCAP-{sub S}) is a negative regulator of the TLR-mediated host defense system in macrophages.

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

Science.gov (United States)

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

2016-11-01

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

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

Science.gov (United States)

Lechner, F; Machado, J; Bertoni, G; Seow, H F; Dobbelaere, D A; Peterhans, E

1997-01-01

Caprine arthritis encephalitis virus (CAEV) is a lentivirus of goats that leads to chronic mononuclear infiltration of various tissues, in particular, the radiocarpal joints. Cells of the monocyte/macrophage lineage are the major host cells of CAEV in vivo. We have shown that infection of cultured goat macrophages with CAEV results in an alteration of cytokine expression in vitro. Constitutive expression of interleukin 8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) was increased in infected macrophages, whereas transforming growth factor beta1 (TGF-beta1) mRNA was down-regulated. When macrophages were infected with a CAEV clone lacking the trans-acting nuclear regulatory gene tat, IL-8 and MCP-1 were also increased. No significant differences from cells infected with the wild-type clone were observed, suggesting that Tat is not required for the increased expression of IL-8 and MCP-1 in infected macrophages. Furthermore, infection with CAEV led to an altered pattern of cytokine expression in response to lipopolysaccharide (LPS), heat-killed Listeria monocytogenes plus gamma interferon, or fixed cells of Staphylococcus aureus Cowan I. In infected macrophages, tumor necrosis factor alpha, IL-1beta, IL-6, and IL-12 p40 mRNA expression was reduced in response to all stimuli tested whereas changes in expression of granulocyte-macrophage colony-stimulating factor depended on the stimulating agent. Electrophoretic mobility shift assays demonstrated that, in contrast to effects of human immunodeficiency virus infection of macrophages, CAEV infection had no effect on the level of constitutive nuclear factor-kappaB (NF-kappaB) activity or on the level of LPS-stimulated NF-kappaB activity, suggesting that NF-kappaB is not involved in altered regulation of cytokine expression in CAEV-infected cells. In contrast, activator protein 1 (AP-1) binding activity was decreased in infected macrophages. These data show that CAEV infection may result in a dysregulation of

Impaired chromatin remodelling at STAT1-regulated promoters leads to global unresponsiveness of Toxoplasma gondii-infected macrophages to IFN-γ.

Directory of Open Access Journals (Sweden)

Christine Lang

2012-01-01

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

MIIP, a cytoskeleton regulator that blocks cell migration and invasion, delays mitosis, and suppresses tumorogenesis.

Science.gov (United States)

Wang, Yingmei; Wen, Jing; Zhang, Wei

2011-02-01

The migration and invasion inhibitory protein (MIIP) was initially discovered in a yeast two-hybrid screen for proteins that interact and inhibit the migration and invasion-promoting protein insulin-like growth factor binding protein 2 (IGFBP2). Recent studies have shown that MIIP not only modulates IGFBP2 but also regulates microtubule by binding to and inhibiting HDAC6, a class 2 histone deacetylase that deacetylates α-tubulin, heat-shock protein 90 (HSP90), and cortactin. In addition, MIIP also regulates the mitosis checkpoint, another microtubule-associated process. The location of the MIIP gene in chromosomal region 1p36, a commonly deleted region in a broad spectrum of human cancers, and the observation that MIIP attenuates tumorigenesis in a mouse model suggest that it functions as a tumor-suppressor gene. This review summarizes the recent progress in characterizing this novel protein, which regulates cell migration and mitosis, two processes that rely on the highly coordinated dynamics of the microtubule and cytoskeleton systems.

Modulating macrophage polarization with divalent cations in nanostructured titanium implant surfaces

International Nuclear Information System (INIS)

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

2016-01-01

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. (paper)

Modulating macrophage polarization with divalent cations in nanostructured titanium implant surfaces

Science.gov (United States)

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.

Alpha1 and Alpha2 Integrins Mediate Invasive Activity of Mouse Mammary Carcinoma Cells through Regulation of Stromelysin-1 Expression

Energy Technology Data Exchange (ETDEWEB)

Lochter, Andre; Navre, Marc; Werb, Zena; Bissell, Mina J

1998-06-29

Tumor cell invasion relies on cell migration and extracellular matrix proteolysis. We investigated the contribution of different integrins to the invasive activity of mouse mammary carcinoma cells. Antibodies against integrin subunits {alpha}6 and {beta}1, but not against {alpha}1 and {alpha}2, inhibited cell locomotion on a reconstituted basement membrane in two-dimensional cell migration assays, whereas antibodies against {beta}1, but not against a6 or {alpha}2, interfered with cell adhesion to basement membrane constituents. Blocking antibodies against {alpha}1 integrins impaired only cell adhesion to type IV collagen. Antibodies against {alpha}1, {alpha}2, {alpha}6, and {beta}1, but not {alpha}5, integrin subunits reduced invasion of a reconstituted basement membrane. Integrins {alpha}1 and {alpha}2, which contributed only marginally to motility and adhesion, regulated proteinase production. Antibodies against {alpha}1 and {alpha}2, but not {alpha}6 and {beta}1, integrin subunits inhibited both transcription and protein expression of the matrix metalloproteinase stromelysin-1. Inhibition of tumor cell invasion by antibodies against {alpha}1 and {alpha}2 was reversed by addition of recombinant stromelysin-1. In contrast, stromelysin-1 could not rescue invasion inhibited by anti-{alpha}6 antibodies. Our data indicate that {alpha}1 and {alpha}2 integrins confer invasive behavior by regulating stromelysin-1 expression, whereas {alpha}6 integrins regulate cell motility. These results provide new insights into the specific functions of integrins during tumor cell invasion.

CpG-ODNs induces up-regulated expression of chemokine CCL9 in mouse macrophages and microglia

Digital Repository Service at National Institute of Oceanography (India)

Ravindran, C.; Cheng, Y.-C.; Liang, S.-M.

G-ODNs on macrophage/microglial cells are investigated. CpG-ODNs enhanced the expression of TLR9 mRNA of RAW264.7 macrophage and BV2 microglia cells time dependently. The expression of CCL9 of macrophages/microglia showed different responsiveness upon stimulation...

Of macrophages and red blood cells; a complex love story.

Directory of Open Access Journals (Sweden)

Djuna Zoe de Back

2014-01-01

Full Text Available Macrophages tightly control the production and clearance of red blood cells (RBC. During steady state haematopoiesis, approximately 1010 red blood cells are produced per hour within erythroblastic islands in humans. In these erythroblastic islands, resident bone marrow macrophages provide erythroblasts with interactions that are essential for erythroid development. New evidence suggests that not only under homeostasis but also under stress conditions, macrophages play an important role in promoting erythropoiesis. Once RBC have matured, these cells remain in circulation for about 120 days. At the end of their life span, RBC are cleared by macrophages residing in the spleen and the liver. Current theories about the removal of senescent RBC and the essential role of macrophages will be discussed as well as the role of macrophages in facilitating the removal of damaged cellular content from the RBC. In this review we will provide an overview on the role of macrophages in the regulation of RBC production, maintenance and clearance. In addition, we will discuss the interactions between these two cell types during transfer of immune complexes and pathogens from RBC to macrophages.

Monocyte to macrophage differentiation-associated (MMD) targeted by miR-140-5p regulates tumor growth in non-small cell lung cancer

International Nuclear Information System (INIS)

Li, Weina; He, Fei

2014-01-01

Highlights: • Expression of MMD is increased in lung cancer tissues. • Knockdown of MMD inhibits growth of A549 and LLC cells in vitro and in vivo. • MMD is a direct functional target of miR-140-5p. • MiR-140-5p/MMD axis regulates Erk1/2 signaling. - Abstract: Monocyte to macrophage differentiation-associated (MMD) is identified in macrophages as a gene associated with the differentiation from monocytes to macrophages. Recent microarray analysis for non-small cell lung cancer (NSCLC) suggests that MMD is an important signature associated with relapse and survival among patients with NSCLC. Therefore, we speculate that MMD likely plays a role in lung cancer. In this study, we found that the protein level of MMD was increased in lung cancer compared to benign lung tissues, and knockdown of MMD inhibited the growth of A549 and Lewis lung cancer cells (LLC) in vitro and in vivo. Integrated analysis demonstrated that MMD was a direct functional target of miR-140-5p. Furthermore, we found that miR-140-5p/MMD axis could affect the cell proliferation of lung cancer cells by regulating Erk signaling. Together, our results highlight the significance of miR-140-5p/MMD axis in lung cancer, and miR-140-5p/MMD axis could serve as new molecular targets for the therapy against lung cancer

SIRT2 ameliorates lipopolysaccharide-induced inflammation in macrophages

International Nuclear Information System (INIS)

Lee, Ae Sin; Jung, Yu Jin; Kim, Dal; Nguyen-Thanh, Tung; Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang; Kim, Won

2014-01-01

Highlights: • Knockout of SIRT2 attenuates lipopolysaccharide-induced iNOS expression. • Lipopolysaccharide-induced NO production is decreased in SIRT2 KO macrophage. • SIRT2 deficiency suppresses lipopolysaccharide-induced ROS production in macrophage. • M1-macrophage related factors are decreased in SIRT2 deficient cells. • SIRT2 deficiency decreases lipopolysaccharide-induced activation of NFκB. - Abstract: Introduction: SIRT2 is a NAD(+)-dependent deacetylases and associated with numerous processes such as infection, carcinogenesis, DNA damage and cell cycle regulation. However, the role of SIRT2 in inflammatory process in macrophage remains unclear. Materials and methods: In the present study, we have evaluated the regulatory effects of SIRT2 in lipopolysaccharide (LPS)-stimulated macrophages isolated from SIRT2 knockout (KO) and wild type (WT) mice or Raw264.7 macrophage cells. As inflammatory parameters, expression of inducible nitric oxide synthase (iNOS), the productions of nitric oxide, reactive oxygen species (ROS) and M1-macrophage-related factors were evaluated. We also examined the effects of SIRT2 on activation of nuclear factor-kappaB (NFκB) signaling. Results: SIRT2 deficiency inhibits LPS-induced iNOS mRNA and protein expression in bone marrow derived macrophages. SIRT2-siRNA transfection also suppressed LPS-induced iNOS expression in Raw264.7 macrophage cells. Bone marrow derived macrophages isolated from SIRT2 KO mice produced lower nitric oxide and expressed lower levels of M1-macrophage related markers including iNOS and CD86 in response to LPS than WT mice. Decrease of SIRT2 reduced the LPS-induced reactive oxygen species production. Deficiency of SIRT2 resulted in inhibition of NFκB activation through reducing the phosphorylation and degradation of IκBα. The phosphorylation and nuclear translocation of p65 was significantly decreased in SIRT2-deficient macrophages after LPS stimulation. Discussion: Our data suggested that

SIRT2 ameliorates lipopolysaccharide-induced inflammation in macrophages

Energy Technology Data Exchange (ETDEWEB)

Lee, Ae Sin; Jung, Yu Jin; Kim, Dal; Nguyen-Thanh, Tung [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Kang, Kyung Pyo [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Research Institute of Clinical Medicine of Chonbuk National University, Chonbuk National University Hospital, Jeonju (Korea, Republic of); Lee, Sik [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Park, Sung Kwang [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Research Institute of Clinical Medicine of Chonbuk National University, Chonbuk National University Hospital, Jeonju (Korea, Republic of); Kim, Won, E-mail: kwon@jbnu.ac.kr [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Research Institute of Clinical Medicine of Chonbuk National University, Chonbuk National University Hospital, Jeonju (Korea, Republic of)

2014-08-08

Highlights: • Knockout of SIRT2 attenuates lipopolysaccharide-induced iNOS expression. • Lipopolysaccharide-induced NO production is decreased in SIRT2 KO macrophage. • SIRT2 deficiency suppresses lipopolysaccharide-induced ROS production in macrophage. • M1-macrophage related factors are decreased in SIRT2 deficient cells. • SIRT2 deficiency decreases lipopolysaccharide-induced activation of NFκB. - Abstract: Introduction: SIRT2 is a NAD(+)-dependent deacetylases and associated with numerous processes such as infection, carcinogenesis, DNA damage and cell cycle regulation. However, the role of SIRT2 in inflammatory process in macrophage remains unclear. Materials and methods: In the present study, we have evaluated the regulatory effects of SIRT2 in lipopolysaccharide (LPS)-stimulated macrophages isolated from SIRT2 knockout (KO) and wild type (WT) mice or Raw264.7 macrophage cells. As inflammatory parameters, expression of inducible nitric oxide synthase (iNOS), the productions of nitric oxide, reactive oxygen species (ROS) and M1-macrophage-related factors were evaluated. We also examined the effects of SIRT2 on activation of nuclear factor-kappaB (NFκB) signaling. Results: SIRT2 deficiency inhibits LPS-induced iNOS mRNA and protein expression in bone marrow derived macrophages. SIRT2-siRNA transfection also suppressed LPS-induced iNOS expression in Raw264.7 macrophage cells. Bone marrow derived macrophages isolated from SIRT2 KO mice produced lower nitric oxide and expressed lower levels of M1-macrophage related markers including iNOS and CD86 in response to LPS than WT mice. Decrease of SIRT2 reduced the LPS-induced reactive oxygen species production. Deficiency of SIRT2 resulted in inhibition of NFκB activation through reducing the phosphorylation and degradation of IκBα. The phosphorylation and nuclear translocation of p65 was significantly decreased in SIRT2-deficient macrophages after LPS stimulation. Discussion: Our data suggested that

MicroRNA 21 Is a Homeostatic Regulator of Macrophage Polarization and Prevents Prostaglandin E2-Mediated M2 Generation

OpenAIRE

Wang, Zhuo; Brandt, Stephanie; Medeiros, Alexandra; Wang, Soujuan; Wu, Hao; Dent, Alexander; Serezani, C. Henrique

2015-01-01

Macrophages dictate both initiation and resolution of inflammation. During acute inflammation classically activated macrophages (M1) predominate, and during the resolution phase alternative macrophages (M2) are dominant. The molecular mechanisms involved in macrophage polarization are understudied. MicroRNAs are differentially expressed in M1 and M2 macrophages that influence macrophage polarization. We identified a role of miR-21 in macrophage polarization, and found that cross-talk between ...

Macrophage-derived insulin-like growth factor-1 affects influenza vaccine efficacy through the regulation of immune cell homeostasis.

Science.gov (United States)

Yoon, Il-Sub; Park, Hyelim; Kwak, Hye-Won; Woo Jung, Yong; Nam, Jae-Hwan

2017-08-24

The level of antibody production induced by a vaccine involves a variety of host factors. One of these, insulin-like growth factor-1 (IGF-1), plays an important role in lymphocyte maturation and antibody expression. Here, we investigated the role of macrophage-derived IGF-1 in the induction of influenza vaccine-specific antibodies using macrophage-derived IGF-1 gene knockout (MIKO) mice. The titers of vaccine-specific total immunoglobulin G (IgG) and IgG1 after immunization were about two- to fourfold lower in MIKO mice than in WT mice. Moreover, MIKO mice showed a relatively weak booster effect of repeated immunization. In contrast, antigen-nonspecific total IgG was about threefold higher in MIKO mice than in WT mice. After viral challenge, the viral titer and the pathological damage in lungs of MIKO mice were higher than those in WT mice despite vaccination. Interestingly, the proportions of proinflammatory immune cells including M1 macrophages, Th1 and Th17 cells was higher in unvaccinated MIKO mice than in unvaccinated WT mice. This suggests that nonspecific activation of immune cells may paradoxically impair the response to the vaccine. In addition, although the proportions of T follicular helper (Tfh) cells and GL-7 + germinal center (GC) B cells were higher in MIKO mice than in WT mice, the population of CD138 + B220 + antibody-secreting plasmablasts was lower in MIKO mice, which may be a cause of the low influenza-specific antibody titer in MIKO mice. Taken together, these results suggest that macrophage-derived IGF-1 might play an important role in the vaccine-triggered immune response by regulating immune cell homeostasis. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

Science.gov (United States)

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

2016-08-19

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

Macrophage ABCA2 deletion modulates intracellular cholesterol deposition, affects macrophage apoptosis, and decreases early atherosclerosis in LDL receptor knockout mice.

Science.gov (United States)

Calpe-Berdiel, Laura; Zhao, Ying; de Graauw, Marjo; Ye, Dan; van Santbrink, Peter J; Mommaas, A Mieke; Foks, Amanda; Bot, Martine; Meurs, Illiana; Kuiper, Johan; Mack, Jody T; Van Eck, Miranda; Tew, Kenneth D; van Berkel, Theo J C

2012-08-01

The ABCA2 transporter shares high structural homology to ABCA1, which is crucial for the removal of excess cholesterol from macrophages and, by extension, in atherosclerosis. It has been suggested that ABCA2 sequesters cholesterol inside the lysosomes, however, little is known of the macrophage-specific role of ABCA2 in regulating lipid homeostasis in vivo and in modulating susceptibility to atherosclerosis. Chimeras with dysfunctional macrophage ABCA2 were generated by transplantation of bone marrow from ABCA2 knockout (KO) mice into irradiated LDL receptor (LDLr) KO mice. Interestingly, lack of ABCA2 in macrophages resulted in a diminished lesion size in the aortic root (-24.5%) and descending thoracic aorta (-36.6%) associated with a 3-fold increase in apoptotic cells, as measured by both caspase 3 and TUNEL. Upon oxidized LDL exposure, macrophages from wildtype (WT) transplanted animals developed filipin-positive droplets in lysosomal-like compartments, corresponding to free cholesterol (FC) accumulation. In contrast, ABCA2-deficient macrophages displayed an abnormal diffuse distribution of FC over peripheral regions. The accumulation of neutral sterols in lipid droplets was increased in ABCA2-deficient macrophages, but primarily in cytoplasmic clusters and not in lysosomes. Importantly, apoptosis of oxLDL loaded macrophages lacking ABCA2 was increased 2.7-fold, probably as a consequence of the broad cellular distribution of FC. Lack of functional ABCA2 generates abnormalities in intracellular lipid distribution/trafficking in macrophages consistent with its lysosomal sequestering role, leading to an increased susceptibility to apoptosis in response to oxidized lipids and reduced atherosclerotic lesion development. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

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

Directory of Open Access Journals (Sweden)

Karl J Staples

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

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

Energy Technology Data Exchange (ETDEWEB)

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

2015-11-27

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

Membrane cholesterol regulates lysosome-plasma membrane fusion events and modulates Trypanosoma cruzi invasion of host cells.

Directory of Open Access Journals (Sweden)

Bárbara Hissa

Full Text Available BACKGROUND: Trypomastigotes of Trypanosoma cruzi are able to invade several types of non-phagocytic cells through a lysosomal dependent mechanism. It has been shown that, during invasion, parasites trigger host cell lysosome exocytosis, which initially occurs at the parasite-host contact site. Acid sphingomyelinase released from lysosomes then induces endocytosis and parasite internalization. Lysosomes continue to fuse with the newly formed parasitophorous vacuole until the parasite is completely enclosed by lysosomal membrane, a process indispensable for a stable infection. Previous work has shown that host membrane cholesterol is also important for the T. cruzi invasion process in both professional (macrophages and non-professional (epithelial phagocytic cells. However, the mechanism by which cholesterol-enriched microdomains participate in this process has remained unclear. METHODOLOGY/PRINCIPAL FINDING: In the present work we show that cardiomyocytes treated with MβCD, a drug able to sequester cholesterol from cell membranes, leads to a 50% reduction in invasion by T. cruzi trypomastigotes, as well as a decrease in the number of recently internalized parasites co-localizing with lysosomal markers. Cholesterol depletion from host membranes was accompanied by a decrease in the labeling of host membrane lipid rafts, as well as excessive lysosome exocytic events during the earlier stages of treatment. Precocious lysosomal exocytosis in MβCD treated cells led to a change in lysosomal distribution, with a reduction in the number of these organelles at the cell periphery, and probably compromises the intracellular pool of lysosomes necessary for T. cruzi invasion. CONCLUSION/SIGNIFICANCE: Based on these results, we propose that cholesterol depletion leads to unregulated exocytic events, reducing lysosome availability at the cell cortex and consequently compromise T. cruzi entry into host cells. The results also suggest that two different pools of

Proteolytic shedding of the macrophage scavenger receptor CD163 in multiple sclerosis

DEFF Research Database (Denmark)

Fabriek, Babs O; Møller, Holger J; Vloet, Rianka P M

2007-01-01

The scavenger receptor CD163 is selectively expressed on tissue macrophages and human monocytes. CD163 has been implicated to play a role in the clearance of hemoglobin and in the regulation of cytokine production by macrophages. Membrane CD163 can be cleaved by matrix metalloproteinases (MMP...

Complement regulation in murine and human hypercholesterolemia and role in the control of macrophage and smooth muscle cell proliferation.

Science.gov (United States)

Verdeguer, Francisco; Castro, Claudia; Kubicek, Markus; Pla, Davinia; Vila-Caballer, Marian; Vinué, Angela; Civeira, Fernando; Pocoví, Miguel; Calvete, Juan José; Andrés, Vicente

2007-11-01

Mounting evidence suggests that activation of complement, an important constituent of innate immunity, contributes to atherosclerosis. Here we investigated the expression of complement components (CCs) in the setting of experimental and clinical hypercholesterolemia, a major risk factor for atherosclerosis, their effects on vascular smooth muscle cell (VSMC) and macrophage proliferation, and the underlying molecular mechanisms. For this study we analyzed the mRNA and protein expression of several CCs in plasma and aorta of hypercholesterolemic atherosclerosis-prone apolipoprotein E-null mice (apoE-KO) and in plasma of normocholesterolemic subjects and familial hypercholesterolemia (FH) patients. We also carried out in vitro molecular studies to assess the role of CCs on the control of macrophage and VSMC proliferation. Fat-fed apoE-KO mice experiencing severe hypercholesterolemia (approximately 400 mg/dL), but not fat-fed wild-type controls with plasma cholesterol levelfeeding when hypercholesterolemia was manifested yet atherosclerotic lesions were absent or incipient. Rapid C3 and C4 protein upregulation was also observed in the plasma of fat-fed apoE-KO mice, and FH patients exhibited higher plasmatic C3a, C4 gamma chain, C1s and C3c alpha chain protein levels than normocholesterolemic subjects. In vitro, C3 and C3a, but not C3a-desArg, C4 and C1q, promoted macrophage and VSMC proliferation through Gi protein-dependent activation of extracellular signal-regulated kinase 1/2 (ERK1/2). We also found that C3-enriched FH plasma evoked a stronger mitogenic response in macrophages than normocholesterolemic plasma, and treatment with anti-C3 antibodies eliminated this difference. Both experimental and clinical hypercholesterolemia coincides with a concerted activation of several CCs. However, only C3 and C3a elicited a mitogenic response in cultured VSMCs and macrophages through Gi protein-dependent ERK1/2 activation. Thus, excess of C3/C3a in hypercholesterolemic apo

Beta 2-adrenergic receptor agonists are novel regulators of macrophage activation in diabetic renal and cardiovascular complications.

Science.gov (United States)

Noh, Hyunjin; Yu, Mi Ra; Kim, Hyun Joo; Lee, Ji Hye; Park, Byoung-Won; Wu, I-Hsien; Matsumoto, Motonobu; King, George L

2017-07-01

Macrophage activation is increased in diabetes and correlated with the onset and progression of vascular complications. To identify drugs that could inhibit macrophage activation, we developed a cell-based assay and screened a 1,040 compound library for anti-inflammatory effects. Beta2-adrenergic receptor (β2AR) agonists were identified as the most potent inhibitors of phorbol myristate acetate-induced tumor necrosis factor-α production in rat bone marrow macrophages. In peripheral blood mononuclear cells isolated from streptozotocin-induced diabetic rats, β2AR agonists inhibited diabetes-induced tumor necrosis factor-α production, which was prevented by co-treatment with a selective β2AR blocker. To clarify the underlying mechanisms, THP-1 cells and bone marrow macrophages were exposed to high glucose. High glucose reduced β-arrestin2, a negative regulator of NF-κB activation, and its interaction with IκBα. This subsequently enhanced phosphorylation of IκBα and activation of NF-κB. The β2AR agonists enhanced β-arrestin2 and its interaction with IκBα, leading to downregulation of NF-κB. A siRNA specific for β-arrestin2 reversed β2AR agonist-mediated inhibition of NF-κB activation and inflammatory cytokine production. Treatment of Zucker diabetic fatty rats with a β2AR agonist for 12 weeks attenuated monocyte activation as well as pro-inflammatory and pro-fibrotic responses in the kidneys and heart. Thus, β2AR agonists might have protective effects against diabetic renal and cardiovascular complications. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Toxoplasma gondii exposes phosphatidylserine inducing a TGF-β1 autocrine effect orchestrating macrophage evasion

International Nuclear Information System (INIS)

Seabra, Sergio H.; Souza, Wanderley de; Matta, Renato A. da

2004-01-01

Toxoplasmosis is a worldwide disease caused by Toxoplasma gondii. Activated macrophages control T. gondii growth by nitric oxide (NO) production. However, T. gondii active invasion inhibits NO production, allowing parasite persistence. Here we show that the mechanism used by T. gondii to inhibit NO production persisting in activated macrophages depends on phosphatidylserine (PS) exposure. Masking PS with annexin-V on parasites or activated macrophages abolished NO production inhibition and parasite persistence. NO production inhibition depended on a transforming growth factor-β 1 (TGF-β 1 ) autocrine effect confirmed by the expression of Smad 2 and 3 in infected macrophages. TGF-β 1 led to inducible nitric oxide synthase (iNOS) degradation, actin filament (F-actin) depolymerization, and lack of nuclear factor-κB (NF-κB) in the nucleus. All these features were reverted by TGF-β 1 neutralizing antibody treatment. Thus, T. gondii mimics the evasion mechanism used by Leishmania amazonensis and also the anti-inflammatory response evoked by apoptotic cells

Regulation of LPS-induced mRNA expression of pro-inflammatory cytokines via alteration of NF-κB activity in mouse peritoneal macrophages exposed to fluoride.

Science.gov (United States)

Tian, Yuhu; Huo, Meijun; Li, Guangsheng; Li, Yanyan; Wang, Jundong

2016-10-01

F toxicity to immune system, especially to macrophage, has been studied a lot recently. Nuclear factor-kappa B (NF-κB), as a transcription factor, plays a central role in immune and inflammatory responses via the regulation of downstream gene expression. Recent studies indicated that fluoride effect on inflammatory cytokine secretion, however, the molecular mechanism was less understood. In our study, peritoneal macrophages (PMs) were divided several groups and were administrated sodium fluoride (NaF, 50, 100, 200, 400, 800 μM) and/or lipopolysaccharide (LPS, 30 ng/mg). The mRNA expression of p65, inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) in macrophages exposed to fluoride was determined by quantitative real-time RT-PCR respectively. The translocation of NF-κB from cytoplasm to nucleus, which in a way reflects NF-κB activity, was demonstrated by Immunofluorescence and ELISA. Our results showed that fluoride had a dose-dependent effect on NF-κB activity, which coincided with LPS-induced mRNA expression of its downstream genes, iNOS and IL-1β. Fluoride alone causes no effect on gene expression. However, the mRNA expression of TNF-α showed non-NF-κB-dependent manner. Therefore, we come to the conclusion that fluoride can regulate LPS-induced mRNA expression of iNOS and IL-1β via NF-κB pathway in mouse peritoneal macrophages. Copyright © 2016 Elsevier Ltd. All rights reserved.

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

Science.gov (United States)

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

2014-04-01

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

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

Energy Technology Data Exchange (ETDEWEB)

Zhan, Chuan, E-mail: zhchuansy@163.com; Ma, Cheng-bin; Yuan, Hong-mou; Cao, Bao-yuan; Zhu, Jia-jun

2015-12-04

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 sciatic 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.

Enhanced SCAP glycosylation by inflammation induces macrophage foam cell formation.

Directory of Open Access Journals (Sweden)

Chao Zhou

Full Text Available Inflammatory stress promotes foam cell formation by disrupting LDL receptor feedback regulation in macrophages. Sterol Regulatory Element Binding Proteins (SREBPs Cleavage-Activating Protein (SCAP glycosylation plays crucial roles in regulating LDL receptor and 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCoAR feedback regulation. The present study was to investigate if inflammatory stress disrupts LDL receptor and HMGCoAR feedback regulation by affecting SCAP glycosylation in THP-1 macrophages. Intracellular cholesterol content was assessed by Oil Red O staining and quantitative assay. The expression of molecules controlling cholesterol homeostasis was examined using real-time quantitative RT-PCR and Western blotting. The translocation of SCAP from the endoplasmic reticulum (ER to the Golgi was detected by confocal microscopy. We demonstrated that exposure to inflammatory cytokines increased lipid accumulation in THP-1 macrophages, accompanying with an increased SCAP expression even in the presence of a high concentration of LDL. These inflammatory cytokines also prolonged the half-life of SCAP by enhancing glycosylation of SCAP due to the elevated expression of the Golgi mannosidase II. This may enhance translocation and recycling of SCAP between the ER and the Golgi, escorting more SREBP2 from the ER to the Golgi for activation by proteolytic cleavages as evidenced by an increased N-terminal of SREBP2 (active form. As a consequence, the LDL receptor and HMGCoAR expression were up-regulated. Interestingly, these effects could be blocked by inhibitors of Golgi mannosidases. Our results indicated that inflammation increased native LDL uptake and endogenous cholesterol de novo synthesis, thereby causing foam cell formation via increasing transcription and protein glycosylation of SCAP in macrophages. These data imply that inhibitors of Golgi processing enzymes might have a potential vascular-protective role in prevention of atherosclerotic foam

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

Science.gov (United States)

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

2016-05-01

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

Radiation induces invasiveness of pancreatic cancer via up-regulation of heparanase

International Nuclear Information System (INIS)

Lerner, I.; Bensoussan, E.; Meirovitz, A.; Elkin, M.; Vlodavsky, I.

2013-01-01

The full text of the publication follows. Pancreatic cancer is one of the most aggressive neoplasms with an extremely low survival rate. Because most pancreatic carcinoma patients miss the opportunity for complete surgical resection at the time of diagnosis, radiotherapy remains a major component of treatment modalities. However, pancreatic cancer often shows resistance to radiation therapy. Ionizing radiation (IR)-induced aggressiveness is emerging as one of the important mechanisms responsible for the limited benefit of radiation therapy in pancreatic cancer, but the identity of downstream effectors responsible for this effect remains poorly investigated. Here we report that IR promotes pancreatic cancer aggressiveness through up-regulation of the heparanase. Heparanase is a predominant mammalian enzyme capable of degrading heparan sulfate (HS), the main polysaccharide component of the basement membrane and other types of extracellular matrix (ECM). Cleavage of HS by heparanase leads to disassembly of ECM, enables cell invasion, releases HS-bound angiogenic and growth factors from the ECM depots, and generates bioactive HS fragments. We found that clinically relevant doses of IR augment invasive ability of pancreatic cells in vitro and in vivo via induction of heparanase. Our results indicate that the effect of IR on heparanase expression is mediated by Egr1 transcription factor. Moreover, specific inhibitor of heparanase enzymatic activity abolished IR-induced invasiveness of pancreatic carcinoma cells in vitro, while combined treatment with IR and the heparanase inhibitor, but not IR alone, attenuated ortho-topic pancreatic tumor progression in vivo. The proposed up-regulation of heparanase by IR represents a new molecular pathway through which IR may promote pancreatic tumor aggressiveness, providing explanation for the limited benefit from radiation therapy in pancreatic cancer. Our research is expected to offer a new approach to improve the efficacy of

Up-regulation of miR-146a contributes to the inhibition of invasion of pancreatic cancer cells

Science.gov (United States)

Li, Yiwei; VandenBoom, Timothy G.; Wang, Zhiwei; Kong, Dejuan; Ali, Shadan; Philip, Philip A.; Sarkar, Fazlul H.

2009-01-01

Pancreatic cancer (PC) is an aggressive malignancy with high mortality and is believed to be in part due to its highly invasive and metastatic behavior, which is associated with over-expression of EGFR and activation of NF-κB. Emerging evidence also suggest critical roles of microRNAs (miRNAs) in the regulation of various pathobiological processes including metastasis in PC and in other human malignancies. In the present study, we found lower expression of miR-146a in PC cells compared to normal human pancreatic duct epithelial (HPDE) cells. Interestingly, re-expression of miR-146a inhibited the invasive capacity of Colo357 and Panc-1 PC cells with concomitant down-regulation of EGFR and IRAK-1. Mechanistic studies including miR-146a re-expression, anti-miR-146 transfection, and EGFR knock-down experiment showed that there was a crosstalk between EGFR, MTA-2, IRAK-1, IκBα and NF-κB. Most importantly, we found that the treatment of PC cells with “natural agents” [3,3′-diinodolylmethane (DIM) or isoflavone] led to an increase in the expression of miR-146a and consequently down-regulated the expression of EGFR, MTA-2, IRAK-1 and NF-κB, resulting in the inhibition of invasion of Colo357 and Panc-1 cells. These results provide experimental evidence in support of the role of DIM and isoflavone as potential non-toxic agents as regulators of miRNA, which could be useful for the inhibition of cancer cell invasion and metastasis, and further suggesting that these agents could be important for designing novel targeted strategy for the treatment of PC. PMID:25242818

miR-664 negatively regulates PLP2 and promotes cell proliferation and invasion in T-cell acute lymphoblastic leukaemia

Energy Technology Data Exchange (ETDEWEB)

Zhu, Hong; Miao, Mei-hua; Ji, Xue-qiang; Xue, Jun; Shao, Xue-jun, E-mail: xuejunshao@hotmail.com

2015-04-03

MicroRNAs (miRNAs) play important roles in the pathogenesis of many types of cancers by negatively regulating gene expression at posttranscriptional level. However, the role of microRNAs in leukaemia, particularly T-cell acute lymphoblastic leukaemia (T-ALL), has remained elusive. Here, we identified miR-664 and its predicted target gene PLP2 were differentially expressed in T-ALL using bioinformatics methods. In T-ALL cell lines, CCK-8 proliferation assay indicated that the cell proliferation was promoted by miR-664, while miR-664 inhibitor could significantly inhibited the proliferation. Moreover, migration and invasion assay showed that overexpression of miR-664 could significantly promoted the migration and invasion of T-ALL cells, whereas miR-664 inhibitor could reduce cell migration and invasion. luciferase assays confirmed that miR-664 directly bound to the 3'untranslated region of PLP2, and western blotting showed that miR-664 suppressed the expression of PLP2 at the protein levels. This study indicated that miR-664 negatively regulates PLP2 and promotes proliferation and invasion of T-ALL cell lines. Thus, miR-664 may represent a potential therapeutic target for T-ALL intervention. - Highlights: • miR-664 mimics promote the proliferation and invasion of T-ALL cells. • miR-664 inhibitors inhibit the proliferation and invasion of T-ALL cells. • miR-664 targets 3′ UTR of PLP2 in T-ALL cells. • miR-664 negatively regulates PLP2 in T-ALL cells.

ADAR1 attenuates allogeneic graft rejection by suppressing miR-21 biogenesis in macrophages and promoting M2 polarization.

Science.gov (United States)

Li, Junjie; Xie, Jiangang; Liu, Shanshou; Li, Xiao; Zhang, Dongliang; Wang, Xianqi; Jiang, Jinquan; Hu, Wei; Zhang, Yuan; Jin, Boquan; Zhuang, Ran; Yin, Wen

2018-04-25

ADAR1 (adenosine deaminase acting on double-stranded RNA 1) is an RNA-editing enzyme that mediates adenosine-to-inosine RNA editing events, an important post-transcriptional modification mechanism that can alter the coding properties of mRNA or regulate microRNA biogenesis. ADAR1 also regulates the innate immune response. Here, we have demonstrated that ADAR1 expression increased in LPS-stimulated macrophages. Silencing ADAR1 by using small interfering RNA in macrophages resulted in the pronounced polarization of macrophages to M1, whereas ADAR1 overexpression promoted M2 polarization, which indicated that ADAR1 can inhibit macrophage hyperpolarization and prevent immune hyperactivity. The RNA-RNP immunoprecipitation binding assay demonstrated a direct interaction between ADAR1 and miR-21 precursor. Significant up-regulation in IL-10 and down-regulation in miR-21 were observed in ADAR1-overexpressing macrophages. We evaluated miR-21 target mRNAs and macrophage polarization signaling pathways and found that forkhead box protein O1 (Foxo1) was up-regulated in cells that overexpressed ADAR1. In a mouse allogeneic skin transplantation model, grafts in the ADAR1-overexpressed group survived longer and suffered less immune cell infiltration. In ADAR1-overexpressed recipients, splenic macrophages were significantly polarized to M2, and levels of sera IL-10 were markedly higher than those in the control group. In summary, ADAR1 modulates macrophage M2 polarization via the ADAR1-miR-21-Foxo1-IL-10 axis, thereby suppressing allogeneic graft rejection.-Li, J., Xie, J., Liu, S., Li, X., Zhang, D., Wang, X., Jiang, J., Hu, W., Zhang, Y., Jin, B., Zhuang, R., Yin, W. ADAR1 attenuates allogeneic graft rejection by suppressing miR-21 biogenesis in macrophages and promoting M2 polarization.

Novel Action of Carotenoids on Non-Alcoholic Fatty Liver Disease: Macrophage Polarization and Liver Homeostasis.

Science.gov (United States)

Ni, Yinhua; Zhuge, Fen; Nagashimada, Mayumi; Ota, Tsuguhito

2016-06-24

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease. It is characterized by a wide spectrum of hepatic changes, which may progress to non-alcoholic steatohepatitis (NASH) and cirrhosis. NAFLD is considered a hepatic manifestation of metabolic syndrome; however, mechanisms underlying the onset and progression of NAFLD are still unclear. Resident and recruited macrophages are key players in the homeostatic function of the liver and in the progression of NAFLD to NASH. Progress has been made in understanding the molecular mechanisms underlying the polarized activation of macrophages. New NAFLD therapies will likely involve modification of macrophage polarization by restraining M1 activation or driving M2 activation. Carotenoids are potent antioxidants and anti-inflammatory micronutrients that have been used to prevent and treat NAFLD. In addition to their antioxidative action, carotenoids can regulate macrophage polarization and thereby halt the progression of NASH. In this review, we summarize the molecular mechanisms of macrophage polarization and the function of liver macrophages/Kupffer cells in NAFLD. From our review, we propose that dietary carotenoids, such as β-cryptoxanthin and astaxanthin, be used to prevent or treat NAFLD through the regulation of macrophage polarization and liver homeostasis.

The ischemic environment drives microglia and macrophage function

Directory of Open Access Journals (Sweden)

Stefano eFumagalli

2015-04-01

Full Text Available Cells of myeloid origin such as microglia and macrophages act at the crossroads of several inflammatory mechanisms during pathophysiology. Besides pro-inflammatory activity (M1 polarization, myeloid cells acquire protective functions (M2 and participate in the neuroprotective innate mechanisms after brain injury. Experimental research is making considerable efforts to understand the rules that regulate the balance between toxic and protective brain innate immunity. Environmental changes affects microglia/macrophage functions. Hypoxia can affect myeloid cell distribution, activity and phenotype. With their intrinsic differences, microglia and macrophages respond differently to hypoxia, the former depending on ATP to activate, the latter switching to anaerobic metabolism and adapting to hypoxia. Myeloid cell functions include homeostasis control, damage-sensing activity, chemotaxis and phagocytosis, all distinctive features of these cells. Specific markers and morphologies enable to recognize each functional state. To ensure homeostasis and activate when needed, microglia/macrophage physiology is finely tuned. Microglia are controlled by several neuron-derived components, including contact-dependent inhibitory signals and soluble molecules. Changes in this control can cause chronic activation or priming with specific functional consequences. Strategies such as stem cell treatment may enhance microglia protective polarization. This review presents data from the literature that has greatly advanced our understanding of myeloid cell action in brain injury. We discuss the selective responses of microglia and macrophages to hypoxia after stroke and review relevant markers with the aim of defining the different subpopulations of myeloid cells that are recruited to the injured site. We also cover the functional consequences of chronically active microglia and review pivotal works on microglia regulation that offer new therapeutic possibilities for acute

IRAK-M regulates chromatin remodeling in lung macrophages during experimental sepsis.

Directory of Open Access Journals (Sweden)

Kenneth Lyn-Kew

2010-06-01

Full Text Available Sepsis results in a profound state of immunosuppression, which is temporally associated with impaired leukocyte function. The mechanism of leukocyte reprogramming in sepsis is incompletely understood. In this study, we explored mechanisms contributing to dysregulated inflammatory cytokine expression by pulmonary macrophages during experimental sepsis. Pulmonary macrophages (PM recovered from the lungs of mice undergoing cecal ligation and puncture (CLP display transiently reduced expression of some, but not all innate genes in response to LPS. Impaired expression of TNF-alpha and iNOS was associated with reduced acetylation and methylation of specific histones (AcH4 and H3K4me3 and reduced binding of RNA polymerase II to the promoters of these genes. Transient impairment in LPS-induced cytokine responses in septic PM temporally correlated with induction of IRAK-M mRNA and protein, which occurred in a MyD88-dependent fashion. PM isolated from IRAK-M(-/- mice were largely refractory to CLP-induced impairment in cytokine expression, chromatin remodeling, recruitment of RNA polymerase II, and induction of histone deacetylase-2 observed during sepsis. Our findings indicate that systemic sepsis induces epigenetic silencing of cytokine gene expression in lung macrophages, and IRAK-M appears to be a critical mediator of this response.

Differentiation of the endometrial macrophage during pregnancy in the cow.

Directory of Open Access Journals (Sweden)

Lilian J Oliveira

Full Text Available BACKGROUND: The presence of conceptus alloantigens necessitates changes in maternal immune function. One player in this process may be the macrophage. In the cow, there is large-scale recruitment of macrophages expressing CD68 and CD14 to the uterine endometrium during pregnancy. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, the function of endometrial macrophages during pregnancy was inferred by comparison of the transcriptome of endometrial CD14(+ cells isolated from pregnant cows as compared to that of blood CD14(+ cells. The pattern of gene expression was largely similar for CD14(+ cells from both sources, suggesting that cells from both tissues are from the monocyte/macrophage lineage. A total of 1,364 unique genes were differentially expressed, with 680 genes upregulated in endometrial CD14(+ cells as compared to blood CD14(+ cells and with 674 genes downregulated in endometrial CD14(+ cells as compared to blood CD14(+ cells. Twelve genes characteristic of M2 activated macrophages (SLCO2B1, GATM, MRC1, ALDH1A1, PTGS1, RNASE6, CLEC7A, DPEP2, CD163, CCL22, CCL24, and CDH1 were upregulated in endometrial CD14(+ cells. M2 macrophages play roles in immune regulation, tissue remodeling, angiogenesis and apoptosis. Consistent with a role in tissue remodeling, there was over-representation of differentially expressed genes in endometrium for three ontologies related to proteolysis. A role in apoptosis is suggested by the observation that the most overrepresented gene in endometrial CD14(+ cells was GZMA. CONCLUSIONS: Results indicate that at least a subpopulation of endometrial macrophages cells differentiates along an M2 activation pathway during pregnancy and that the cells are likely to play roles in immune regulation, tissue remodeling, angiogenesis, and apoptosis.

Macrophage migration inhibitory factor and autism spectrum disorders

NARCIS (Netherlands)

Grigorenko, Elena L.; Han, Summer S.; Yrigollen, Carolyn M.; Leng, Lin; Mizue, Yuka; Anderson, George M.; Mulder, Erik J.; de Bildt, Annelies; Minderaa, Ruud B.; Volkmar, Fred R.; Chang, Joseph T.; Bucala, Richard

OBJECTIVE. Autistic spectrum disorders are childhood neurodevelopmental disorders characterized by social and communicative impairment and repetitive and stereotypical behavior. Macrophage migration inhibitory factor (MIF) is an upstream regulator of innate immunity that promotes

Regulation of HGF and SDF-1 expression by oral fibroblasts--implications for invasion of oral cancer.

Science.gov (United States)

Daly, Aisling J; McIlreavey, Leanne; Irwin, Chris R

2008-07-01

Invasion and metastasis of oral squamous cell carcinoma (OSCC) is dependent on signals received from stromal fibroblasts present in the surrounding connective tissue. The aim of this study was to investigate the regulation of expression of two important signaling molecules--HGF and SDF-1--by both stromal fibroblasts and their 'activated' form, myofibroblasts, and to determine the role of these two factors in stimulating OSCC cell invasion in vitro. Fibroblasts and myofibroblasts produced similar levels of HGF and SDF-1. IL-1alpha and OSCC cell conditioned medium both stimulated HGF and SDF-1 expression, while TGF-beta(1) inhibited production of each factor. Myofibroblast-derived conditioned medium stimulated OSCC cell invasion through matrigel. Blocking antibodies to both HGF and SDF-1 reduced the level of invasion. In fibroblast-free organotypic raft cultures, addition of HGF and SDF-1 stimulated OSCC cell invasion into the underlying collagen gel, although the pattern of invasion differed from that induced by fibroblasts. Fibroblast-derived HGF and SDF-1 appear to play central roles in the reciprocal interactions between OSCC cells and underlying stromal fibroblasts leading to the local invasion of oral cancer.

CFTR-dependent defect in alternatively-activated macrophages in cystic fibrosis.

Science.gov (United States)

Tarique, Abdullah A; Sly, Peter D; Holt, Patrick G; Bosco, Anthony; Ware, Robert S; Logan, Jayden; Bell, Scott C; Wainwright, Claire E; Fantino, Emmanuelle

2017-07-01

The role of the macrophages in cystic fibrosis (CF) lung disease has been poorly studied. We hypothesized that alternatively activated M2 macrophages are abnormal in CF lung disease. Blood samples were collected from adults (n=13) children (n=27) with CF on admission for acute pulmonary exacerbation and when clinically stable. Monocytes were differentiated into macrophages and polarized into classical (M1) and alternatively-activated (M2) phenotypes, function determined ex-vivo and compared with healthy controls. In the absence of functional cystic fibrosis trans-membrane conductance regulator (CFTR), either naturally in patients with CF or induced with CFTR inhibitors, monocyte-derived macrophages do not respond to IL-13/IL-4, fail to polarize into M2s associated with a post-transcriptional failure to produce and express IL-13Rα1 on the macrophage surface Polarization to the M1 phenotype was unaffected. CFTR-dependent imbalance of macrophage phenotypes and functions could contribute to the exaggerated inflammatory response seen in CF lung disease. Copyright © 2017 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.

Proteomic Analysis Reveals Distinct Metabolic Differences Between Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) and Macrophage Colony Stimulating Factor (M-CSF) Grown Macrophages Derived from Murine Bone Marrow Cells.

Science.gov (United States)

Na, Yi Rang; Hong, Ji Hye; Lee, Min Yong; Jung, Jae Hun; Jung, Daun; Kim, Young Won; Son, Dain; Choi, Murim; Kim, Kwang Pyo; Seok, Seung Hyeok

2015-10-01

Macrophages are crucial in controlling infectious agents and tissue homeostasis. Macrophages require a wide range of functional capabilities in order to fulfill distinct roles in our body, one being rapid and robust immune responses. To gain insight into macrophage plasticity and the key regulatory protein networks governing their specific functions, we performed quantitative analyses of the proteome and phosphoproteome of murine primary GM-CSF and M-CSF grown bone marrow derived macrophages (GM-BMMs and M-BMMs, respectively) using the latest isobaric tag based tandem mass tag (TMT) labeling and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Strikingly, metabolic processes emerged as a major difference between these macrophages. Specifically, GM-BMMs show significant enrichment of proteins involving glycolysis, the mevalonate pathway, and nitrogen compound biosynthesis. This evidence of enhanced glycolytic capability in GM-BMMs is particularly significant regarding their pro-inflammatory responses, because increased production of cytokines upon LPS stimulation in GM-BMMs depends on their acute glycolytic capacity. In contrast, M-BMMs up-regulate proteins involved in endocytosis, which correlates with a tendency toward homeostatic functions such as scavenging cellular debris. Together, our data describes a proteomic network that underlies the pro-inflammatory actions of GM-BMMs as well as the homeostatic functions of M-BMMs. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Metabolic reprogramming through fatty acid transport protein 1 (FATP1 regulates macrophage inflammatory potential and adipose inflammation

Directory of Open Access Journals (Sweden)

Amy R. Johnson

2016-07-01

Full Text Available Objective: A novel approach to regulate obesity-associated adipose inflammation may be through metabolic reprogramming of macrophages (MΦs. Broadly speaking, MΦs dependent on glucose are pro-inflammatory, classically activated MΦs (CAM, which contribute to adipose inflammation and insulin resistance. In contrast, MΦs that primarily metabolize fatty acids are alternatively activated MΦs (AAM and maintain tissue insulin sensitivity. In actuality, there is much flexibility and overlap in the CAM-AAM spectrum in vivo dependent upon various stimuli in the microenvironment. We hypothesized that specific lipid trafficking proteins, e.g. fatty acid transport protein 1 (FATP1, would direct MΦ fatty acid transport and metabolism to limit inflammation and contribute to the maintenance of adipose tissue homeostasis. Methods: Bone marrow derived MΦs (BMDMs from Fatp1−/− and Fatp1+/+ mice were used to investigate FATP1-dependent substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. We also generated C57BL/6J chimeric mice by bone marrow transplant specifically lacking hematopoetic FATP1 (Fatp1B−/− and controls Fatp1B+/+. Mice were challenged by high fat diet (HFD or low fat diet (LFD and analyses including MRI, glucose and insulin tolerance tests, flow cytometric, histologic, and protein quantification assays were conducted. Finally, an FATP1-overexpressing RAW 264.7 MΦ cell line (FATP1-OE and empty vector control (FATP1-EV were developed as a gain of function model to test effects on substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. Results: Fatp1 is downregulated with pro-inflammatory stimulation of MΦs. Fatp1−/− BMDMs and FATP1-OE RAW 264.7 MΦs demonstrated that FATP1 reciprocally controled metabolic flexibility, i.e. lipid and glucose metabolism, which was associated with inflammatory response. Supporting our previous work demonstrating the positive relationship between glucose

The Flavonoid Quercetin Ameliorates Liver Inflammation and Fibrosis by Regulating Hepatic Macrophages Activation and Polarization in Mice

Directory of Open Access Journals (Sweden)

Xi Li

2018-02-01

Full Text Available At present, there are no effective antifibrotic drugs for patients with chronic liver disease; hence, the development of antifibrotic therapies is urgently needed. Here, we performed an experimental and translational study to investigate the potential and underlying mechanism of quercetin treatment in liver fibrosis, mainly focusing on the impact of quercetin on macrophages activation and polarization. BALB/c mice were induced liver fibrosis by carbon tetrachloride (CCl4 for 8 weeks and concomitantly treated with quercetin (50 mg/kg or vehicle by daily gavage. Liver inflammation, fibrosis, and hepatic stellate cells (HSCs activation were examined. Moreover, massive macrophages accumulation, M1 macrophages and their related markers, such as tumor necrosis factor (TNF-α, interleukin (IL-1β, IL-6, and monocyte chemotactic protein-1 (MCP-1 in livers were analyzed. In vitro, we used Raw 264.7 cells to examine the effect of quercetin on M1-polarized macrophages activation. Our results showed that quercetin dramatically ameliorated liver inflammation, fibrosis, and inhibited HSCs activation. These results were attributed to the reductive recruitment of macrophages (F4/80+ and CD68+ into the liver in quercetin-treated fibrotic mice confirmed by immunostaining and expression levels of marker molecules. Importantly, quercetin strongly inhibited M1 polarization and M1-related inflammatory cytokines in fibrotic livers when compared with vehicle-treated mice. In vitro, studies further revealed that quercetin efficiently inhibited macrophages activation and M1 polarization, as well as decreased the mRNA expression of M1 macrophage markers such as TNF-α, IL-1β, IL-6, and nitric oxide synthase 2. Mechanistically, the inhibition of M1 macrophages by quercetin was associated with the decreased levels of Notch1 expression on macrophages both in vivo and in vitro. Taken together, our data indicated that quercetin attenuated CCl4-induced liver inflammation and

Human immunodeficiency virus impairs reverse cholesterol transport from macrophages.

Directory of Open Access Journals (Sweden)

Zahedi Mujawar

2006-10-01

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

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

Science.gov (United States)

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

2015-08-01

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

Obesity, expression of adipocytokines, and macrophage infiltration in canine mammary tumors.

Science.gov (United States)

Lim, H Y; Im, K S; Kim, N H; Kim, H W; Shin, J I; Sur, J H

2015-03-01

Obesity influences the development, progression and prognosis of human breast cancer and canine mammary cancer (MC) but the precise underlying mechanism is not well-documented in the fields of either human or veterinary oncology. In the present study, the expression of major adipocytokines, including leptin, adiponectin, and leptin receptor (ObR) in benign (n = 28) and malignant (n = 70) canine mammary tumors was investigated by immunohistochemistry and on the basis of the subject's body condition score (BCS). To evaluate the relationship between obesity and chronic inflammation of the mammary gland, macrophages infiltrating within and around tumoral areas were counted. The mean age of MC development was lower in overweight or obese dogs (9.0 ± 1.8 years) than in lean dogs or optimal bodyweight (10.2 ± 2.9 years), and the evidence of lymphatic invasion of carcinoma cells was found more frequently in overweight or obese group than in lean or optimal groups. Decreased adiponectin expression and increased macrophage numbers in overweight or obese subjects were significantly correlated with factors related to a poor prognosis, such as high histological grade and lymphatic invasion. Leptin expression was correlated with progesterone receptor status, and ObR expression was correlated with estrogen receptor status of MCs, regardless of BCS. Macrophage infiltration within and around the tumor may play an important role in tumor progression and metastasis in obese female dogs and may represent a prognostic factor for canine MCs. Copyright © 2015 Elsevier Ltd. All rights reserved.

Clonorchis sinensis antigens alter hepatic macrophage polarization in vitro and in vivo.

Directory of Open Access Journals (Sweden)

Eun-Min Kim

2017-05-01

Full Text Available Clonorchis sinensis infection elicits hepatic inflammation, which can lead to cholangitis, periductal hepatic fibrosis, liver cirrhosis, and even cholangiocarcinoma. Hepatic macrophages are an intrinsic element of both innate and acquired immunity. This study was conducted to demonstrate the dynamics of hepatic macrophage polarization during C. sinensis infection in mice and to identify factors regulating this polarization. Treatment of hepatic macrophages isolated from normal mice with C. sinensis excretory/secretory products (ESPs resulted in the preferential generation of classically activated hepatic macrophages (M1 macrophages and the production of pro-inflammatory cytokines. Additionally, cells stimulated with C. sinensis ESPs exhibited changes in cellular morphology. During the early stages of C. sinensis infection, hepatic macrophages preferentially differentiated into M1 macrophages; however, during the C. sinensis mature worm stage, when eggs are released, there were significant increases in the abundance of both M1 macrophages and alternatively activated hepatic macrophages (M2 macrophages. Moreover, there was a further increase in the M2 macrophage count during the fibrotic and cirrhotic stage of infection. Notably, this fibrotic and cirrhotic stage promoted a strong increase in the proportion of Arg-1-producing macrophages (M2 phenotype, which were associated with fibrosis and tissue repair in the liver. Our results suggest that the dynamic polarization of hepatic macrophages as C. sinensis infection progresses is related to the histological lesions present in liver tissue. Hepatic macrophages thus play an important role in local immunity during C. sinensis infection.

Clonorchis sinensis antigens alter hepatic macrophage polarization in vitro and in vivo.

Science.gov (United States)

Kim, Eun-Min; Kwak, You Shine; Yi, Myung-Hee; Kim, Ju Yeong; Sohn, Woon-Mok; Yong, Tai-Soon

2017-05-01

Clonorchis sinensis infection elicits hepatic inflammation, which can lead to cholangitis, periductal hepatic fibrosis, liver cirrhosis, and even cholangiocarcinoma. Hepatic macrophages are an intrinsic element of both innate and acquired immunity. This study was conducted to demonstrate the dynamics of hepatic macrophage polarization during C. sinensis infection in mice and to identify factors regulating this polarization. Treatment of hepatic macrophages isolated from normal mice with C. sinensis excretory/secretory products (ESPs) resulted in the preferential generation of classically activated hepatic macrophages (M1 macrophages) and the production of pro-inflammatory cytokines. Additionally, cells stimulated with C. sinensis ESPs exhibited changes in cellular morphology. During the early stages of C. sinensis infection, hepatic macrophages preferentially differentiated into M1 macrophages; however, during the C. sinensis mature worm stage, when eggs are released, there were significant increases in the abundance of both M1 macrophages and alternatively activated hepatic macrophages (M2 macrophages). Moreover, there was a further increase in the M2 macrophage count during the fibrotic and cirrhotic stage of infection. Notably, this fibrotic and cirrhotic stage promoted a strong increase in the proportion of Arg-1-producing macrophages (M2 phenotype), which were associated with fibrosis and tissue repair in the liver. Our results suggest that the dynamic polarization of hepatic macrophages as C. sinensis infection progresses is related to the histological lesions present in liver tissue. Hepatic macrophages thus play an important role in local immunity during C. sinensis infection.

Mena invasive (MenaINV) promotes multicellular streaming motility and transendothelial migration in a mouse model of breast cancer.

Science.gov (United States)

Roussos, Evanthia T; Balsamo, Michele; Alford, Shannon K; Wyckoff, Jeffrey B; Gligorijevic, Bojana; Wang, Yarong; Pozzuto, Maria; Stobezki, Robert; Goswami, Sumanta; Segall, Jeffrey E; Lauffenburger, Douglas A; Bresnick, Anne R; Gertler, Frank B; Condeelis, John S

2011-07-01

We have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (Mena(INV)) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated Mena(INV) increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by Mena(INV) is dependent on a macrophage-tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of Mena(INV) and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells.

Biological role of granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) on cells of the myeloid lineage

Science.gov (United States)

Ushach, Irina; Zlotnik, Albert

2016-01-01

M-CSF and GM-CSF are 2 important cytokines that regulate macrophage numbers and function. Here, we review their known effects on cells of the macrophage-monocyte lineage. Important clues to their function come from their expression patterns. M-CSF exhibits a mostly homeostatic expression pattern, whereas GM-CSF is a product of cells activated during inflammatory or pathologic conditions. Accordingly, M-CSF regulates the numbers of various tissue macrophage and monocyte populations without altering their "activation" status. Conversely, GM-CSF induces activation of monocytes/macrophages and also mediates differentiation to other states that participate in immune responses [i.e., dendritic cells (DCs)]. Further insights into their function have come from analyses of mice deficient in either cytokine. M-CSF signals through its receptor (CSF-1R). Interestingly, mice deficient in CSF-1R expression exhibit a more significant phenotype than mice deficient in M-CSF. This observation was explained by the discovery of a novel cytokine (IL-34) that represents a second ligand of CSF-1R. Information about the function of these ligands/receptor system is still developing, but its complexity is intriguing and strongly suggests that more interesting biology remains to be elucidated. Based on our current knowledge, several therapeutic molecules targeting either the M-CSF or the GM-CSF pathways have been developed and are currently being tested in clinical trials targeting either autoimmune diseases or cancer. It is intriguing to consider how evolution has directed these pathways to develop; their complexity likely mirrors the multiple functions in which cells of the monocyte/macrophage system are involved. PMID:27354413

Characterization of the growth-inhibitory and apoptosis-inducing activities of a triterpene saponin, securioside B against BAC1.2F5 macrophages

Directory of Open Access Journals (Sweden)

Satoru Yui

2003-01-01

Full Text Available Background: Since the growth state of macrophages in local pathological sites is considered a factor that regulates the processes of many disease, such as tumors, inflammation, and atherosclerosis, the substances that regulate macrophage growth or survival may be useful for disease control. We previously reported that securiosides A and B, novel triterpene saponins, exerted macrophage-oriented cytotoxicity in the presence of a L-cell-conditioned medium containing macrophage colony-stimulating factor (M-CSF, while the compounds did not exhibit an effect on macrophages in the absence of the growth-stimulating factors.

The MARVEL domain protein Nce102 regulates actin organization and invasive growth of Candida albicans.

Science.gov (United States)

Douglas, Lois M; Wang, Hong X; Konopka, James B

2013-11-26

Invasive growth of the fungal pathogen Candida albicans into tissues promotes disseminated infections in humans. The plasma membrane is essential for pathogenesis because this important barrier mediates morphogenesis and invasive growth, as well as secretion of virulence factors, cell wall synthesis, nutrient import, and other processes. Previous studies showed that the Sur7 tetraspan protein that localizes to MCC (membrane compartment occupied by Can1)/eisosome subdomains of the plasma membrane regulates a broad range of key functions, including cell wall synthesis, morphogenesis, and resistance to copper. Therefore, a distinct tetraspan protein found in MCC/eisosomes, Nce102, was investigated. Nce102 belongs to the MARVEL domain protein family, which is implicated in regulating membrane structure and function. Deletion of NCE102 did not cause the broad defects seen in sur7Δ cells. Instead, the nce102Δ mutant displayed a unique phenotype in that it was defective in forming hyphae and invading low concentrations of agar but could invade well in higher agar concentrations. This phenotype was likely due to a defect in actin organization that was observed by phalloidin staining. In support of this, the invasive growth defect of a bni1Δ mutant that mislocalizes actin due to lack of the Bni1 formin was also reversed at high agar concentrations. This suggests that a denser matrix provides a signal that compensates for the actin defects. The nce102Δ mutant displayed decreased virulence and formed abnormal hyphae in mice. These studies identify novel ways that Nce102 and the physical environment surrounding C. albicans regulate morphogenesis and pathogenesis. The plasma membrane promotes virulence of the human fungal pathogen Candida albicans by acting as a protective barrier around the cell and mediating dynamic activities, such as morphogenesis, cell wall synthesis, secretion of virulence factors, and nutrient uptake. To better understand how the plasma membrane

Growth of Listeria monocytogenes on a RTE-meat matrix enhances cell invasiveness to mouse J774A.1 macrophages.

Science.gov (United States)

Lin, Chen-Si; Wang, Chinling; Tsai, Hsiang-Jung; Chou, Chung-Hsi

2010-11-15

It remains unclear whether the growth of Listeria monocytogenes on a ready-to-eat (RTE) meat matrix has an impact on the bacterium's pathogenic abilities. In this study, we investigated the impact of environments on virulence by growing L. monocytogenes (F2365 strain) on brain heart infusion agar (BHI), tryptic soy agar (TSA), and RTE turkey meat matrices. Bacteria cultured from these media were harvested and used to infect mouse macrophage cell line J774A.1 with different MOIs to examine their invasion ability. At MOI=10 and 50, the numbers of bacteria recovered from cells infected with turkey-meat-grown Listeria were significantly higher than those from the two nutrient-rich growth media. Additionally, MOI played a role in determining L. monocytogenes recovery rates, since significant differences were found amongst all three groups at low MOI, while no significant differences were found between BHI and TSA groups at high MOI. These results indicate that environmental changes affect the ability of L. monocytogenes to invade and survive intracellularly while grown on RTE-meat matrix. Copyright © 2010 Elsevier B.V. All rights reserved.

CD147, CD44, and the epidermal growth factor receptor (EGFR) signaling pathway cooperate to regulate breast epithelial cell invasiveness.

Science.gov (United States)

Grass, G Daniel; Tolliver, Lauren B; Bratoeva, Momka; Toole, Bryan P

2013-09-06

The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777-788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer.

CD147, CD44, and the Epidermal Growth Factor Receptor (EGFR) Signaling Pathway Cooperate to Regulate Breast Epithelial Cell Invasiveness*

Science.gov (United States)

Grass, G. Daniel; Tolliver, Lauren B.; Bratoeva, Momka; Toole, Bryan P.

2013-01-01

The immunoglobulin superfamily glycoprotein CD147 (emmprin; basigin) is associated with an invasive phenotype in various types of cancers, including malignant breast cancer. We showed recently that up-regulation of CD147 in non-transformed, non-invasive breast epithelial cells is sufficient to induce an invasive phenotype characterized by membrane type-1 matrix metalloproteinase (MT1-MMP)-dependent invadopodia activity (Grass, G. D., Bratoeva, M., and Toole, B. P. (2012) Regulation of invadopodia formation and activity by CD147. J. Cell Sci. 125, 777–788). Here we found that CD147 induces breast epithelial cell invasiveness by promoting epidermal growth factor receptor (EGFR)-Ras-ERK signaling in a manner dependent on hyaluronan-CD44 interaction. Furthermore, CD147 promotes assembly of signaling complexes containing CD147, CD44, and EGFR in lipid raftlike domains. We also found that oncogenic Ras regulates CD147 expression, hyaluronan synthesis, and formation of CD147-CD44-EGFR complexes, thus forming a positive feedback loop that may amplify invasiveness. Last, we showed that malignant breast cancer cells are heterogeneous in their expression of surface-associated CD147 and that high levels of membrane CD147 correlate with cell surface EGFR and CD44 levels, activated EGFR and ERK1, and activated invadopodia. Future studies should evaluate CD147 as a potential therapeutic target and disease stratification marker in breast cancer. PMID:23888049

Molecular Characterization of Macrophage-Biomaterial Interactions.

Science.gov (United States)

Moore, Laura Beth; Kyriakides, Themis R

2015-01-01

Implantation of biomaterials in vascularized tissues elicits the sequential engagement of molecular and cellular elements that constitute the foreign body response. Initial events include the non-specific adsorption of proteins to the biomaterial surface that render it adhesive for cells such as neutrophils and macrophages. The latter undergo unique activation and in some cases undergo cell-cell fusion to form foreign body giant cells that contribute to implant damage and fibrotic encapsulation. In this review, we discuss the molecular events that contribute to macrophage activation and fusion with a focus on the role of the inflammasome, signaling pathways such as JAK/STAT and NF-κB, and the putative involvement of micro RNAs in the regulation of these processes.

Isoginkgetin inhibits tumor cell invasion by regulating phosphatidylinositol 3-kinase/Akt-dependent matrix metalloproteinase-9 expression.

Science.gov (United States)

Yoon, Sang-Oh; Shin, Sejeong; Lee, Ho-Jae; Chun, Hyo-Kon; Chung, An-Sik

2006-11-01

Matrix metalloproteinase (MMP)-9 plays a key role in tumor invasion. Inhibitors of MMP-9 were screened from Metasequoia glyptostroboides (Dawn redwood) and one potent inhibitor, isoginkgetin, a biflavonoid, was identified. Noncytotoxic levels of isoginkgetin decreased MMP-9 production profoundly, but up-regulated the level of tissue inhibitor of metalloproteinase (TIMP)-1, an inhibitor of MMP-9, in HT1080 human fibrosarcoma cells. The major mechanism of Ras-dependent MMP-9 production in HT1080 cells was phosphatidylinositol 3-kinase (PI3K)/Akt/nuclear factor-kappaB (NF-kappaB) activation. Expression of dominant-active H-Ras and p85 (a subunit of PI3K) increased MMP-9 activity, whereas dominant-negative forms of these molecules decreased the level of MMP-9. H-Ras did not increase MMP-9 in the presence of a PI3K inhibitor, LY294002, and a NF-kappaB inhibitor, SN50. Further studies showed that isoginkgetin regulated MMP-9 production via PI3K/Akt/NF-kappaB pathway, as evidenced by the findings that isoginkgetin inhibited activities of both Akt and NF-kappaB. PI3K/Akt is a well-known key pathway for cell invasion, and isoginkgetin inhibited HT1080 tumor cell invasion substantially. Isoginkgetin was also quite effective in inhibiting the activities of Akt and MMP-9 in MDA-MB-231 breast carcinomas and B16F10 melanoma. Moreover, isoginkgetin treatment resulted in marked decrease in invasion of these cells. In summary, PI3K/Akt is a major pathway for MMP-9 expression and isoginkgetin markedly decreased MMP-9 expression and invasion through inhibition of this pathway. This suggests that isoginkgetin could be a potential candidate as a therapeutic agent against tumor invasion.

Immune Evasion Strategies of Pathogens in Macrophages: the Potential for Limiting Pathogen Transmission.

Science.gov (United States)

Ren, Yuwei; Khan, Faheem Ahmed; Pandupuspitasari, Nuruliarizki Shinta; Zhang, Shujun

2017-01-01

Preventing pathogen transmission to a new host is of major interest to the immunologist and could benefit from a detailed investigation of pathogen immune evasion strategies. The first line of defense against pathogen invasion is provided by macrophages. When they sense pathogens, macrophages initiate signals to inflammatory and pro-inflammatory cytokines through pattern recognition receptors (PRRs) subsequently mediating phagocytosis and inflammation. The macrophage immune machinery classically includes two subsets: the activated M1 and the activated M2 that respond accordingly in diverse immune challenges. The lipid and glycogen metabolic pathways work together with the lysosome to help the mature phagosome to degrade and eliminate intracellular pathogens in macrophages. The viral evasion strategies are even more complex due to the interplay between autophagy and apoptosis. However, pathogens evolve several strategies to camouflage themselves against immune responses in order to ensure their survival, replication and transmission. These strategies include the muting of PRRs initiated inflammatory responses, attenuation of M1 and/or induction of M2 macrophages, suppression of autophago-lysosomal formation, interference with lipid and glycogen metabolism, and viral mediation of autophagy and apoptosis cross-talk to enhance viral replication. This review focuses on pathogen immune evasion methods and on the strategies used by the host against camouflaged pathogens.

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

Science.gov (United States)

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.

Collaborating with the Enemy: Function of Macrophages in the Development of Neoplastic Disease

Directory of Open Access Journals (Sweden)

Andrzej Eljaszewicz

2013-01-01

Full Text Available 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.

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

Science.gov (United States)

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

2016-04-01

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

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

Directory of Open Access Journals (Sweden)

Sredni Benjamin

2010-01-01

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

Inflammation and ER Stress Downregulate BDH2 Expression and Dysregulate Intracellular Iron in Macrophages

Directory of Open Access Journals (Sweden)

Susu M. Zughaier

2014-01-01

Full Text Available Macrophages play a very important role in host defense and in iron homeostasis by engulfing senescent red blood cells and recycling iron. Hepcidin is the master iron regulating hormone that limits dietary iron absorption from the gut and limits iron egress from macrophages. Upon infection macrophages retain iron to limit its bioavailability which limits bacterial growth. Recently, a short chain butyrate dehydrogenase type 2 (BDH2 protein was reported to contain an iron responsive element and to mediate cellular iron trafficking by catalyzing the synthesis of the mammalian siderophore that binds labile iron; therefore, BDH2 plays a crucial role in intracellular iron homeostasis. However, BDH2 expression and regulation in macrophages have not yet been described. Here we show that LPS-induced inflammation combined with ER stress led to massive BDH2 downregulation, increased the expression of ER stress markers, upregulated hepcidin expression, downregulated ferroportin expression, caused iron retention in macrophages, and dysregulated cytokine release from macrophages. We also show that ER stress combined with inflammation synergistically upregulated the expression of the iron carrier protein NGAL and the stress-inducible heme degrading enzyme heme oxygenase-1 (HO-1 leading to iron liberation. This is the first report to show that inflammation and ER stress downregulate the expression of BDH2 in human THP-1 macrophages.

MicroRNA-223 Is Upregulated in Active Tuberculosis Patients and Inhibits Apoptosis of Macrophages by Targeting FOXO3.

Science.gov (United States)

Xi, Xiue; Zhang, Chunxiao; Han, Wei; Zhao, Huayang; Zhang, Huiqiang; Jiao, Junhua

2015-12-01

Macrophage apoptosis is a host innate defense mechanism against tuberculosis (TB). In this study, we aimed to investigate the role of microRNA-223 (miR-223) in macrophage apoptosis of TB. We analyzed apoptosis in peripheral blood macrophages of active TB patients, infected human macrophages (TDMs and MDMs) with the Mycobacterium tuberculosis (Mtb) strain H37Rv, and observed the expression of miR-223 to investigate the relationship between miR-223 and macrophage apoptosis induced by Mtb. The apoptosis rate of peripheral blood macrophages decreased in active TB patients compared with healthy controls, and miR-223 expression increased significantly in macrophages after H37Rv infection. Transfection of human macrophages (TDMs and MDMs) with miR-223 inhibited macrophage apoptosis. We also demonstrated that miR-223 directly suppressed forkhead box O3 (FOXO3), and FOXO3 played a critical role as a mediator of the biological effects of miR-223 in macrophage apoptosis. The overexpression of FOXO3 remarkably reversed the apoptosis inhibitory effect of miR-223. Our data provide new clues for the essential role of miR-223 in the regulation of anti-Mtb-directed immune responses, which relies on the regulation of FOXO3 expression.

Tolerance of monocytes and macrophages in response to bacterial endotoxin

Directory of Open Access Journals (Sweden)

Ewelina Wiśnik

2017-03-01

Full Text Available Monocytes belong to myeloid effector cells, which constitute the first line of defense against pathogens, also called the nonspecific immune system and play an important role in the maintenance of tissue homeostasis. In response to stimulation, monocytes differentiate into macrophages capable of microorganism phagocytosis and secrete factors that play a key role in the regulation of immune responses. However excessive exposure of monocytes/macrophages to the lipopolysaccharide (LPS of Gram negative bacteria leads to the acquisition of immune tolerance by these cells. Such state results from disruption of different biological processes, for example intracellular signaling pathways and is accompanied by a number of disease states (immune, inflammatory or neoplastic conditions. Regulation of monocytes/macrophages activity is controlled by miRNAs, which are involved in the modulation of immune tolerance acquired by these cells. Moreover, the tolerance to endotoxin is conditioned by the posttranscriptional processes and posttranslational epigenetic modifications leading to the impairment of normal immune response for example by alterations in the expression of many genes encoding immune signaling mediators. The aim of this paper is to provide an overview existing knowledge on the modulation of activity of monocytes/macrophages in response to bacterial endotoxin and impaired immune responses.

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

Directory of Open Access Journals (Sweden)

Marien Willem Johan Adriaan Fieren

2012-01-01

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

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

Science.gov (United States)

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.

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

Lifescience Database Archive (English)

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

Cathepsin E deficiency impairs autophagic proteolysis in macrophages.

Directory of Open Access Journals (Sweden)

Takayuki Tsukuba

Full Text Available Cathepsin E is an endosomal aspartic proteinase that is predominantly expressed in immune-related cells. Recently, we showed that macrophages derived from cathepsin E-deficient (CatE(-/- mice display accumulation of lysosomal membrane proteins and abnormal membrane trafficking. In this study, we demonstrated that CatE(-/- macrophages exhibit abnormalities in autophagy, a bulk degradation system for aggregated proteins and damaged organelles. CatE(-/- macrophages showed increased accumulation of autophagy marker proteins such as LC3 and p62, and polyubiquitinated proteins. Cathepsin E deficiency also altered autophagy-related signaling pathways such as those mediated by the mammalian target of rapamycin (mTOR, Akt, and extracellular signal-related kinase (ERK. Furthermore, immunofluorescence microscopy analyses showed that LC3-positive vesicles were merged with acidic compartments in wild-type macrophages, but not in CatE(-/- macrophages, indicating inhibition of fusion of autophagosome with lysosomes in CatE(-/- cells. Delayed degradation of LC3 protein was also observed under starvation-induced conditions. Since the autophagy system is involved in the degradation of damaged mitochondria, we examined the accumulation of damaged mitochondria in CatE(-/- macrophages. Several mitochondrial abnormalities such as decreased intracellular ATP levels, depolarized mitochondrial membrane potential, and decreased mitochondrial oxygen consumption were observed. Such mitochondrial dysfunction likely led to the accompanying oxidative stress. In fact, CatE(-/- macrophages showed increased reactive oxygen species (ROS production and up-regulation of oxidized peroxiredoxin-6, but decreased antioxidant glutathione. These results indicate that cathepsin E deficiency causes autophagy impairment concomitantly with increased aberrant mitochondria as well as increased oxidative stress.

Inhibiting epigenetic enzymes to improve atherogenic macrophage functions

NARCIS (Netherlands)

van den Bossche, Jan; Neele, Annette E.; Hoeksema, Marten A.; de Heij, Femke; Boshuizen, Marieke C. S.; van der Velden, Saskia; de Boer, Vincent C.; Reedquist, Kris A.; de Winther, Menno P. J.

2014-01-01

Macrophages determine the outcome of atherosclerosis by propagating inflammatory responses, foam cell formation and eventually necrotic core development. Yet, the pathways that regulate their atherogenic functions remain ill-defined. It is now apparent that chromatin remodeling chromatin modifying

The role of medicaments, exosomes and miRNA molecules in modulation of macrophage immune activity

Directory of Open Access Journals (Sweden)

Katarzyna Nazimek

2015-01-01

Full Text Available Macrophages play an important role in innate immunity, in induction and orchestration of acquired immune response as well as in the maintenance of tissue homeostasis. Macrophages as antigen presenting cells induce or inhibit the development of immune response and as effector cells play an important role in innate immunity to infectious agents and in delayed--type hypersensitivity as well. Thus, either up- or down-regulation of their activity leads to the impairment of different biological processes. This often results in the development of immunological diseases or inflammatory response associated with metabolic, cardiovascular or neuroendocrine disorders. Therefore, the possibility of modulation of macrophage function should allow for elaboration of new effective therapeutic strategies. Noteworthy, interaction of medicaments with macrophages may directly mediate their therapeutic activity or is an additional beneficial effect increasing efficacy of treatment. Further, macrophage differentiation is regulated by miRNA-223, while expression of miRNA-146 and miRNA-155 may modulate and/or be a result of the current cell phenotype. Present review is focused on the current knowledge about the action of medicaments, microRNA molecules, exosomes and related vesicles on macrophages leading to modulation of their biological activity.

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

Science.gov (United States)

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

2014-11-07

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

Macrophages are necessary for epimorphic regeneration in African spiny mice.

Science.gov (United States)

Simkin, Jennifer; Gawriluk, Thomas R; Gensel, John C; Seifert, Ashley W

2017-05-16

How the immune system affects tissue regeneration is not well understood. In this study, we used an emerging mammalian model of epimorphic regeneration, the African spiny mouse, to examine cell-based inflammation and tested the hypothesis that macrophages are necessary for regeneration. By directly comparing inflammatory cell activation in a 4 mm ear injury during regeneration ( Acomys cahirinus ) and scarring ( Mus musculus ), we found that both species exhibited an acute inflammatory response, with scarring characterized by stronger myeloperoxidase activity. In contrast, ROS production was stronger and more persistent during regeneration. By depleting macrophages during injury, we demonstrate a functional requirement for these cells to stimulate regeneration. Importantly, the spatial distribution of activated macrophage subtypes was unique during regeneration with pro-inflammatory macrophages failing to infiltrate the regeneration blastema. Together, our results demonstrate an essential role for inflammatory cells to regulate a regenerative response.

Erythropoietin protects against rhabdomyolysis-induced acute kidney injury by modulating macrophage polarization

Science.gov (United States)

Wang, Shuo; Zhang, Chao; Li, Jiawei; Niyazi, Sidikejiang; Zheng, Long; Xu, Ming; Rong, Ruiming; Yang, Cheng; Zhu, Tongyu

2017-01-01

Erythropoietin (EPO) is a well-known hormone that is clinically used for the treatment of anemia. Very recently, an increasing body of evidence showed that EPO could still regulate bioactivities of macrophages. However, the details about the immunomodulatory effect of EPO on macrophages are not fully delineated, particularly in the setting of renal damages. Therefore, in the present study, we determined whether EPO could exert an impact on the dynamics of macrophages in a well-established model of rhabdomyolysis-induced acute kidney injury and explored the potential mechanisms. EPO was found to ameliorate kidney injuries by reducing macrophages recruitment and promoting phenotype switch toward M2 macrophages in vivo. It was also confirmed that EPO could directly suppress pro-inflammatory responses of M1 macrophages and promote M2 marker expression in vitro. Data indicated the possible involvement of Jak2/STAT3/STAT6 pathway in the augmentation of EPO on M2 polarization. These results improved the understanding of the immunoregulatory capacity of EPO on macrophages, which might optimize the therapeutic modalities of EPO. PMID:28383559

MicroRNA 27a-3p Regulates Antimicrobial Responses of Murine Macrophages Infected by Mycobacterium avium subspecies paratuberculosis by Targeting Interleukin-10 and TGF-β-Activated Protein Kinase 1 Binding Protein 2

Directory of Open Access Journals (Sweden)

Tariq Hussain

2018-01-01

Full Text Available Mycobacterium avium subspecies paratuberculosis (MAP persistently survive and replicate in mononuclear phagocytic cells by adopting various strategies to subvert host immune response. Interleukin-10 (IL-10 upregulation via inhibition of macrophage bactericidal activity is a critical step for MAP survival and pathogenesis within the host cell. Mitogen-activated protein kinase p38 signaling cascade plays a crucial role in the elevation of IL-10 and progression of MAP pathogenesis. The contribution of microRNAs (miRNAs and their influence on the activation of macrophages during MAP pathogenesis are still unclear. In the current study, we found that miRNA-27a-3p (miR-27a expression is downregulated during MAP infection both in vivo and in vitro. Moreover, miR-27a is also downregulated in toll-like receptor 2 (TLR2-stimulated murine macrophages (RAW264.7 and bone marrow-derived macrophage. ELISA and real-time qRT-PCR results confirm that overexpression of miR-27a inhibited MAP-induced IL-10 production in macrophages and upregulated pro-inflammatory cytokines, while miR-27a inhibitor counteracted these effects. Luciferase reporter assay results revealed that IL-10 and TGF-β-activated protein kinase 1 binding protein 2 (TAB 2 are potential targets of miR-27a. In addition, we demonstrated that miR-27a negatively regulates TAB 2 expression and diminishes TAB 2-dependent p38/JNK phosphorylation, ultimately downregulating IL-10 expression in MAP-infected macrophages. Furthermore, overexpression of miR-27a significantly inhibited the intracellular survival of MAP in infected macrophages. Our data show that miR-27a augments antimicrobial activities of macrophages and inhibits the expression of IL-10, demonstrating that miR-27a regulates protective innate immune responses during MAP infection and can be exploited as a novel therapeutic target in the control of intracellular pathogens, including paratuberculosis.

Proline-rich tyrosine kinase 2 (Pyk2 regulates IGF-I-induced cell motility and invasion of urothelial carcinoma cells.

Directory of Open Access Journals (Sweden)

Marco Genua

Full Text Available The insulin-like growth factor receptor I (IGF-IR plays an essential role in transformation by promoting cell growth and protecting cancer cells from apoptosis. We have recently demonstrated that the IGF-IR is overexpressed in invasive bladder cancer tissues and promotes motility and invasion of urothelial carcinoma cells. These effects require IGF-I-induced Akt- and MAPK-dependent activation of paxillin. The latter co-localizes with focal adhesion kinases (FAK at dynamic focal adhesions and is critical for promoting motility of urothelial cancer cells. FAK and its homolog Proline-rich tyrosine kinase 2 (Pyk2 modulate paxillin activation; however, their role in regulating IGF-IR-dependent signaling and motility in bladder cancer has not been established. In this study we demonstrate that FAK was not required for IGF-IR-dependent signaling and motility of invasive urothelial carcinoma cells. On the contrary, Pyk2, which was strongly activated by IGF-I, was critical for IGF-IR-dependent motility and invasion and regulated IGF-I-dependent activation of the Akt and MAPK pathways. Using immunofluorescence and AQUA analysis we further discovered that Pyk2 was overexpressed in bladder cancer tissues as compared to normal tissue controls. Significantly, in urothelial carcinoma tissues there was increased Pyk2 localization in the nuclei as compared to normal tissue controls. These results provide the first evidence of a specific Pyk2 activity in regulating IGF-IR-dependent motility and invasion of bladder cancer cells suggesting that Pyk2 and the IGF-IR may play a critical role in the invasive phenotype in urothelial neoplasia. In addition, Pyk2 and the IGF-IR may serve as novel biomarkers with diagnostic and prognostic significance in bladder cancer.

miR-214 down-regulates ARL2 and suppresses growth and invasion of cervical cancer cells

International Nuclear Information System (INIS)

Peng, Ruiqing; Men, Jianlong; Ma, Rui; Wang, Qian; Wang, Yang; Sun, Ying; Ren, Jing

2017-01-01

Increasing evidence has shown that miRNAs are implicated in carcinogenesis and can function as oncogenes or tumor suppressor genes in human cancers. In this study, we confirmed that miR-214 is frequently down-regulated in cervical cancer compared with normal cervical tissues. Ectopic expression of miR-214 suppressed proliferation, migration and invasion of HeLa and C33A cervical cancer cells. Bioinformatics analysis revealed that ADP ribosylation factor like 2 (ARL2) was a potential target of miR-214 and was remarkably up-regulated in cervical cancer. Knockdown of ARL2 markedly inhibited cervical cancer cell proliferation, migration and invasion, similarly to over-expression of miR-214, indicating that ARL2 may function as an oncogene in cervical cancer. In conclusion, our study revealed that miR-214 acts as a tumor suppressor via inhibiting proliferation, migration and invasion of cervical cancer cells through targeting ARL2, and that both miR-214 and ARL2 may serve as prognostic or therapeutic targets for cervical cancer. - Highlights: • miR-214 targets ARL2. • ARL2 maybe an oncogene in cervical cancer. • ARL2 rescues miR-214.

Immunohistochemical study of macrophage migration inhibitory factor in rat liver fibrosis induced by thioacetamide

OpenAIRE

Y Hori; S Sato; J Yamate; M Kurasaki

2009-01-01

Macrophage migration inhibitory factor (MIF) is a molecule known to regulate macrophage accumulation at sites of inflammation. To elucidate the role of MIF in progression of liver fibrosis, the immunohistochemical localization of MIF and macrophages in the liver were examined. Male Wistar rats received thioacetamide (TA) injections (200 mg/kg, i.p.) for 1 or 6 weeks. In biochemical and histological tests, it was confirmed that liver fibrosis was induced. In immunohistochemical analyses, the e...

NRF2 Signaling Negatively Regulates Phorbol-12-Myristate-13-Acetate (PMA-Induced Differentiation of Human Monocytic U937 Cells into Pro-Inflammatory Macrophages.

Directory of Open Access Journals (Sweden)

Min-Gu Song

Full Text Available Blood monocytes are recruited to injured tissue sites and differentiate into macrophages, which protect against pathogens and repair damaged tissues. Reactive oxygen species (ROS are known to be an important contributor to monocytes' differentiation and macrophages' function. NF-E2-related factor 2 (NRF2, a transcription factor regulating cellular redox homeostasis, is known to be a critical modulator of inflammatory responses. We herein investigated the role of NRF2 in macrophage differentiation using the human monocytic U937 cell line and phorbol-12-myristate-13-acetate (PMA. In U937 cells with NRF2 silencing, PMA-stimulated cell adherence was significantly facilitated when compared to control U937 cells. Both transcript and protein levels for pro-inflammatory cytokines, including interleukine-1β (IL-1β, IL-6, and tumor necrosis factor-α (TNFα were highly elevated in PMA-stimulated NRF2-silenced U937 compared to the control. In addition, PMA-inducible secretion of monocyte chemotactic protein 1 (MCP-1 was significantly high in NRF2-silenced U937. As an underlying mechanism, we showed that NRF2-knockdown U937 retained high levels of cellular ROS and endoplasmic reticulum (ER stress markers expression; and subsequently, PMA-stimulated levels of Ca2+ and PKCα were greater in NRF2-knockdown U937 cells, which caused enhanced nuclear accumulation of nuclear factor-ҡB (NFҡB p50 and extracellular signal-regulated kinase (ERK-1/2 phosphorylation. Whereas the treatment of NRF2-silenced U937 cells with pharmacological inhibitors of NFҡB or ERK1/2 largely blocked PMA-induced IL-1β and IL-6 expression, indicating that these pathways are associated with cell differentiation. Taken together, our results suggest that the NRF2 system functions to suppress PMA-stimulated U937 cell differentiation into pro-inflammatory macrophages and provide evidence that the ROS-PKCα-ERK-NFҡB axis is involved in PMA-facilitated differentiation of NRF2-silenced U937

The β-hemolysin and intracellular survival of Streptococcus agalactiae in human macrophages.

Science.gov (United States)

Sagar, Anubha; Klemm, Carolin; Hartjes, Lara; Mauerer, Stefanie; van Zandbergen, Ger; Spellerberg, Barbara

2013-01-01

S. agalactiae (group B streptococci, GBS) is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches.

The Upregulation of Integrin αDβ2 (CD11d/CD18) on Inflammatory Macrophages Promotes Macrophage Retention in Vascular Lesions and Development of Atherosclerosis.

Science.gov (United States)

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

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

Science.gov (United States)

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

2016-02-12

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

The role of HFE genotype in macrophage phenotype.

Science.gov (United States)

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.

MiR-34a regulates the invasive capacity of canine osteosarcoma cell lines.

Science.gov (United States)

Lopez, Cecilia M; Yu, Peter Y; Zhang, Xiaoli; Yilmaz, Ayse Selen; London, Cheryl A; Fenger, Joelle M

2018-01-01

Osteosarcoma (OSA) is the most common bone tumor in children and dogs; however, no substantial improvement in clinical outcome has occurred in either species over the past 30 years. MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and play a fundamental role in cancer. The purpose of this study was to investigate the potential contribution of miR-34a loss to the biology of canine OSA, a well-established spontaneous model of the human disease. RT-qPCR demonstrated that miR-34a expression levels were significantly reduced in primary canine OSA tumors and canine OSA cell lines as compared to normal canine osteoblasts. In canine OSA cell lines stably transduced with empty vector or pre-miR-34a lentiviral constructs, overexpression of miR-34a inhibited cellular invasion and migration but had no effect on cell proliferation or cell cycle distribution. Transcriptional profiling of canine OSA8 cells possessing enforced miR-34a expression demonstrated dysregulation of numerous genes, including significant down-regulation of multiple putative targets of miR-34a. Moreover, gene ontology analysis of down-regulated miR-34a target genes showed enrichment of several biological processes related to cell invasion and motility. Lastly, we validated changes in miR-34a putative target gene expression, including decreased expression of KLF4, SEM3A, and VEGFA transcripts in canine OSA cells overexpressing miR-34a and identified KLF4 and VEGFA as direct target genes of miR-34a. Concordant with these data, primary canine OSA tumor tissues demonstrated increased expression levels of putative miR-34a target genes. These data demonstrate that miR-34a contributes to invasion and migration in canine OSA cells and suggest that loss of miR-34a may promote a pattern of gene expression contributing to the metastatic phenotype in canine OSA.

MiR-34a regulates the invasive capacity of canine osteosarcoma cell lines.

Directory of Open Access Journals (Sweden)

Cecilia M Lopez

Full Text Available Osteosarcoma (OSA is the most common bone tumor in children and dogs; however, no substantial improvement in clinical outcome has occurred in either species over the past 30 years. MicroRNAs (miRNAs are small non-coding RNAs that regulate gene expression and play a fundamental role in cancer. The purpose of this study was to investigate the potential contribution of miR-34a loss to the biology of canine OSA, a well-established spontaneous model of the human disease.RT-qPCR demonstrated that miR-34a expression levels were significantly reduced in primary canine OSA tumors and canine OSA cell lines as compared to normal canine osteoblasts. In canine OSA cell lines stably transduced with empty vector or pre-miR-34a lentiviral constructs, overexpression of miR-34a inhibited cellular invasion and migration but had no effect on cell proliferation or cell cycle distribution. Transcriptional profiling of canine OSA8 cells possessing enforced miR-34a expression demonstrated dysregulation of numerous genes, including significant down-regulation of multiple putative targets of miR-34a. Moreover, gene ontology analysis of down-regulated miR-34a target genes showed enrichment of several biological processes related to cell invasion and motility. Lastly, we validated changes in miR-34a putative target gene expression, including decreased expression of KLF4, SEM3A, and VEGFA transcripts in canine OSA cells overexpressing miR-34a and identified KLF4 and VEGFA as direct target genes of miR-34a. Concordant with these data, primary canine OSA tumor tissues demonstrated increased expression levels of putative miR-34a target genes.These data demonstrate that miR-34a contributes to invasion and migration in canine OSA cells and suggest that loss of miR-34a may promote a pattern of gene expression contributing to the metastatic phenotype in canine OSA.

Normal autophagic activity in macrophages from mice lacking Gαi3, AGS3, or RGS19.

Directory of Open Access Journals (Sweden)

Ali Vural

Full Text Available In macrophages autophagy assists antigen presentation, affects cytokine release, and promotes intracellular pathogen elimination. In some cells autophagy is modulated by a signaling pathway that employs Gαi3, Activator of G-protein Signaling-3 (AGS3/GPSM1, and Regulator of G-protein Signaling 19 (RGS19. As macrophages express each of these proteins, we tested their importance in regulating macrophage autophagy. We assessed LC3 processing and the formation of LC3 puncta in bone marrow derived macrophages prepared from wild type, Gnai3(-/-, Gpsm1(-/-, or Rgs19(-/- mice following amino acid starvation or Nigericin treatment. In addition, we evaluated rapamycin-induced autophagic proteolysis rates by long-lived protein degradation assays and anti-autophagic action after rapamycin induction in wild type, Gnai3(-/-, and Gpsm1(-/- macrophages. In similar assays we compared macrophages treated or not with pertussis toxin, an inhibitor of GPCR (G-protein couple receptor triggered Gαi nucleotide exchange. Despite previous findings, the level of basal autophagy, autophagic induction, autophagic flux, autophagic degradation and the anti-autophagic action in macrophages that lacked Gαi3, AGS3, or RGS19; or had been treated with pertussis toxin, were similar to controls. These results indicate that while Gαi signaling may impact autophagy in some cell types it does not in macrophages.

Mitochondrial function is involved in regulation of cholesterol efflux to apolipoprotein (apoA-I from murine RAW 264.7 macrophages

Directory of Open Access Journals (Sweden)

Allen Anne Marie

2012-12-01

Full Text Available Abstract Background Mitochondrial DNA damage, increased production of reactive oxygen species and progressive respiratory chain dysfunction, together with increased deposition of cholesterol and cholesteryl esters, are hallmarks of atherosclerosis. This study investigated the role of mitochondrial function in regulation of macrophage cholesterol efflux to apolipoprotein A-I, by the addition of established pharmacological modulators of mitochondrial function. Methods Murine RAW 264.7 macrophages were treated with a range of concentrations of resveratrol, antimycin, dinitrophenol, nigericin and oligomycin, and changes in viability, cytotoxicity, membrane potential and ATP, compared with efflux of [3H]cholesterol to apolipoprotein (apo A-I. The effect of oligomycin treatment on expression of genes implicated in macrophage cholesterol homeostasis were determined by quantitative polymerase chain reaction, and immunoblotting, relative to the housekeeping enzyme, Gapdh, and combined with studies of this molecule on cholesterol esterification, de novo lipid biosynthesis, and induction of apoptosis. Significant differences were determined using analysis of variance, and Dunnett’s or Bonferroni post t-tests, as appropriate. Results The positive control, resveratrol (24 h, significantly enhanced cholesterol efflux to apoA-I at concentrations ≥30 μM. By contrast, cholesterol efflux to apoA-I was significantly inhibited by nigericin (45%; ppAbca1 mRNA. Oligomycin treatment did not affect cholesterol biosynthesis, but significantly inhibited cholesterol esterification following exposure to acetylated LDL, and induced apoptosis at ≥30 μM. Finally, oligomycin induced the expression of genes implicated in both cholesterol efflux (Abca1, Abcg4, Stard1 and cholesterol biosynthesis (Hmgr, Mvk, Scap, Srebf2, indicating profound dysregulation of cholesterol homeostasis. Conclusions Acute loss of mitochondrial function, and in particular Δψm, reduces

Incorporation of cerium oxide into hydroxyapatite coating regulates osteogenic activity of mesenchymal stem cell and macrophage polarization.

Science.gov (United States)

Li, Kai; Shen, Qingyi; Xie, Youtao; You, Mingyu; Huang, Liping; Zheng, Xuebin

2017-02-01

Biomedical coatings for orthopedic implants should facilitate osseointegration and mitigate implant-induced inflammatory reactions. Cerium oxide (CeO 2 ) ceramics possess anti-oxidative properties and can be used to decrease mediators of inflammation, which makes them attractive for biomedical applications. In our work, two kinds of CeO 2 incorporated hydroxyapatite coatings (HA-10Ce and HA-30Ce) were prepared via plasma spraying technique and the effects of CeO 2 addition on the responses of bone mesenchymal stem cells (BMSCs) and RAW264.7 macrophages were investigated. An increase in CeO 2 content in the HA coatings resulted in better osteogenic behaviors of BMSCs in terms of cell proliferation, alkaline phosphatase (ALP) activity and mineralized nodule formation. RT-PCR and western blot analysis suggested that the incorporation of CeO 2 may promote the osteogenic differentiation of BMSCs through the Smad-dependent BMP signaling pathway, which activated Runx2 expression and subsequently enhanced the expression of ALP and OCN. The expression profiles of macrophages cultured on the CeO 2 modified coating revealed a tendency toward a M2 phenotype, because of an upregulation of M2 surface markers (CD163 and CD206), anti-inflammatory cytokines (TNF-α and IL-6) and osteoblastogenesis-related genes (BMP2 and TGF-β1) as well as a downregulation of M1 surface markers (CCR7 and CD11c), proinflammatory cytokines (IL-10 and IL-1ra) and reactive oxygen species production. The results suggested the regulation of BMSCs behaviors and macrophage-mediated responses at the coating's surface were associated with CeO 2 incorporation. The incorporation of CeO 2 in HA coatings can be a valuable strategy to promote osteogenic responses and reduce inflammatory reactions.

Specific macrophage subtypes influence the progression of rhabdomyolysis-induced kidney injury.

Science.gov (United States)

Belliere, Julie; Casemayou, Audrey; Ducasse, Laure; Zakaroff-Girard, Alexia; Martins, Frédéric; Iacovoni, Jason S; Guilbeau-Frugier, Céline; Buffin-Meyer, Bénédicte; Pipy, Bernard; Chauveau, Dominique; Schanstra, Joost P; Bascands, Jean-Loup

2015-06-01

Rhabdomyolysis can be life threatening if complicated by AKI. Macrophage infiltration has been observed in rat kidneys after glycerol-induced rhabdomyolysis, but the role of macrophages in rhabdomyolysis-induced AKI remains unknown. Here, in a patient diagnosed with rhabdomyolysis, we detected substantial macrophage infiltration in the kidney. In a mouse model of rhabdomyolysis-induced AKI, diverse renal macrophage phenotypes were observed depending on the stage of the disease. Two days after rhabdomyolysis, F4/80(low)CD11b(high)Ly6b(high)CD206(low) kidney macrophages were dominant, whereas by day 8, F4/80(high)CD11b(+)Ly6b(low)CD206(high) cells became the most abundant. Single-cell gene expression analyses of FACS-sorted macrophages revealed that these subpopulations were heterogeneous and that individual cells simultaneously expressed both M1 and M2 markers. Liposomal clodronate-mediated macrophage depletion significantly reduced the early infiltration of F4/80(low)CD11b(high)Ly6b(high)CD206(low) macrophages. Furthermore, transcriptionally regulated targets potentially involved in disease progression, including fibronectin, collagen III, and chemoattractants that were identified via single-cell analysis, were verified as macrophage-dependent in situ. In vitro, myoglobin treatment induced proximal tubular cells to secrete chemoattractants and macrophages to express proinflammatory markers. At day 30, liposomal clodronate-mediated macrophage depletion reduced fibrosis and improved both kidney repair and mouse survival. Seven months after rhabdomyolysis, histologic lesions were still present but were substantially reduced with prior depletion of macrophages. These results suggest an important role for macrophages in rhabdomyolysis-induced AKI progression and advocate the utility of long-term follow-up for patients with this disease. Copyright © 2015 by the American Society of Nephrology.

Dihydro-CDDO-trifluoroethyl amide suppresses inflammatory responses in macrophages via activation of Nrf2

International Nuclear Information System (INIS)

Li, Bin; Abdalrahman, Akram; Lai, Yimu; Janicki, Joseph S.; Ward, Keith W.; Meyer, Colin J.; Wang, Xing Li; Tang, Dongqi; Cui, Taixing

2014-01-01

Highlights: • Dh404 suppresses the expression of a selected set of pro-inflammatory cytokines in inflamed macrophages via activating Nrf2. • Dh404 activates Nrf2 while keeping Keap1 function intact in macrophages. • Dh404 minimally regulates NF-κB pathway in macrophages. - Abstract: Nuclear factor erythroid 2-related factor (Nrf2) is the major regulator of cellular defenses against various pathological stresses in a variety of organ systems, thus Nrf2 has evolved to be an attractive drug target for the treatment and/or prevention of human disease. Several synthetic oleanolic triterpenoids including dihydro-CDDO-trifluoroethyl amide (dh404) appear to be potent activators of Nrf2 and exhibit chemopreventive promises in multiple disease models. While the pharmacological efficacy of Nrf2 activators may be dependent on the nature of Nrf2 activation in specific cell types of target organs, the precise role of Nrf2 in mediating biological effects of Nrf2 activating compounds in various cell types remains to be further explored. Herein we report a unique and Nrf2-dependent anti-inflammatory profile of dh404 in inflamed macrophages. In lipopolysaccharide (LPS)-inflamed RAW264.7 macrophages, dh404 dramatically suppressed the expression of pro-inflammatory cytokines including inducible nitric oxide synthase (iNOS), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 beta (MIP-1β), while minimally regulating the expression of interleulin-6 (IL-6), IL-1β, and tumor necrosis factor alpha (TNFα). Dh404 potently activated Nrf2 signaling; however, it did not affect LPS-induced NF-κB activity. Dh404 did not interrupt the interaction of Nrf2 with its endogenous inhibitor Kelch-like ECH associating protein 1 (Keap1) in macrophages. Moreover, knockout of Nrf2 blocked the dh404-induced anti-inflammatory responses in LPS-inflamed macrophages. These results demonstrated that dh404 suppresses pro-inflammatory responses in macrophages via an activation

Dihydro-CDDO-trifluoroethyl amide suppresses inflammatory responses in macrophages via activation of Nrf2

Energy Technology Data Exchange (ETDEWEB)

Li, Bin [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Abdalrahman, Akram; Lai, Yimu; Janicki, Joseph S. [Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Ward, Keith W.; Meyer, Colin J. [Department of Pharmacology, Reata Pharmaceuticals, Inc., Irving, TX 75063 (United States); Wang, Xing Li [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Tang, Dongqi, E-mail: Dongqi.Tang@uscmed.sc.edu [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States); Cui, Taixing, E-mail: taixing.cui@uscmed.sc.edu [Shandong University Qilu Hospital Research Center for Cell Therapy, Key Laboratory of Cardiovascular Remodeling and Function Research, Qilu Hospital of Shandong University, Jinan 250012 (China); Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208 (United States)

2014-02-21

Highlights: • Dh404 suppresses the expression of a selected set of pro-inflammatory cytokines in inflamed macrophages via activating Nrf2. • Dh404 activates Nrf2 while keeping Keap1 function intact in macrophages. • Dh404 minimally regulates NF-κB pathway in macrophages. - Abstract: Nuclear factor erythroid 2-related factor (Nrf2) is the major regulator of cellular defenses against various pathological stresses in a variety of organ systems, thus Nrf2 has evolved to be an attractive drug target for the treatment and/or prevention of human disease. Several synthetic oleanolic triterpenoids including dihydro-CDDO-trifluoroethyl amide (dh404) appear to be potent activators of Nrf2 and exhibit chemopreventive promises in multiple disease models. While the pharmacological efficacy of Nrf2 activators may be dependent on the nature of Nrf2 activation in specific cell types of target organs, the precise role of Nrf2 in mediating biological effects of Nrf2 activating compounds in various cell types remains to be further explored. Herein we report a unique and Nrf2-dependent anti-inflammatory profile of dh404 in inflamed macrophages. In lipopolysaccharide (LPS)-inflamed RAW264.7 macrophages, dh404 dramatically suppressed the expression of pro-inflammatory cytokines including inducible nitric oxide synthase (iNOS), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 beta (MIP-1β), while minimally regulating the expression of interleulin-6 (IL-6), IL-1β, and tumor necrosis factor alpha (TNFα). Dh404 potently activated Nrf2 signaling; however, it did not affect LPS-induced NF-κB activity. Dh404 did not interrupt the interaction of Nrf2 with its endogenous inhibitor Kelch-like ECH associating protein 1 (Keap1) in macrophages. Moreover, knockout of Nrf2 blocked the dh404-induced anti-inflammatory responses in LPS-inflamed macrophages. These results demonstrated that dh404 suppresses pro-inflammatory responses in macrophages via an activation

Radiation Therapy Induces Macrophages to Suppress T-Cell Responses Against Pancreatic Tumors in Mice.

Science.gov (United States)

Seifert, Lena; Werba, Gregor; Tiwari, Shaun; Giao Ly, Nancy Ngoc; Nguy, Susanna; Alothman, Sara; Alqunaibit, Dalia; Avanzi, Antonina; Daley, Donnele; Barilla, Rocky; Tippens, Daniel; Torres-Hernandez, Alejandro; Hundeyin, Mautin; Mani, Vishnu R; Hajdu, Cristina; Pellicciotta, Ilenia; Oh, Philmo; Du, Kevin; Miller, George

2016-06-01

The role of radiation therapy in the treatment of patients with pancreatic ductal adenocarcinoma (PDA) is controversial. Randomized controlled trials investigating the efficacy of radiation therapy in patients with locally advanced unresectable PDA have reported mixed results, with effects ranging from modest benefit to worse outcomes compared with control therapies. We investigated whether radiation causes inflammatory cells to acquire an immune-suppressive phenotype that limits the therapeutic effects of radiation on invasive PDAs and accelerates progression of preinvasive foci. We investigated the effects of radiation therapy in p48(Cre);LSL-Kras(G12D) (KC) and p48(Cre);LSLKras(G12D);LSL-Trp53(R172H) (KPC) mice, as well as in C57BL/6 mice with orthotopic tumors grown from FC1242 cells derived from KPC mice. Some mice were given neutralizing antibodies against macrophage colony-stimulating factor 1 (CSF1 or MCSF) or F4/80. Pancreata were exposed to doses of radiation ranging from 2 to 12 Gy and analyzed by flow cytometry. Pancreata of KC mice exposed to radiation had a higher frequency of advanced pancreatic intraepithelial lesions and more foci of invasive cancer than pancreata of unexposed mice (controls); radiation reduced survival time by more than 6 months. A greater proportion of macrophages from radiation treated invasive and preinvasive pancreatic tumors had an immune-suppressive, M2-like phenotype compared with control mice. Pancreata from mice exposed to radiation had fewer CD8(+) T cells than controls, and greater numbers of CD4(+) T cells of T-helper 2 and T-regulatory cell phenotypes. Adoptive transfer of T cells from irradiated PDA to tumors of control mice accelerated tumor growth. Radiation induced production of MCSF by PDA cells. A neutralizing antibody against MCSF prevented radiation from altering the phenotype of macrophages in tumors, increasing the anti-tumor T-cell response and slowing tumor growth. Radiation treatment causes macrophages

Ginger extract inhibits LPS induced macrophage activation and function

Directory of Open Access Journals (Sweden)

Bruch David

2008-01-01

Full Text Available Abstract Background Macrophages play a dual role in host defence. They act as the first line of defence by mounting an inflammatory response to antigen exposure and also act as antigen presenting cells and initiate the adaptive immune response. They are also the primary infiltrating cells at the site of inflammation. Inhibition of macrophage activation is one of the possible approaches towards modulating inflammation. Both conventional and alternative approaches are being studied in this regard. Ginger, an herbal product with broad anti inflammatory actions, is used as an alternative medicine in a number of inflammatory conditions like rheumatic disorders. In the present study we examined the effect of ginger extract on macrophage activation in the presence of LPS stimulation. Methods Murine peritoneal macrophages were stimulated by LPS in presence or absence of ginger extract and production of proinflammatory cytokines and chemokines were observed. We also studied the effect of ginger extract on the LPS induced expression of MHC II, B7.1, B7.2 and CD40 molecules. We also studied the antigen presenting function of ginger extract treated macrophages by primary mixed lymphocyte reaction. Results We observed that ginger extract inhibited IL-12, TNF-α, IL-1β (pro inflammatory cytokines and RANTES, MCP-1 (pro inflammatory chemokines production in LPS stimulated macrophages. Ginger extract also down regulated the expression of B7.1, B7.2 and MHC class II molecules. In addition ginger extract negatively affected the antigen presenting function of macrophages and we observed a significant reduction in T cell proliferation in response to allostimulation, when ginger extract treated macrophages were used as APCs. A significant decrease in IFN-γ and IL-2 production by T cells in response to allostimulation was also observed. Conclusion In conclusion ginger extract inhibits macrophage activation and APC function and indirectly inhibits T cell activation.

Effects of microparticle size and Fc density on macrophage phagocytosis.

Directory of Open Access Journals (Sweden)

Patricia Pacheco

Full Text Available Controlled induction of phagocytosis in macrophages offers the ability to therapeutically regulate the immune system as well as improve delivery of chemicals or biologicals for immune processing. Maximizing particle uptake by macrophages through Fc receptor-mediated phagocytosis could lead to new delivery mechanisms in drug or vaccine development. Fc ligand density and particle size were examined independently and in combination in order to optimize and tune the phagocytosis of opsonized microparticles. We show the internalization efficiency of small polystyrene particles (0.5 µm to 2 µm is significantly affected by changes in Fc ligand density, while particles greater than 2 µm show little correlation between internalization and Fc density. We found that while macrophages can efficiently phagocytose a large number of smaller particles, the total volume of phagocytosed particles is maximized through the non-specific uptake of larger microparticles. Therefore, larger microparticles may be more efficient at delivering a greater therapeutic payload to macrophages, but smaller opsonized microparticles can deliver bio-active substances to a greater percentage of the macrophage population. This study is the first to treat as independent variables the physical and biological properties of Fc density and microparticle size that initiate macrophage phagocytosis. Defining the physical and biological parameters that affect phagocytosis efficiency will lead to improved methods of microparticle delivery to macrophages.

CAPN 7 promotes the migration and invasion of human endometrial stromal cell by regulating matrix metalloproteinase 2 activity.

Science.gov (United States)

Liu, Hongyu; Jiang, Yue; Jin, Xiaoyan; Zhu, Lihua; Shen, Xiaoyue; Zhang, Qun; Wang, Bin; Wang, Junxia; Hu, Yali; Yan, Guijun; Sun, Haixiang

2013-07-15

Matrix metalloproteinase 2 (MMP-2) has been reported to be an important regulator of cell migration and invasion through degradation of the extracellular matrix (ECM) in many diseases, such as cancer and endometriosis. Here, we found calcium-activated neutral protease 7 (CAPN 7) expression was markedly upregulated in the eutopic endometrium and endometrial stromal cells of women diagnosed with endometriosis. Our studies were carried out to detect the effects of CAPN 7 on human endometrial stromal cell (hESC) migration and invasion. Western blotting and quantitative real-time PCR were used to detect the expression of CAPN 7 in endometriosis patients and normal fertile women. Scratch-wound-healing and invasion chamber assay were used to investigate the role of CAPN 7 in hESC migration and invasion. Western blotting, quantitative real-time PCR and zymography were carried out to detect the effect of CAPN 7 on the expressions and activity of MMP-2. CAPN 7 was markedly up-regulated in endometriosis, thereby promoting the migration and invasion of hESC. CAPN 7 overexpression led to increased expression of MMP-2 and tissue inhibitor of metalloproteinases 2 (TIMP-2); CAPN 7 knockdown reversed these changes. CAPN 7 increased MMP-2 activity by increasing the ratio of MMP-2 to TIMP-2. We also found that OA-Hy (an MMP-2 inhibitor) decreased the effects of CAPN 7 overexpression on hESC migration and invasion by approximately 50% and 55%, respectively. Additionally, a coimmunoprecipitation assay demonstrated that CAPN 7 interacted with activator protein 2α (AP-2α): an important transcription factor of MMP-2. CAPN 7 promotes hESC migration and invasion by increasing the activity of MMP-2 via an increased ratio of MMP-2 to TIMP-2.

Functional Roles of Syk in Macrophage-Mediated Inflammatory Responses

Science.gov (United States)

Yi, Young-Su; Son, Young-Jin; Ryou, Chongsuk; Sung, Gi-Ho; Kim, Jong-Hoon; Cho, Jae Youl

2014-01-01

Inflammation is a series of complex biological responses to protect the host from pathogen invasion. Chronic inflammation is considered a major cause of diseases, such as various types of inflammatory/autoimmune diseases and cancers. Spleen tyrosine kinase (Syk) was initially found to be highly expressed in hematopoietic cells and has been known to play crucial roles in adaptive immune responses. However, recent studies have reported that Syk is also involved in other biological functions, especially in innate immune responses. Although Syk has been extensively studied in adaptive immune responses, numerous studies have recently presented evidence that Syk has critical functions in macrophage-mediated inflammatory responses and is closely related to innate immune response. This review describes the characteristics of Syk-mediated signaling pathways, summarizes the recent findings supporting the crucial roles of Syk in macrophage-mediated inflammatory responses and diseases, and discusses Syk-targeted drug development for the therapy of inflammatory diseases. PMID:25045209

Functional Roles of Syk in Macrophage-Mediated Inflammatory Responses

Directory of Open Access Journals (Sweden)

Young-Su Yi

2014-01-01

Full Text Available Inflammation is a series of complex biological responses to protect the host from pathogen invasion. Chronic inflammation is considered a major cause of diseases, such as various types of inflammatory/autoimmune diseases and cancers. Spleen tyrosine kinase (Syk was initially found to be highly expressed in hematopoietic cells and has been known to play crucial roles in adaptive immune responses. However, recent studies have reported that Syk is also involved in other biological functions, especially in innate immune responses. Although Syk has been extensively studied in adaptive immune responses, numerous studies have recently presented evidence that Syk has critical functions in macrophage-mediated inflammatory responses and is closely related to innate immune response. This review describes the characteristics of Syk-mediated signaling pathways, summarizes the recent findings supporting the crucial roles of Syk in macrophage-mediated inflammatory responses and diseases, and discusses Syk-targeted drug development for the therapy of inflammatory diseases.

The macrophage scavenger receptor (CD163): a double-edged sword in treatment of malignant disease

DEFF Research Database (Denmark)

Maniecki, Maciej Bogdan

2009-01-01

of inflammatory processes. The receptor is expressed by circulatory monocytes and it is highly expressed on tissue-resident macrophages. CD163 is also expressed on leukemic blast cells of AML type M4/M5 and tumor cells in malignant melanoma and breast cancer. Although circumstantial evidence of the potential...... was investigated in biopsies from bladder cancer patients. We demonstrated that CD163 mRNA expression was significantly elevated in muscle invasive tumors (T2-T4) compared with superficial tumors (Ta), and that a high level of CD163 mRNA expression in tumor biopsies was significantly associated with poor 13-year......The hemoglobin scavenger receptor CD163 is a transmembrane glycoprotein belonging to the scavenger receptor cysteine-rich (SRCR) domain family. It mediates the clearance of hemoglobin released to the circulation during intravascular hemolysis, and it is also involved in the regulation...

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

Directory of Open Access Journals (Sweden)

Miguel Pinilla-Vera

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

NFIB Mediates BRN2 Driven Melanoma Cell Migration and Invasion Through Regulation of EZH2 and MITF

Directory of Open Access Journals (Sweden)

Mitchell E. Fane

2017-02-01

Full Text Available While invasion and metastasis of tumour cells are the principle factor responsible for cancer related deaths, the mechanisms governing the process remain poorly defined. Moreover, phenotypic divergence of sub-populations of tumour cells is known to underpin alternative behaviors linked to tumour progression such as proliferation, survival and invasion. In the context of melanoma, heterogeneity between two transcription factors, BRN2 and MITF, has been associated with phenotypic switching between predominantly invasive and proliferative behaviors respectively. Epigenetic changes, in response to external cues, have been proposed to underpin this process, however the mechanism by which the phenotypic switch occurs is unclear. Here we report the identification of the NFIB transcription factor as a novel downstream effector of BRN2 function in melanoma cells linked to the migratory and invasive characteristics of these cells. Furthermore, the function of NFIB appears to drive an invasive phenotype through an epigenetic mechanism achieved via the upregulation of the polycomb group protein EZH2. A notable target of NFIB mediated up-regulation of EZH2 is decreased MITF expression, which further promotes a less proliferative, more invasive phenotype. Together our data reveal that NFIB has the ability to promote dynamic changes in the chromatin state of melanoma cells to facilitate migration, invasion and metastasis.

Neopterin negatively regulates expression of ABCA1 and ABCG1 by the LXRα signaling pathway in THP-1 macrophage-derived foam cells.

Science.gov (United States)

Yan, Jin-quan; Tan, Chun-zhi; Wu, Jin-hua; Zhang, Dong-cui; Chen, Ji-ling; Zeng, Bin-yuan; Jiang, Yu-ping; Nie, Jin; Liu, Wei; Liu, Qin; Dai, Hao

2013-07-01

To investigate the effects of neopterin on ABCA1 expression and cholesterol efflux in human THP-1 macrophage-derived foam cells, and to explore the role of the liver X receptor alpha (LXRα) involved. In the present study, THP-1 cells were pre-incubated with ox-LDL to become foam cells. The protein and mRNA expression were examined by Western blot assays and real-time quantitative PCR, respectively. Liquid scintillation counting and high performance liquid chromatography assays were used to test cellular cholesterol efflux and cholesterol content. Neopterin decreased ABCA1 expression and cholesterol efflux in a time- and concentration-dependent manner in THP-1 macrophage-derived foam cells, and the LXRα siRNA can reverse the inhibitory effects induced by neopterin. Neoterin has a negative regulation on ABCA1 expression via the LXRα signaling pathway, which suggests the aggravated effects of neopterin on atherosclerosis.

Major Vault Protein Regulates Class A Scavenger Receptor-mediated Tumor Necrosis Factor-α Synthesis and Apoptosis in Macrophages*

Science.gov (United States)

Ben, Jingjing; Zhang, Yan; Zhou, Rongmei; Zhang, Haiyang; Zhu, Xudong; Li, Xiaoyu; Zhang, Hanwen; Li, Nan; Zhou, Xiaodan; Bai, Hui; Yang, Qing; Li, Donghai; Xu, Yong; Chen, Qi

2013-01-01

Atherosclerosis is considered a disease of chronic inflammation largely initiated and perpetuated by macrophage-dependent synthesis and release of pro-inflammatory mediators. Class A scavenger receptor (SR-A) expressed on macrophages plays a key role in this process. However, how SR-A-mediated pro-inflammatory response is modulated in macrophages remains ill defined. Here through immunoprecipitation coupled with mass spectrometry, we reported major vault protein (MVP) as a novel binding partner for SR-A. The interaction between SR-A and MVP was confirmed by immunofluorescence staining and chemical cross-linking assay. Treatment of macrophages with fucoidan, a SR-A ligand, led to a marked increase in TNF-α production, which was attenuated by MVP depletion. Further analysis revealed that SR-A stimulated TNF-α synthesis in macrophages via the caveolin- instead of clathrin-mediated endocytic pathway linked to p38 and JNK, but not ERK, signaling pathways. Importantly, fucoidan invoked an enrichment of MVP in lipid raft, a caveolin-reliant membrane structure, and enhanced the interaction among SR-A, caveolin, and MVP. Finally, we demonstrated that MVP elimination ameliorated SR-A-mediated apoptosis in macrophages. As such, MVP may fine-tune SR-A activity in macrophages which contributes to the development of atherosclerosis. PMID:23703615

Controlled release of cytokines using silk-biomaterials for macrophage polarization.

Science.gov (United States)

Reeves, Andrew R D; Spiller, Kara L; Freytes, Donald O; Vunjak-Novakovic, Gordana; Kaplan, David L

2015-12-01

Polarization of macrophages into an inflammatory (M1) or anti-inflammatory (M2) phenotype is important for clearing pathogens and wound repair, however chronic activation of either type of macrophage has been implicated in several diseases. Methods to locally control the polarization of macrophages is of great interest for biomedical implants and tissue engineering. To that end, silk protein was used to form biopolymer films that release either IFN-γ or IL-4 to control the polarization of macrophages. Modulation of the solubility of the silk films through regulation of β-sheet (crystalline) content enabled a short-term release (4-8 h) of either cytokine, with smaller amounts released out to 24 h. Altering the solubility of the films was accomplished by varying the time that the films were exposed to water vapor. The released IFN-γ or IL-4 induced polarization of THP-1 derived macrophages into the M1 or M2 phenotypes, respectively. The silk biomaterials were able to release enough IFN-γ or IL-4 to repolarize the macrophage from M1 to M2 and vice versa, demonstrating the well-established plasticity of macrophages. High β-sheet content films that are not soluble and do not release the trapped cytokines were also able to polarize macrophages that adhered to the surface through degradation of the silk protein. Chemically conjugating IFN-γ to silk films through disulfide bonds allowed for longer-term release to 10 days. The release of covalently attached IFN-γ from the films was also able to polarize M1 macrophages in vitro. Thus, the strategy described here offers new approaches to utilizing biomaterials for directing the polarization of macrophages. Copyright © 2015 Elsevier Ltd. All rights reserved.

CENPI is overexpressed in colorectal cancer and regulates cell migration and invasion.

Science.gov (United States)

Ding, Na; Li, Rongxin; Shi, Wenhao; He, Cui

2018-06-21

Centromere protein I (CENPI),an important member of centromere protein family, has been suggest to serve as a oncogene in breast cancer, but the clinical significance and biological function of CENPI in colorectal cancer (CRC) is still unclear. In our results, we found CENPI was overexpressed in CRC tissues and cells, and associated with clinical stage, tumor depth, lymph node metastasis, distant metastasis and differentiation in CRC patients. However, there was no significant association between CENPI protein expression and overall survival time in colon cancer patients and rectal cancer patients through analyzing TCGA survival data. Moreover, CENPI mRNA and protein were increased in metastatic lymph nodes compared with primary CRC tissues. Down-regulation of CENPI expression suppresses CRC cell migration, invasion and epithelial mesenchymal transition process. In conclusion, CENPI is overexpressed in CRC and functions as oncogene in modulating CRC cell migration, invasion and EMT process. Copyright © 2018. Published by Elsevier B.V.

Matrix Metallopeptidase 14 Plays an Important Role in Regulating Tumorigenic Gene Expression and Invasion Ability of HeLa Cells.

Science.gov (United States)

Zhang, Ying-Hui; Wang, Juan-Juan; Li, Min; Zheng, Han-Xi; Xu, Lan; Chen, You-Guo

2016-03-01

The objectives of this study were to investigate the functional effect of matrix metallopeptidase 14 (MMP14) on cell invasion in cervical cancer cells (HeLa line) and to study the underlying molecular mechanisms. Expression vector of short hairpin RNA targeting MMP14 was treated in HeLa cells, and then, transfection efficiency was verified by a florescence microscope. Transwell assay was used to investigate cell invasion ability in HeLa cells. Quantitative polymerase chain reaction and Western blotting analysis were used to detect the expression of MMP14 and relative factors in messenger RNA and protein levels, respectively. Matrix metallopeptidase 14 short hairpin RNA expression vector transfection obviously decreased MMP14 expression in messenger RNA and protein levels. Down-regulation of MMP14 suppressed invasion ability of HeLa cells and reduced transforming growth factor β1 and vascular endothelial growth factor B expressions. Furthermore, MMP14 knockdown decreased bone sialoprotein and enhanced forkhead box protein L2 expression in both RNA and protein levels. Matrix metallopeptidase 14 plays an important role in regulating invasion of HeLa cells. Matrix metallopeptidase 14 knockdown contributes to attenuating the malignant phenotype of cervical cancer cell.

Rhinovirus infection induces distinct transcriptome profiles in polarized human macrophages.

Science.gov (United States)

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

2018-05-01

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

Adventitial fibroblasts induce a distinct proinflammatory/profibrotic macrophage phenotype in pulmonary hypertension.

Science.gov (United States)

El Kasmi, Karim C; Pugliese, Steven C; Riddle, Suzette R; Poth, Jens M; Anderson, Aimee L; Frid, Maria G; Li, Min; Pullamsetti, Soni S; Savai, Rajkumar; Nagel, Maria A; Fini, Mehdi A; Graham, Brian B; Tuder, Rubin M; Friedman, Jacob E; Eltzschig, Holger K; Sokol, Ronald J; Stenmark, Kurt R

2014-07-15

Macrophage accumulation is not only a characteristic hallmark but is also a critical component of pulmonary artery remodeling associated with pulmonary hypertension (PH). However, the cellular and molecular mechanisms that drive vascular macrophage activation and their functional phenotype remain poorly defined. Using multiple levels of in vivo (bovine and rat models of hypoxia-induced PH, together with human tissue samples) and in vitro (primary mouse, rat, and bovine macrophages, human monocytes, and primary human and bovine fibroblasts) approaches, we observed that adventitial fibroblasts derived from hypertensive pulmonary arteries (bovine and human) regulate macrophage activation. These fibroblasts activate macrophages through paracrine IL-6 and STAT3, HIF1, and C/EBPβ signaling to drive expression of genes previously implicated in chronic inflammation, tissue remodeling, and PH. This distinct fibroblast-activated macrophage phenotype was independent of IL-4/IL-13-STAT6 and TLR-MyD88 signaling. We found that genetic STAT3 haplodeficiency in macrophages attenuated macrophage activation, complete STAT3 deficiency increased macrophage activation through compensatory upregulation of STAT1 signaling, and deficiency in C/EBPβ or HIF1 attenuated fibroblast-driven macrophage activation. These findings challenge the current paradigm of IL-4/IL-13-STAT6-mediated alternative macrophage activation as the sole driver of vascular remodeling in PH, and uncover a cross-talk between adventitial fibroblasts and macrophages in which paracrine IL-6-activated STAT3, HIF1α, and C/EBPβ signaling are critical for macrophage activation and polarization. Thus, targeting IL-6 signaling in macrophages by completely inhibiting C/EBPβ or HIF1α or by partially inhibiting STAT3 may hold therapeutic value for treatment of PH and other inflammatory conditions characterized by increased IL-6 and absent IL-4/IL-13 signaling. Copyright © 2014 by The American Association of Immunologists

Suppression of TLR4-mediated inflammatory response by macrophage class A scavenger receptor (CD204)

Energy Technology Data Exchange (ETDEWEB)

Ohnishi, Koji; Komohara, Yoshihiro; Fujiwara, Yukio; Takemura, Kenichi [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Lei, XiaoFeng [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Department of Biochemistry, Showa University School of Medicine, Tokyo (Japan); Nakagawa, Takenobu [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Sakashita, Naomi [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan); Department of Human Pathology, Institute of Health Biosciences, The University of Tokushima, Tokushima (Japan); Takeya, Motohiro, E-mail: takeya@kumamoto-u.ac.jp [Department of Cell Pathology, Graduate School of Medical Sciences, Faculty of Life Sciences, Kumamoto University, Kumamoto (Japan)

2011-08-05

Highlights: {yields} We focused on the interaction between SR-A and TLR4 signaling in this study. {yields} SR-A deletion promoted NF{kappa}B activation in macrophages in septic model mouse. {yields} SR-A suppresses both MyD88-dependent and -independent TLR4 signaling in vitro. {yields} SR-A clears LPS binding to TLR4 which resulting in the suppression of TLR4 signals. -- Abstract: The class A scavenger receptor (SR-A, CD204), one of the principal receptors expressed on macrophages, has been found to regulate inflammatory response and attenuate septic endotoxemia. However, the detailed mechanism of this process has not yet been well characterized. To clarify the regulative mechanisms of lipopolysaccharide (LPS)-induced macrophage activation by SR-A, we evaluated the activation of Toll-like receptor 4 (TLR4)-mediated signaling molecules in SR-A-deficient (SR-A{sup -/-}) macrophages. In a septic shock model, the blood levels of tumor necrosis factor (TNF)-{alpha}, interleukin (IL)-6 and interferon (IFN)-{beta} were significantly increased in SR-A{sup -/-} mice compared to wild-type mice, and elevated nuclear factor kappa B (NF{kappa}B) activation was detected in SR-A{sup -/-} macrophages. SR-A deletion increased the production of pro-inflammatory cytokines, and the phosphorylation of mitogen-activated protein kinase (MAPK) and NF{kappa}B in vitro. SR-A deletion also promoted the nuclear translocation of NF{kappa}B and IFN regulatory factor (IRF)-3. In addition, a competitive binding assay with acetylated low-density lipoprotein, an SR-A-specific ligand, and anti-SR-A antibody induced significant activation of TLR4-mediated signaling molecules in wild-type macrophages but not in SR-A{sup -/-} macrophages. These results suggest that SR-A suppresses the macrophage activation by inhibiting the binding of LPS to TLR4 in a competitive manner and it plays a pivotal role in the regulation of the LPS-induced inflammatory response.

Reduced host cell invasiveness and oxidative stress tolerance in double and triple csp gene family deletion mutants of Listeria monocytogenes.

Science.gov (United States)

Loepfe, Chantal; Raimann, Eveline; Stephan, Roger; Tasara, Taurai

2010-07-01

The cold shock protein (Csp) family comprises small, highly conserved proteins that bind nucleic acids to modulate various bacterial gene expressions. In addition to cold adaptation functions, this group of proteins is thought to facilitate various cellular processes to promote normal growth and stress adaptation responses. Three proteins making up the Listeria monocytogenes Csp family (CspA, CspB, and CspD) promote both cold and osmotic stress adaptation functions in this bacterium. The contribution of these three Csps in the host cell invasion processes of L. monocytogenes was investigated based on human Caco-2 and murine macrophage in vitro cell infection models. The DeltacspB, DeltacspD, DeltacspAB, DeltacspAD, DeltacspBD, and DeltacspABD strains were all significantly impaired in Caco-2 cell invasion compared with the wild-type strain, whereas in the murine macrophage infection assay only, the double (DeltacspBD) and triple (DeltacspABD) csp mutants were also significantly impaired in cell invasion compared with the wild-type strain. The DeltacspBD and DeltacspABD mutants displayed the most severely impaired invasion phenotypes. The invasion ability of these two mutant strains was also further analyzed using cold-stress-exposed organisms. In both cell infection models a significant reduction in invasiveness was observed after cold stress exposure of Listeria organisms. The negative impact of cold stress on subsequent cell invasion ability was, however, more severe in cold-sensitive csp mutants (DeltacspBD and DeltacspABD) compared with the wild type. The impaired macrophage invasion and intracellular growth of DeltacspBD and DeltacspABD also led us to examine oxidative stress resistance capacity in these two mutant strains. Both strains also displayed higher oxidative stress sensitivity relative to the wild-type strain. Our data indicate that besides cold and osmotic stress adaptation roles, Csp family proteins also promote efficient host cell invasion and

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

Science.gov (United States)

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.

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

International Nuclear Information System (INIS)

Liu, Chun; Ge, Beihai; He, Chao; Zhang, Yi; Liu, Xiaowen; Liu, Kejian; Qian, Cuiping; Zhang, Yu; Peng, Wenzhong; Guo, Xiaomei

2014-01-01

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, an 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

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

Energy Technology Data Exchange (ETDEWEB)

Liu, Chun; Ge, Beihai [Department of Cardiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan 430030 (China); He, Chao [Department of Cardiology, China Three Gorges University, Yichang 433000 (China); Zhang, Yi; Liu, Xiaowen [Department of Cardiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan 430030 (China); Liu, Kejian [Department of Cardiology, The First Affiliated Hospital of Medical College, Shihezi University (China); Qian, Cuiping; Zhang, Yu; Peng, Wenzhong [Department of Cardiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan 430030 (China); Guo, Xiaomei, E-mail: xmguo@tjh.tjmu.edu.cn [Department of Cardiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan 430030 (China)

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, an 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.

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

Science.gov (United States)

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

1995-01-01

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

Interleukin-6 Contributes to Age-Related Alteration of Cytokine Production by Macrophages

Science.gov (United States)

Gomez, Christian R.; Karavitis, John; Palmer, Jessica L.; Faunce, Douglas E.; Ramirez, Luis; Nomellini, Vanessa; Kovacs, Elizabeth J.

2010-01-01

Here, we studied in vitro cytokine production by splenic macrophages obtained from young and aged BALB/c wild type (WT) and IL-6 knockout (IL-6 KO) mice. Relative to macrophages obtained from young WT mice given lipopolysaccharide (LPS), those from aged WT mice had decreased production of proinflammatory cytokines. In contrast, when compared to macrophages from young IL-6 KO mice, LPS stimulation yielded higher levels of these cytokines by cells from aged IL-6 KO mice. Aging or IL-6 deficiency did not affected the percentage of F4/80+ macrophages, or the surface expression of Toll-like receptor 4 (TLR4) and components of the IL-6 receptor. Overall, our results indicate that IL-6 plays a role in regulating the age-related defects in macrophages through alteration of proinflammatory cytokines, adding to the complexity of IL-6-mediated impairment of immune cell function with increasing age. PMID:20671912

Regulation of endothelial cell adhesion molecule expression by mast cells, macrophages, and neutrophils.

Directory of Open Access Journals (Sweden)

Jie Zhang

2011-01-01

Full Text Available Leukocyte adhesion to the vascular endothelium and subsequent transendothelial migration play essential roles in the pathogenesis of cardiovascular diseases such as atherosclerosis. The leukocyte adhesion is mediated by localized activation of the endothelium through the action of inflammatory cytokines. The exact proinflammatory factors, however, that activate the endothelium and their cellular sources remain incompletely defined.Using bone marrow-derived mast cells from wild-type, Tnf(-/-, Ifng(-/-, Il6(-/- mice, we demonstrated that all three of these pro-inflammatory cytokines from mast cells induced the expression of vascular cell adhesion molecule-1 (VCAM-1, intercellular adhesion molecule-1 (ICAM-1, P-selectin, and E-selectin in murine heart endothelial cells (MHEC at both mRNA and protein levels. Compared with TNF-α and IL6, IFN-γ appeared weaker in the induction of the mRNA levels, but at protein levels, both IL6 and IFN-γ were weaker inducers than TNF-α. Under physiological shear flow conditions, mast cell-derived TNF-α and IL6 were more potent than IFN-γ in activating MHEC and in promoting neutrophil adhesion. Similar observations were made when neutrophils or macrophages were used. Neutrophils and macrophages produced the same sets of pro-inflammatory cytokines as did mast cells to induce MHEC adhesion molecule expression, with the exception that macrophage-derived IFN-γ showed negligible effect in inducing VCAM-1 expression in MHEC.Mast cells, neutrophils, and macrophages release pro-inflammatory cytokines such as TNF-α, IFN-γ, and IL6 that induce expression of adhesion molecules in endothelium and recruit of leukocytes, which is essential to the pathogenesis of vascular inflammatory diseases.

ANDROGENS REGULATE T47D CELLS MOTILITY AND INVASION THROUGH ACTIN CYTOSKELETON REMODELLING

Directory of Open Access Journals (Sweden)

Maria Magdalena Montt-Guevara

2016-09-01

Full Text Available The relationship between androgens and breast cancer is controversial. Androgens have complex effects on breast cancer progression and metastasis. Moreover, androgens receptor (AR is expressed in approximately 70% to 90% of invasive breast carcinomas, which has prognostic relevance in basal-like cancers and in triple negative breast cancers. Recent studies have associated the actin-binding proteins of the Ezrin-Radixin-Moesin (ERM family with metastasis in endocrine-sensitive cancers. We studied on T47D breast cancer cells whether androgens with different characteristics, such as testosterone (T, dihydrotestosterone (DHT and dehydroepiandrosterone (DHEA may regulate breast cancer cell motility and invasion through the control of actin remodelling. We demonstrate that androgens promote migration and invasion in T47D via Moesin activation. We show that T and DHEA exert their actions via the AR and estrogen receptor (ER, while the non aromatizable androgen – DHT only recruits AR. We further report that androgen induced significant changes in actin organization with pseudopodia along with membrane ruffles formation, and this process is mediated by Moesin. Our work identifies novel mechanisms of action of androgens on breast cancer cells. Through the modulation of Moesin, androgens alter the architecture of cytoskeleton in T47D breast cancer cell and promote cell migration and invasion. These results could help to understand the biological actions of androgens on breast cancer, and eventually to develop new strategies for treatment of breast cancer.

Corticotropin-Releasing Hormone (CRH Promotes Macrophage Foam Cell Formation via Reduced Expression of ATP Binding Cassette Transporter-1 (ABCA1.

Directory of Open Access Journals (Sweden)

Wonkyoung Cho

Full Text Available Atherosclerosis, the major pathology of cardiovascular disease, is caused by multiple factors involving psychological stress. Corticotropin-releasing hormone (CRH, which is released by neurosecretory cells in the hypothalamus, peripheral nerve terminals and epithelial cells, regulates various stress-related responses. Our current study aimed to verify the role of CRH in macrophage foam cell formation, the initial critical stage of atherosclerosis. Our quantitative real-time reverse transcriptase PCR (qRT-PCR, semi-quantitative reverse transcriptase PCR, and Western blot results indicate that CRH down-regulates ATP-binding cassette transporter-1 (ABCA1 and liver X receptor (LXR-α, a transcription factor for ABCA1, in murine peritoneal macrophages and human monocyte-derived macrophages. Oil-red O (ORO staining and intracellular cholesterol measurement of macrophages treated with or without oxidized LDL (oxLDL and with or without CRH (10 nM in the presence of apolipoprotein A1 (apoA1 revealed that CRH treatment promotes macrophage foam cell formation. The boron-dipyrromethene (BODIPY-conjugated cholesterol efflux assay showed that CRH treatment reduces macrophage cholesterol efflux. Western blot analysis showed that CRH-induced down-regulation of ABCA1 is dependent on phosphorylation of Akt (Ser473 induced by interaction between CRH and CRH receptor 1(CRHR1. We conclude that activation of this pathway by CRH accelerates macrophage foam cell formation and may promote stress-related atherosclerosis.

Macrophage polarization differs between apical granulomas, radicular cysts, and dentigerous cysts.

Science.gov (United States)

Weber, Manuel; Schlittenbauer, Tilo; Moebius, Patrick; Büttner-Herold, Maike; Ries, Jutta; Preidl, Raimund; Geppert, Carol-Immanuel; Neukam, Friedrich W; Wehrhan, Falk

2018-01-01

Apical periodontitis can appear clinically as apical granulomas or radicular cysts. There is evidence that immunologic factors are involved in the pathogenesis of both pathologies. In contrast to radicular cysts, the dentigerous cysts have a developmental origin. Macrophage polarization (M1 vs M2) is a main regulator of tissue homeostasis and differentiation. There are no studies comparing macrophage polarization in apical granulomas, radicular cysts, and dentigerous cysts. Forty-one apical granulomas, 23 radicular cysts, and 23 dentigerous cysts were analyzed in this study. A tissue microarray (TMA) of the 87 consecutive specimens was created, and CD68-, CD11c-, CD163-, and MRC1-positive macrophages were detected by immunohistochemical methods. TMAs were digitized, and the expression of macrophage markers was quantitatively assessed. Radicular cysts are characterized by M1 polarization of macrophages while apical granulomas show a significantly higher degree of M2 polarization. Dentigerous cysts have a significantly lower M1 polarization than both analyzed periapical lesions (apical granulomas and radicular cysts) and accordingly, a significantly higher M2 polarization than radicular cysts. Macrophage cell density in dentigerous cysts is significantly lower than in the periapical lesions. The development of apical periodontitis towards apical granulomas or radicular cysts might be directed by macrophage polarization. Radicular cyst formation is associated with an increased M1 polarization of infiltrating macrophages. In contrast to radicular cysts, dentigerous cysts are characterized by a low macrophage infiltration and a high degree of M2 polarization, possibly reflecting their developmental rather than inflammatory origin. As M1 polarization of macrophages is triggered by bacterial antigens, these results underline the need for sufficient bacterial clearance during endodontic treatment to prevent a possible M1 macrophage-derived stimulus for radicular cyst

Regulation of the arachidonic acid mobilization in macrophages by combustion-derived particles

Directory of Open Access Journals (Sweden)

Weiss Carsten

2011-08-01

Full Text Available Abstract Background Acute exposure to elevated levels of environmental particulate matter (PM is associated with increasing morbidity and mortality rates. These adverse health effects, e.g. culminating in respiratory and cardiovascular diseases, have been demonstrated by a multitude of epidemiological studies. However, the underlying mechanisms relevant for toxicity are not completely understood. Especially the role of particle-induced reactive oxygen species (ROS, oxidative stress and inflammatory responses is of particular interest. In this in vitro study we examined the influence of particle-generated ROS on signalling pathways leading to activation of the arachidonic acid (AA cascade. Incinerator fly ash particles (MAF02 were used as a model for real-life combustion-derived particulate matter. As macrophages, besides epithelial cells, are the major targets of particle actions in the lung murine RAW264.7 macrophages and primary human macrophages were investigated. Results The interaction of fly ash particles with macrophages induced both the generation of ROS and as part of the cellular inflammatory responses a dose- and time-dependent increase of free AA, prostaglandin E2/thromboxane B2 (PGE2/TXB2, and 8-isoprostane, a non-enzymatically formed oxidation product of AA. Additionally, increased phosphorylation of the mitogen-activated protein kinases (MAPK JNK1/2, p38 and ERK1/2 was observed, the latter of which was shown to be involved in MAF02-generated AA mobilization and phosphorylation of the cytosolic phospolipase A2. Using specific inhibitors for the different phospolipase A2 isoforms the MAF02-induced AA liberation was shown to be dependent on the cytosolic phospholipase A2, but not on the secretory and calcium-independent phospholipase A2. The initiation of the AA pathway due to MAF02 particle exposure was demonstrated to depend on the formation of ROS since the presence of the antioxidant N-acetyl-cysteine (NAC prevented the MAF02

Altered CXCR3 isoform expression regulates prostate cancer cell migration and invasion

Directory of Open Access Journals (Sweden)

Wu Qian

2012-01-01

Full Text Available Abstract Background Carcinoma cells must circumvent the normally suppressive signals to disseminate. While often considered 'stop' signals for adherent cells, CXCR3-binding chemokines have recently been correlated positively with cancer progression though the molecular basis remains unclear. Results Here, we examined the expression and function of two CXCR3 variants in human prostate cancer biopsies and cell lines. Globally, both CXCR3 mRNA and protein were elevated in localized and metastatic human cancer biopsies compared to normal. Additionally, CXCR3A mRNA level was upregulated while CXCR3B mRNA was downregulated in these prostate cancer specimens. In contrast to normal prostate epithelial cells (RWPE-1, CXCR3A was up to half the receptor in the invasive and metastatic DU-145 and PC-3 prostate cancer cells, but not in the localized LNCaP cells. Instead of inhibiting cell migration as in RWPE-1 cells, the CXCR3 ligands CXCL4/PF4 and CXCL10/IP10 promoted cell motility and invasiveness in both DU-145 and PC-3 cells via PLCβ3 and μ-calpain activation. CXCR3-mediated diminution of cell motility in RWPE-1 cells is likely a result of cAMP upregulation and m-calpain inhibition via CXCR3B signal transduction. Interestingly, overexpression of CXCR3B in DU-145 cells decreased cell movement and invasion. Conclusion These data suggest that the aberrant expression of CXCR3A and down-regulation of CXCR3B may switch a progression "stop" to a "go" signal to promote prostate tumor metastasis via stimulating cell migration and invasion.

Prostate-Specific G-Protein Coupled Receptor, an Emerging Biomarker Regulating Inflammation and Prostate Cancer Invasion.

Science.gov (United States)

Rodriguez, M; Siwko, S; Liu, M

2016-01-01

Prostate cancer is highly prevalent among men in developed countries, but a significant proportion of detected cancers remain indolent, never progressing into aggressive carcinomas. This highlights the need to develop refined biomarkers that can distinguish between indolent and potentially dangerous cases. The prostate-specific G-protein coupled receptor (PSGR, or OR51E2) is an olfactory receptor family member with highly specific expression in human prostate epithelium that is highly overexpressed in PIN and prostate cancer. PSGR has been functionally implicated in prostate cancer cell invasiveness, suggesting a potential role in the transition to metastatic PCa. Recently, transgenic mice overexpressing PSGR in the prostate were reported to develop an acute inflammatory response followed by emergence of low grade PIN, whereas mice with compound PSGR overexpression and loss of PTEN exhibited accelerated formation of invasive prostate adenocarcinoma. This article will review recent PSGR findings with a focus on its role as a potential prostate cancer biomarker and regulator of prostate cancer invasion and inflammation.

Human genetic variation in VAC14 regulates Salmonella invasion and typhoid fever through modulation of cholesterol.

Science.gov (United States)

Alvarez, Monica I; Glover, Luke C; Luo, Peter; Wang, Liuyang; Theusch, Elizabeth; Oehlers, Stefan H; Walton, Eric M; Tram, Trinh Thi Bich; Kuang, Yu-Lin; Rotter, Jerome I; McClean, Colleen M; Chinh, Nguyen Tran; Medina, Marisa W; Tobin, David M; Dunstan, Sarah J; Ko, Dennis C

2017-09-12

Risk, severity, and outcome of infection depend on the interplay of pathogen virulence and host susceptibility. Systematic identification of genetic susceptibility to infection is being undertaken through genome-wide association studies, but how to expeditiously move from genetic differences to functional mechanisms is unclear. Here, we use genetic association of molecular, cellular, and human disease traits and experimental validation to demonstrate that genetic variation affects expression of VAC14, a phosphoinositide-regulating protein, to influence susceptibility to Salmonella enterica serovar Typhi ( S Typhi) infection. Decreased VAC14 expression increased plasma membrane cholesterol, facilitating Salmonella docking and invasion. This increased susceptibility at the cellular level manifests as increased susceptibility to typhoid fever in a Vietnamese population. Furthermore, treating zebrafish with a cholesterol-lowering agent, ezetimibe, reduced susceptibility to S Typhi. Thus, coupling multiple genetic association studies with mechanistic dissection revealed how VAC14 regulates Salmonella invasion and typhoid fever susceptibility and may open doors to new prophylactic/therapeutic approaches.

MicroRNAs Control Macrophage Formation and Activation: The Inflammatory Link between Obesity and Cardiovascular Diseases

Directory of Open Access Journals (Sweden)

Richard Cheng-An Chang

2014-07-01

Full Text Available Activation and recruitment of resident macrophages in tissues in response to physiological stress are crucial regulatory processes in promoting the development of obesity-associated metabolic disorders and cardiovascular diseases. Recent studies have provided compelling evidence that microRNAs play important roles in modulating monocyte formation, macrophage maturation, infiltration into tissues and activation. Macrophage-dependent systemic physiological and tissue-specific responses also involve cell-cell interactions between macrophages and host tissue niche cell components, including other tissue-resident immune cell lineages, adipocytes, vascular smooth muscle and others. In this review, we highlight the roles of microRNAs in regulating the development and function of macrophages in the context of obesity, which could provide insights into the pathogenesis of obesity-related metabolic syndrome and cardiovascular diseases.

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

DEFF Research Database (Denmark)

Falchi, Mario; Varricchio, Lilian; Martelli, Fabrizio

2015-01-01

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

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

Directory of Open Access Journals (Sweden)

Hai B Tran

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

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

International Nuclear Information System (INIS)

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

2014-01-01

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

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

Science.gov (United States)

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

2014-01-01

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

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

Science.gov (United States)

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

2013-11-01

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

Agmatine Reduces Lipopolysaccharide-Mediated Oxidant Response via Activating PI3K/Akt Pathway and Up-Regulating Nrf2 and HO-1 Expression in Macrophages.

Directory of Open Access Journals (Sweden)

Jianshen Chai

Full Text Available Macrophages are key responders of inflammation and are closely related with oxidative stress. Activated macrophages can enhance oxygen depletion, which causes an overproduction of reactive oxygen species (ROS and leads to further excessive inflammatory response and tissue damage. Agmatine, an endogenous metabolite of L-arginine, has recently been shown to have neuroprotective effects based on its antioxidant properties. However, the antioxidant effects of agmatine in peripheral tissues and cells, especially macrophages, remain unclear. In this study we explored the role of agmatine in mediating antioxidant effects in RAW 264.7 cells and studied its antioxidant mechanism. Our data demonstrate that agmatine is an activator of Nrf2 signaling that markedly enhances Nrf2 nuclear translocation, increases nuclear Nrf2 protein level, up-regulates the expression of the Nrf2 downstream effector HO-1, and attenuates ROS generation induced by Lipopolysaccharide (LPS. We further demonstrated that the agmatine-induced activation of Nrf2 is likely through the PI3K/Akt pathway. LY294002, a specific PI3K/Akt inhibitor, abolished agmatine-induced HO-1 up-regulation and ROS suppression significantly. Inhibiting HO-1 pathway significantly attenuated the antioxidant effect of agmatine which the products of HO-1 enzymatic activity contributed to. Furthermore, the common membrane receptors of agmatine were evaluated, revealing that α2-adrenoceptor, I1-imidazoline receptor or I2-imidazoline receptor are not required by the antioxidant properties of agmatine. Taken together, our findings revealed that agmatine has antioxidant activity against LPS-induced ROS accumulation in RAW 264.7 cells involving HO-1 expression induced by Nrf2 via PI3K/Akt pathway activation.

Scavenger receptor B1 facilitates macrophage uptake of silver nanoparticles and cellular activation

Energy Technology Data Exchange (ETDEWEB)

Aldossari, Abdullah A.; Shannahan, Jonathan H. [The University of Colorado Anschutz Medical Campus, Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences (United States); Podila, Ramakrishna [Clemson University, Department of Physics and Astronomy (United States); Brown, Jared M., E-mail: jared.brown@ucdenver.edu [The University of Colorado Anschutz Medical Campus, Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences (United States)

2015-07-15

Due to increased use of silver nanoparticles (AgNPs) for their antimicrobial activity, concerns have risen regarding potential adverse human health effects. Scavenger receptor B1 (SR-B1), a major receptor for high-density lipoprotein (HDL), is expressed by macrophages and has also been reported to play a role in recognition of negatively charged particles. We, therefore, hypothesized that SR-B1 mediates macrophage uptake of AgNPs and inflammatory activation. To test this hypothesis, we exposed a mouse macrophage cell line RAW264.7 (RAW) and bone marrow-derived macrophages (BMDM) to 20 nm citrate-suspended AgNPs. To verify the role of the SR-B1 receptor, we utilized a SR-B1 inhibitor (Blt2). In vitro studies demonstrated uptake of AgNPs and HDL-coated AgNPs by macrophages which were significantly reduced following pretreatment with Blt2. Inflammatory cytokine arrays revealed that macrophages exposed to AgNPs up-regulated expression of Tnf-α, Oncostatin m (OSM), Ccl4, Il17f, Ccl7, and Ccl2, whereas Il16 was found to be down-regulated. Macrophage activation was observed following AgNP and HDL-coated AgNP exposure as measured by OSM protein production and increased surface expression of CD86. These markers of activation were reduced with Blt2 pretreatment. The in vitro findings were confirmed in vivo through pulmonary instillation of AgNPs in mice. Pulmonary instillation of AgNPs resulted in a recruitment of inflammatory cells that were reduced in SR-B1-deficient mice or following Blt2 pretreatment. This study suggests that SR-B1 plays a major role in cellular recognition of AgNPs and the induction of cell responses that could contribute to inflammation caused by AgNP exposure.

Deubiquitinase USP12 promotes LPS induced macrophage responses through inhibition of IκBα

International Nuclear Information System (INIS)

Nayak, Tapan Kumar Singh; Alamuru-Yellapragada, Neeraja P.; Parsa, Kishore V.L.

2017-01-01

Post translational modifications, ubiquitination and its reversal by deubiquitination play an important role in regulating innate immune system. USP12 is a poorly studied deubiquitinase reported to regulate T-cell receptor signalling however the functional role of USP12 in macrophages, the principal architects of inflammation, is unknown. Thus, in this study we probed the involvement of USP12 in macrophage mediated inflammatory responses using bacterial endotoxin, LPS, as the model system. Here, we observed that the expression of USP12 was altered in time dependent manner in LPS stimulated RAW 264.7 macrophages at both mRNA and protein levels as revealed by qPCR and western blot analysis, respectively. Further analysis showed that LPS reduced the levels of Sp1 which enhanced the transcriptional levels of USP12. We observed that siRNA mediated ablation of USP12 expression in mouse macrophages suppressed the induction of LPS-induced iNOS and IL-6 expression but failed to alter IFN-β synthesis, oxidative stress and phagocytic ability of macrophages. Mechanistic analysis suggest that USP12 may be required for the activation of NFκB pathway as knockdown of USP12 reduced the inhibitory phosphorylation of IκBα, a well characterized inhibitor of NFκB nuclear translocation. Further, USP12 was observed to be required for LPS elicited phosphorylation of ERK1/2 and p38. Collectively, our data suggest that USP12 may be a key mediator of LPS stimulated macrophage responses. - Highlights: • USP12 levels are significantly altered in LPS stimulated macrophages. • USP12 is required for LPS induced iNOS and IL6 expression. • USP12 is crucial for LPS induced phosphorylation of IκBα, ERK1/2, p38.

Scavenger receptor B1 facilitates macrophage uptake of silver nanoparticles and cellular activation

Science.gov (United States)

Aldossari, Abdullah A.; Shannahan, Jonathan H.; Podila, Ramakrishna; Brown, Jared M.

2015-07-01

Due to increased use of silver nanoparticles (AgNPs) for their antimicrobial activity, concerns have risen regarding potential adverse human health effects. Scavenger receptor B1 (SR-B1), a major receptor for high-density lipoprotein (HDL), is expressed by macrophages and has also been reported to play a role in recognition of negatively charged particles. We, therefore, hypothesized that SR-B1 mediates macrophage uptake of AgNPs and inflammatory activation. To test this hypothesis, we exposed a mouse macrophage cell line RAW264.7 (RAW) and bone marrow-derived macrophages (BMDM) to 20 nm citrate-suspended AgNPs. To verify the role of the SR-B1 receptor, we utilized a SR-B1 inhibitor (Blt2). In vitro studies demonstrated uptake of AgNPs and HDL-coated AgNPs by macrophages which were significantly reduced following pretreatment with Blt2. Inflammatory cytokine arrays revealed that macrophages exposed to AgNPs up-regulated expression of Tnf- α, Oncostatin m (OSM), Ccl4, Il17f, Ccl7, and Ccl2, whereas Il16 was found to be down-regulated. Macrophage activation was observed following AgNP and HDL-coated AgNP exposure as measured by OSM protein production and increased surface expression of CD86. These markers of activation were reduced with Blt2 pretreatment. The in vitro findings were confirmed in vivo through pulmonary instillation of AgNPs in mice. Pulmonary instillation of AgNPs resulted in a recruitment of inflammatory cells that were reduced in SR-B1-deficient mice or following Blt2 pretreatment. This study suggests that SR-B1 plays a major role in cellular recognition of AgNPs and the induction of cell responses that could contribute to inflammation caused by AgNP exposure.

The β-Hemolysin and Intracellular Survival of Streptococcus agalactiae in Human Macrophages

Science.gov (United States)

Sagar, Anubha; Klemm, Carolin; Hartjes, Lara; Mauerer, Stefanie; van Zandbergen, Ger; Spellerberg, Barbara

2013-01-01

S. agalactiae (group B streptococci, GBS) is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches. PMID:23593170

The β-hemolysin and intracellular survival of Streptococcus agalactiae in human macrophages.

Directory of Open Access Journals (Sweden)

Anubha Sagar

Full Text Available S. agalactiae (group B streptococci, GBS is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches.

A role for the RNA chaperone Hfq in controlling adherent-invasive Escherichia coli colonization and virulence.

Directory of Open Access Journals (Sweden)

Karina T Simonsen

Full Text Available Adherent-invasive Escherichia coli (AIEC has been linked with the onset and perpetuation of inflammatory bowel diseases. The AIEC strain LF82 was originally isolated from an ileal biopsy from a patient with Crohn's disease. The pathogenesis of LF82 results from its abnormal adherence to and subsequent invasion of the intestinal epithelium coupled with its ability to survive phagocytosis by macrophages once it has crossed the intestinal barrier. To gain further insight into AIEC pathogenesis we employed the nematode Caenorhabditis elegans as an in vivo infection model. We demonstrate that AIEC strain LF82 forms a persistent infection in C. elegans, thereby reducing the host lifespan significantly. This host killing phenotype was associated with massive bacterial colonization of the nematode intestine and damage to the intestinal epithelial surface. C. elegans killing was independent of known LF82 virulence determinants but was abolished by deletion of the LF82 hfq gene, which encodes an RNA chaperone involved in mediating posttranscriptional gene regulation by small non-coding RNAs. This finding reveals that important aspects of LF82 pathogenesis are controlled at the posttranscriptional level by riboregulation. The role of Hfq in LF82 virulence was independent of its function in regulating RpoS and RpoE activity. Further, LF82Δhfq mutants were non-motile, impaired in cell invasion and highly sensitive to various chemical stress conditions, reinforcing the multifaceted function of Hfq in mediating bacterial adaptation. This study highlights the usefulness of simple non-mammalian infection systems for the identification and analysis of bacterial virulence factors.

ER-α36 mediates estrogen-stimulated MAPK/ERK activation and regulates migration, invasion, proliferation in cervical cancer cells

International Nuclear Information System (INIS)

Sun, Qing; Liang, Ying; Zhang, Tianli; Wang, Kun; Yang, Xingsheng

2017-01-01

Objective: Estrogen receptor alpha 36 (ER-α36), a truncated variant of ER-α, is different from other nuclear receptors of the ER-α family. Previous findings indicate that ER-α36 might be involved in cell growth, proliferation, and differentiation in carcinomas and primarily mediates non-genomic estrogen signaling. However, studies on ER-α36 and cervical cancer are rare. This study aimed to detect the expression of ER-α36 in cervical cancer; the role of ER-α36 in 17-β-estradiol (E2)-induced invasion, migration and proliferation of cervical cancer; and their probable molecular mechanisms. Methods: Immunohistochemistry and immunofluorescence were used to determine the location of ER-α36 in cervical cancer tissues and cervical cell lines. CaSki and HeLa cell lines were transfected with lentiviruses to establish stable cell lines with knockdown and overexpression of ER-α36. Wound healing assay, transwell invasion assay, and EdU incorporation proliferation assay were performed to evaluate the migration, invasion, and proliferation ability. The phosphorylation levels of mitogen-activated protein kinases/extracellular signal-regulated kinase (MAPK/ERK) signaling molecules were examined with western blot analysis. Results: ER-α36 expression was detected in both cervical cell lines and cervical cancer tissues. Downregulation of ER-α36 significantly inhibited cell invasion, migration, and proliferation. Moreover, upregulation of ER-α36 increased the invasion, migration, and proliferation ability of CaSki and HeLa cell lines. ER-α36 mediates estrogen-stimulated MAPK/ERK activation. Conclusion: ER-α36 is localized on the plasma membrane and cytoplasm in both cervical cancer tissues and cell lines. ER-α36 mediates estrogen-stimulated MAPK/ERK activation and regulates migration, invasion, proliferation in cervical cancer cells. - Highlights: • ER-α36 is expressed on both cervical cell lines and cervical cancer tissues. • ER-α36 mediates estrogen

Chemokine receptor CXCR7 regulates the invasion, angiogenesis and tumor growth of human hepatocellular carcinoma cells

Directory of Open Access Journals (Sweden)

Li Fan

2010-04-01

Full Text Available Abstract Background In spite of recent advances in diagnostic and therapeutic measures, the prognosis of hepatocellular carcinoma (HCC patients remains poor. Therefore, it is crucial to understand what factors are involved in promoting development of HCC. Evidence is accumulating that members of the chemokine receptor family are viewed as promising therapeutic targets in the fight against cancer. More recent studies have revealed that chemokine receptor CXCR7 plays an important role in cancer development. However, little is known about the effect of CXCR7 on the process of HCC cell invasion and angiogenesis. The aim of this study is to investigate the expression of CXCR7 in hepatocellular carcinoma tissues and cell lines and to evaluate the role of CXCR7 in tumor growth, angiogenesis and invasion of HCC cells. Methods We constructed CXCR7 expressing shRNA, and CXCR7shRNA was subsequently stably transfected into human HCC cells. We evaluated the effect of CXCR7 inhibition on cell invasion, adhesion, VEGF secretion, tube formation and tumor growth. Immunohistochemistry was done to assess the expression of CXCR7 in human hepatocellular carcinoma tissues and CD31 in tumor of mice. We also evaluated the effect of VEGF stimulation on expression of CXCR7. Results CXCR7 was overexpressed in hepatocellular carcinoma tissues. We showed that high invasive potential HCC cell lines express high levels of CXCR7. In vitro, CXCL12 was found to induce invasion, adhesion, tube formation, and VEGF secretion in SMMC-7721 cells. These biological effects were inhibited by silencing of CXCR7 in SMMC-7721 cells. In addition, we also found that VEGF stimulation can up-regulate CXCR7 expression in SMMC-7721 cells and HUVECs. More importantly, enhanced expression of CXCR7 by VEGF was founctional. In vivo, tumor growth and angiogenesis were suppressed by knockdown of CXCR7 in SMMC-7721 cells. However, silencing of CXCR7 did not affect metastasis of tumor in vivo

Phenylbutyrate Is Bacteriostatic against Mycobacterium tuberculosis and Regulates the Macrophage Response to Infection, Synergistically with 25-Hydroxy-Vitamin D3.

Directory of Open Access Journals (Sweden)

Anna K Coussens

2015-07-01

Full Text Available Adjunctive vitamin D treatment for pulmonary tuberculosis enhances resolution of inflammation but has modest effects on bacterial clearance. Sodium 4-phenylbutyrate (PBA is in clinical use for a range of conditions and has been shown to synergise with vitamin D metabolites to upregulate cathelicidin antimicrobial peptide (CAMP expression. We investigated whether clinically attainable plasma concentrations of PBA (0.4-4 mM directly affect Mycobacterium tuberculosis (Mtb growth and human macrophage and PBMC response to infection. We also tested the ability of PBA to enhance the immunomodulatory actions of the vitamin D metabolite 25(OHD3 during infection and synergistically inhibit intracellular Mtb growth. PBA inhibited Mtb growth in broth with an MIC99 of 1 mM, which was reduced to 0.25 mM by lowering pH. During human macrophage infection, PBA treatment restricted Mtb uptake, phagocytic receptor expression and intracellular growth in a dose-dependent manner. PBA independently regulated CCL chemokine secretion and induced expression of the antimicrobial LTF (lactoferrin, the anti-inflammatory PROC (protein C and multiple genes within the NLRP3 inflammasome pathway. PBA co-treatment with 25(OHD3 synergistically modulated expression of numerous vitamin D-response genes, including CAMP, CYP24A1, CXCL10 and IL-37. This synergistic effect was dependent on MAPK signalling, while the effect of PBA on LTF, PROC and NLRP3 was MAPK-independent. During PBA and 25(OHD3 co-treatment of human macrophages, in the absence of exogenous proteinase 3 (PR3 to activate cathelicidin, Mtb growth restriction was dominated by the effect of PBA, while the addition of PR3 enhanced growth restriction by 25(OHD3 and PBA co-treatment. This suggests that PBA augments vitamin D-mediated cathelicidin-dependent Mtb growth restriction by human macrophages and independently induces antimicrobial and anti-inflammatory action. Therefore through both host-directed and bacterial