Fractional Factorial Design to Investigate Stromal Cell Regulation of Macrophage Plasticity

PubMed Central

Barminko, Jeffrey A.; Nativ, Nir I.; Schloss, Rene; Yarmush, Martin L.

2018-01-01

Understanding the regulatory networks which control specific macrophage phenotypes is essential in identifying novel targets to correct macrophage mediated clinical disorders, often accompanied by inflammatory events. Since mesenchymal stromal cells (MSCs) have been shown to play key roles in regulating immune functions predominantly via a large number of secreted products, we used a fractional factorial approach to streamline experimental evaluation of MSC mediated inflammatory macrophage regulation. Our macrophage reprogramming metrics, human bone marrow MSC attenuation of macrophage pro-inflammatory M1 TNFα secretion and simultaneous enhanced expression of the M2 macrophage marker, CD206, were used as analysis endpoints. Objective evaluation of a panel of MSC secreted mediators indicated that PGE2 alone was sufficient in facilitating macrophage reprogramming, while IL4 only provided partial reprogramming. Inhibiting stromal cell PGE2 secretion with Indomethacin, reversed the macrophage reprogramming effect. PGE2 reprogramming was mediated through the EP4 receptor and indirectly through the CREB signaling pathway as GSK3 specific inhibitors induced M1 macrophages to express CD206. This reprogramming pathway functioned independently from the M1 suppression pathway, as neither CREB nor GSK3 inhibition reversed PGE2 TNF-α secretion attenuation. In conclusion, fractional factorial experimental design identified stromal derived PGE2 as the factor most important in facilitating macrophage reprogramming, albeit via two unique pathways. PMID:24891120

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

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

Cheng, Yahui; Tian, Yuanyao; Xia, Jialu

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 significantlymore » 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.« less

Lysine Deacetylases and Regulated Glycolysis in Macrophages.

PubMed

Shakespear, Melanie R; Iyer, Abishek; Cheng, Catherine Youting; Das Gupta, Kaustav; Singhal, Amit; Fairlie, David P; Sweet, Matthew J

2018-06-01

Regulated cellular metabolism has emerged as a fundamental process controlling macrophage functions, but there is still much to uncover about the precise signaling mechanisms involved. Lysine acetylation regulates the activity, stability, and/or localization of metabolic enzymes, as well as inflammatory responses, in macrophages. Two protein families, the classical zinc-dependent histone deacetylases (HDACs) and the NAD-dependent HDACs (sirtuins, SIRTs), mediate lysine deacetylation. We describe here mechanisms by which classical HDACs and SIRTs directly regulate specific glycolytic enzymes, as well as evidence that links these protein deacetylases to the regulation of glycolysis-related genes. In these contexts, we discuss HDACs and SIRTs as key control points for regulating immunometabolism and inflammatory outputs from macrophages. Copyright © 2018 Elsevier Ltd. All rights reserved.

The role of the local environment and epigenetics in shaping macrophage identity and their effect on tissue homeostasis.

PubMed

Amit, Ido; Winter, Deborah R; Jung, Steffen

2016-01-01

Macrophages provide a critical systemic network cells of the innate immune system. Emerging data suggest that in addition, they have important tissue-specific functions that range from clearance of surfactant from the lungs to neuronal pruning and establishment of gut homeostasis. The differentiation and tissue-specific activation of macrophages require precise regulation of gene expression, a process governed by epigenetic mechanisms such as DNA methylation, histone modification and chromatin structure. We argue that epigenetic regulation of macrophages is determined by lineage- and tissue-specific transcription factors controlled by the built-in programming of myeloid development in combination with signaling from the tissue environment. Perturbation of epigenetic mechanisms of tissue macrophage identity can affect normal macrophage tissue function and contribute to pathologies ranging from obesity and autoimmunity to neurodegenerative diseases.

Mesenchymal stem cells use extracellular vesicles to outsource mitophagy and shuttle microRNAs

PubMed Central

Phinney, Donald G.; Di Giuseppe, Michelangelo; Njah, Joel; Sala, Ernest; Shiva, Sruti; St Croix, Claudette M.; Stolz, Donna B.; Watkins, Simon C.; Di, Y. Peter; Leikauf, George D.; Kolls, Jay; Riches, David W. H.; Deiuliis, Giuseppe; Kaminski, Naftali; Boregowda, Siddaraju V.; McKenna, David H.; Ortiz, Luis A.

2015-01-01

Mesenchymal stem cells (MSCs) and macrophages are fundamental components of the stem cell niche and function coordinately to regulate haematopoietic stem cell self-renewal and mobilization. Recent studies indicate that mitophagy and healthy mitochondrial function are critical to the survival of stem cells, but how these processes are regulated in MSCs is unknown. Here we show that MSCs manage intracellular oxidative stress by targeting depolarized mitochondria to the plasma membrane via arrestin domain-containing protein 1-mediated microvesicles. The vesicles are then engulfed and re-utilized via a process involving fusion by macrophages, resulting in enhanced bioenergetics. Furthermore, we show that MSCs simultaneously shed micro RNA-containing exosomes that inhibit macrophage activation by suppressing Toll-like receptor signalling, thereby de-sensitizing macrophages to the ingested mitochondria. Collectively, these studies mechanistically link mitophagy and MSC survival with macrophage function, thereby providing a physiologically relevant context for the innate immunomodulatory activity of MSCs. PMID:26442449

Mesenchymal stem cells use extracellular vesicles to outsource mitophagy and shuttle microRNAs.

PubMed

Phinney, Donald G; Di Giuseppe, Michelangelo; Njah, Joel; Sala, Ernest; Shiva, Sruti; St Croix, Claudette M; Stolz, Donna B; Watkins, Simon C; Di, Y Peter; Leikauf, George D; Kolls, Jay; Riches, David W H; Deiuliis, Giuseppe; Kaminski, Naftali; Boregowda, Siddaraju V; McKenna, David H; Ortiz, Luis A

2015-10-07

Mesenchymal stem cells (MSCs) and macrophages are fundamental components of the stem cell niche and function coordinately to regulate haematopoietic stem cell self-renewal and mobilization. Recent studies indicate that mitophagy and healthy mitochondrial function are critical to the survival of stem cells, but how these processes are regulated in MSCs is unknown. Here we show that MSCs manage intracellular oxidative stress by targeting depolarized mitochondria to the plasma membrane via arrestin domain-containing protein 1-mediated microvesicles. The vesicles are then engulfed and re-utilized via a process involving fusion by macrophages, resulting in enhanced bioenergetics. Furthermore, we show that MSCs simultaneously shed micro RNA-containing exosomes that inhibit macrophage activation by suppressing Toll-like receptor signalling, thereby de-sensitizing macrophages to the ingested mitochondria. Collectively, these studies mechanistically link mitophagy and MSC survival with macrophage function, thereby providing a physiologically relevant context for the innate immunomodulatory activity of MSCs.

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

PubMed

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.

Inhibition of IRF8 Negatively Regulates Macrophage Function and Impairs Cutaneous Wound Healing.

PubMed

Guo, Yuanyuan; Yang, Zhiyin; Wu, Shan; Xu, Peng; Peng, Yinbo; Yao, Min

2017-02-01

The inflammatory response is essential for normal cutaneous wound healing. Macrophages, as critical inflammatory cells, coordinate inflammation and angiogenesis phases during wound healing. It has been reported that the transcription factor interferon regulatory factor 8 (IRF8), a member of the IRF family, plays a critical role in the development and function of macrophages and is associated with inflammation. However, the role of IRF8 in cutaneous wound healing and its underlying mechanism remain elusive. Through immunohistochemical (IHC) staining, we showed that IRF8 is involved in the wound repair process in mice and patients. Furthermore, we ascertain that the repression of IRF8 by small interfering RNA (siRNA) leads to delayed wound healing. To explore the mechanism by which IRF8 impacts wound healing, we observed its effect on macrophage-related mediators by IHC or real-time PCR. The results demonstrated that the inhibition of IRF8 decreases the mRNA expression of inflammatory mediators associated with M1 macrophage (il-1b, il-6, inos, and tnf-a) but no impact on M2 macrophage-related mediators (arg-1, mrc-1, and il-10) and the number of macrophages in the wounds. Furthermore, the inhibition of IRF8 induced apoptosis in the wounds. In summary, this study demonstrates that the down-regulation of IRF8 in the wound leads to impaired wound healing possibly through the regulation of macrophage function and apoptosis in skin wound.

p21 mediates macrophage reprogramming through regulation of p50-p50 NF-κB and IFN-β

PubMed Central

Hernández-Jiménez, Enrique; Shokri, Rahman; Carmona-Rodríguez, Lorena; Mañes, Santos; Álvarez-Mon, Melchor; López-Collazo, Eduardo; Martínez-A, Carlos

2016-01-01

M1 and M2 macrophage phenotypes, which mediate proinflammatory and antiinflammatory functions, respectively, represent the extremes of immunoregulatory plasticity in the macrophage population. This plasticity can also result in intermediate macrophage states that support a balance between these opposing functions. In sepsis, M1 macrophages can compensate for hyperinflammation by acquiring an M2-like immunosuppressed status that increases the risk of secondary infection and death. The M1 to M2 macrophage reprogramming that develops during LPS tolerance resembles the pathological antiinflammatory response to sepsis. Here, we determined that p21 regulates macrophage reprogramming by shifting the balance between active p65-p50 and inhibitory p50-p50 NF-κB pathways. p21 deficiency reduced the DNA-binding affinity of the p50-p50 homodimer in LPS-primed and -rechallenged macrophages, impairing their ability to attenuate IFN-β production and acquire an M2-like hyporesponsive status. High p21 levels in sepsis patients correlated with low IFN-β expression, and p21 knockdown in human monocytes corroborated its role in IFN-β regulation. The data demonstrate that p21 adjusts the equilibrium between p65-p50 and p50-p50 NF-κB pathways to mediate macrophage plasticity in LPS tolerance. Identifying p21-related pathways involved in monocyte reprogramming may lead to potential targets for sepsis treatment. PMID:27427981

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

PubMed

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.

Akt-mTORC1 signaling regulates Acly to integrate metabolic input to control of macrophage activation

PubMed Central

Covarrubias, Anthony J; Aksoylar, Halil Ibrahim; Yu, Jiujiu; Snyder, Nathaniel W; Worth, Andrew J; Iyer, Shankar S; Wang, Jiawei; Ben-Sahra, Issam; Byles, Vanessa; Polynne-Stapornkul, Tiffany; Espinosa, Erika C; Lamming, Dudley; Manning, Brendan D; Zhang, Yijing; Blair, Ian A; Horng, Tiffany

2016-01-01

Macrophage activation/polarization to distinct functional states is critically supported by metabolic shifts. How polarizing signals coordinate metabolic and functional reprogramming, and the potential implications for control of macrophage activation, remains poorly understood. Here we show that IL-4 signaling co-opts the Akt-mTORC1 pathway to regulate Acly, a key enzyme in Ac-CoA synthesis, leading to increased histone acetylation and M2 gene induction. Only a subset of M2 genes is controlled in this way, including those regulating cellular proliferation and chemokine production. Moreover, metabolic signals impinge on the Akt-mTORC1 axis for such control of M2 activation. We propose that Akt-mTORC1 signaling calibrates metabolic state to energetically demanding aspects of M2 activation, which may define a new role for metabolism in supporting macrophage activation. DOI: http://dx.doi.org/10.7554/eLife.11612.001 PMID:26894960

Elevated Mirc1/Mir17-92 cluster expression negatively regulates autophagy and CFTR (cystic fibrosis transmembrane conductance regulator) function in CF macrophages.

PubMed

Tazi, Mia F; Dakhlallah, Duaa A; Caution, Kyle; Gerber, Madelyn M; Chang, Sheng-Wei; Khalil, Hany; Kopp, Benjamin T; Ahmed, Amr E; Krause, Kathrin; Davis, Ian; Marsh, Clay; Lovett-Racke, Amy E; Schlesinger, Larry S; Cormet-Boyaka, Estelle; Amer, Amal O

2016-11-01

Cystic fibrosis (CF) is a fatal, genetic disorder that critically affects the lungs and is directly caused by mutations in the CF transmembrane conductance regulator (CFTR) gene, resulting in defective CFTR function. Macroautophagy/autophagy is a highly regulated biological process that provides energy during periods of stress and starvation. Autophagy clears pathogens and dysfunctional protein aggregates within macrophages. However, this process is impaired in CF patients and CF mice, as their macrophages exhibit limited autophagy activity. The study of microRNAs (Mirs), and other noncoding RNAs, continues to offer new therapeutic targets. The objective of this study was to elucidate the role of Mirs in dysregulated autophagy-related genes in CF macrophages, and then target them to restore this host-defense function and improve CFTR channel function. We identified the Mirc1/Mir17-92 cluster as a potential negative regulator of autophagy as CF macrophages exhibit decreased autophagy protein expression and increased cluster expression when compared to wild-type (WT) counterparts. The absence or reduced expression of the cluster increases autophagy protein expression, suggesting the canonical inverse relationship between Mirc1/Mir17-92 and autophagy gene expression. An in silico study for targets of Mirs that comprise the cluster suggested that the majority of the Mirs target autophagy mRNAs. Those targets were validated by luciferase assays. Notably, the ability of macrophages expressing mutant F508del CFTR to transport halide through their membranes is compromised and can be restored by downregulation of these inherently elevated Mirs, via restoration of autophagy. In vivo, downregulation of Mir17 and Mir20a partially restored autophagy expression and hence improved the clearance of Burkholderia cenocepacia. Thus, these data advance our understanding of mechanisms underlying the pathobiology of CF and provide a new therapeutic platform for restoring CFTR function and autophagy in patients with CF.

Emodin Bidirectionally Modulates Macrophage Polarization and Epigenetically Regulates Macrophage Memory.

PubMed

Iwanowycz, Stephen; Wang, Junfeng; Altomare, Diego; Hui, Yvonne; Fan, Daping

2016-05-27

Macrophages are pleiotropic cells capable of performing a broad spectrum of functions. Macrophage phenotypes are classified along a continuum between the extremes of proinflammatory M1 macrophages and anti-inflammatory M2 macrophages. The seemingly opposing functions of M1 and M2 macrophages must be tightly regulated for an effective and proper response to foreign molecules or damaged tissue. Excessive activation of either M1 or M2 macrophages contributes to the pathology of many diseases. Emodin is a Chinese herb-derived compound and has shown potential to inhibit inflammation in various settings. In this study, we tested the ability of emodin to modulate the macrophage response to both M1 and M2 stimuli. Primary mouse macrophages were stimulated with LPS/IFNγ or IL4 with or without emodin, and the effects of emodin on gene transcription, cell signaling pathways, and histone modifications were examined by a variety of approaches, including microarray, quantitative real-time PCR, Western blotting, chromatin immunoprecipitation, and functional assays. We found that emodin bidirectionally tunes the induction of LPS/IFNγ- and IL4-responsive genes through inhibiting NFκB/IRF5/STAT1 signaling and IRF4/STAT6 signaling, respectively. Thereby, emodin modulates macrophage phagocytosis, migration, and NO production. Furthermore, emodin inhibited the removal of H3K27 trimethylation (H3K27m3) marks and the addition of H3K27 acetylation (H3K27ac) marks on genes required for M1 or M2 polarization of macrophages. In conclusion, our data suggest that emodin is uniquely able to suppress the excessive response of macrophages to both M1 and M2 stimuli and therefore has the potential to restore macrophage homeostasis in various pathologies. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Emodin Bidirectionally Modulates Macrophage Polarization and Epigenetically Regulates Macrophage Memory*

PubMed Central

Iwanowycz, Stephen; Wang, Junfeng; Altomare, Diego; Hui, Yvonne; Fan, Daping

2016-01-01

Macrophages are pleiotropic cells capable of performing a broad spectrum of functions. Macrophage phenotypes are classified along a continuum between the extremes of proinflammatory M1 macrophages and anti-inflammatory M2 macrophages. The seemingly opposing functions of M1 and M2 macrophages must be tightly regulated for an effective and proper response to foreign molecules or damaged tissue. Excessive activation of either M1 or M2 macrophages contributes to the pathology of many diseases. Emodin is a Chinese herb-derived compound and has shown potential to inhibit inflammation in various settings. In this study, we tested the ability of emodin to modulate the macrophage response to both M1 and M2 stimuli. Primary mouse macrophages were stimulated with LPS/IFNγ or IL4 with or without emodin, and the effects of emodin on gene transcription, cell signaling pathways, and histone modifications were examined by a variety of approaches, including microarray, quantitative real-time PCR, Western blotting, chromatin immunoprecipitation, and functional assays. We found that emodin bidirectionally tunes the induction of LPS/IFNγ- and IL4-responsive genes through inhibiting NFκB/IRF5/STAT1 signaling and IRF4/STAT6 signaling, respectively. Thereby, emodin modulates macrophage phagocytosis, migration, and NO production. Furthermore, emodin inhibited the removal of H3K27 trimethylation (H3K27m3) marks and the addition of H3K27 acetylation (H3K27ac) marks on genes required for M1 or M2 polarization of macrophages. In conclusion, our data suggest that emodin is uniquely able to suppress the excessive response of macrophages to both M1 and M2 stimuli and therefore has the potential to restore macrophage homeostasis in various pathologies. PMID:27008857

[Immune regulation activity and mechanism of Tibetan Kefir exopolysaccharide fractions].

PubMed

Meng, Li; Zhang, Lanwei

2009-12-01

To investigate the effects and mechanism on immune regulation activity in mice of two Tibetan Kefir exoploysaccharides (EPS) with different molecular weight of 0.1 x 10(5) - 3 x 10(5) (fraction 1) and 1.8 x 10(3) (fraction 2). The immune regulation activity experiment was carried out in vitro based on the Functional Assessment Procedure and Test Methods of Health Food, which was issued by Ministry of Health of China. First, we treated mice subjects with EPS at doses of 40 mg/kg, 80 mg/kg, 120 mg/kg through ig. Then we detected the index of immune organs, the ability of antibody production (tested by HC50), activity of NK cell, delayed type hypersensitivity (DTH) and phagocytosis of macrophage in mice. Finally, we examined the expression of Erk protein in Macrophages by Western Blot assay. Fraction 1 could promote HC50, activity of NK cell and DTH in mice which low dose showed better. Fraction 2 could promote DTH, phagocytosis of macrophage which high dose showed better. The expression of Erk and COX-2 had the same trend with Phagocytic index. We verified the two fractions of Tibetan Kefir EPS could enhance immune functions in mice. Fraction 1 regulated immune function through NK cell and B cell while fraction 2 through macrophage cell and T cell. The effects to macrophage of Tibetan Kefir EPS in mice may realize through extra cellular signal-regulated kinase Erk pathway.

Chemoattractant signaling between tumor cells and macrophages regulates cancer cell migration, metastasis and neovascularization.

PubMed

Green, Chad E; Liu, Tiffany; Montel, Valerie; Hsiao, Gene; Lester, Robin D; Subramaniam, Shankar; Gonias, Steven L; Klemke, Richard L

2009-08-21

Tumor-associated macrophages are known to influence cancer progression by modulation of immune function, angiogenesis, and cell metastasis, however, little is known about the chemokine signaling networks that regulate this process. Utilizing CT26 colon cancer cells and RAW 264.7 macrophages as a model cellular system, we demonstrate that treatment of CT26 cells with RAW 264.7 conditioned medium induces cell migration, invasion and metastasis. Inflammatory gene microarray analysis indicated CT26-stimulated RAW 264.7 macrophages upregulate SDF-1alpha and VEGF, and that these cytokines contribute to CT26 migration in vitro. RAW 264.7 macrophages also showed a robust chemotactic response towards CT26-derived chemokines. In particular, microarray analysis and functional testing revealed CSF-1 as the major chemoattractant for RAW 264.7 macrophages. Interestingly, in the chick CAM model of cancer progression, RAW 264.7 macrophages localized specifically to the tumor periphery where they were found to increase CT26 tumor growth, microvascular density, vascular disruption, and lung metastasis, suggesting these cells home to actively invading areas of the tumor, but not the hypoxic core of the tumor mass. In support of these findings, hypoxic conditions down regulated CSF-1 production in several tumor cell lines and decreased RAW 264.7 macrophage migration in vitro. Together our findings suggest a model where normoxic tumor cells release CSF-1 to recruit macrophages to the tumor periphery where they secrete motility and angiogenic factors that facilitate tumor cell invasion and metastasis.

α- and β-d-Glucans from the edible mushroom Pleurotus albidus differentially regulate lipid-induced inflammation and foam cell formation in human macrophage-like THP-1 cells.

PubMed

Castro-Alves, Victor Costa; Nascimento, João Roberto Oliveira do

2018-05-01

Macrophages play an essential role in lipid metabolism; however, the excessive uptake of modified lipids and cholesterol crystals (CC) leads to the formation of pro-inflammatory lipid-laden macrophages called foam cells. Since the α-1,6- and β-1,3-d-glucans from the basidiome and the mycelium of the edible mushroom Pleurotus albidus have previously been shown to regulate macrophage function, these glucans were tested in macrophage-like THP-1 cells previously exposed to acetylated low-density lipoproteins (acLDL) or CC. The glucans inhibited lipid-induced inflammation, but only the β-1,3-d-glucan regulated both the NLRP3 inflammasome activation and the expression of genes involved on lipid efflux in acLDL- or CC-pretreated cells, thereby reducing foam cell formation. In contrast, the two α-1,6-glucans tested inhibited foam cell formation only in acLDL-pretreated cells and had no effect on the expression of the peroxisome proliferator-activated receptor gamma and liver X receptor alpha genes, suggesting that these glucans regulate lipid influx rather than lipid efflux. Thus, α- and β-d-glucans differentially regulate lipid-induced inflammation and foam cell formation in macrophage-like cells. Furthermore, results emphasize that P. albidus has potential to be used as a functional food or as a source for the extraction of biologically-active glucans. Copyright © 2018 Elsevier B.V. All rights reserved.

Spleen lymphocyte function modulated by a cocoa-enriched diet.

PubMed

Ramiro-Puig, E; Pérez-Cano, F J; Ramírez-Santana, C; Castellote, C; Izquierdo-Pulido, M; Permanyer, J; Franch, A; Castell, M

2007-09-01

Previous studies have shown the down-regulating in vitro effect of cocoa flavonoids on lymphocyte and macrophage activation. In the present paper, we report the capacity of a long-term rich cocoa diet to modulate macrophage cytokine secretion and lymphocyte function in young rats. Weaned rats received natural cocoa (4% or 10% food intake), containing 32 mg flavonoids/g, for 3 weeks. Spleen immune function was then evaluated through the analysis of lymphocyte composition, their proliferative response and their ability to secrete cytokines and Ig. In addition, the status of activated peritoneal macrophages was established through tumour necrosis factor (TNF)-alpha secretion. The richest cocoa diet (10%) caused a reduction of TNF-alpha secretion by peritoneal macrophages showing anti-inflammatory activity. Similarly, although a 10% cocoa diet increased lymphocyte proliferation rate, it down-regulated T helper 2 (Th2)-related cytokines and decreased Ig secretion. These changes were accompanied by an increase in spleen B cell proportion and a decrease in Th cell percentage. In summary, these results demonstrate the functional activity of a cocoa-high dosage in down-regulating the immune response that might be beneficial in hypersensitivity and autoimmunity.

Spleen lymphocyte function modulated by a cocoa-enriched diet

PubMed Central

Ramiro-Puig, E; Pérez-Cano, F J; Ramírez-Santana, C; Castellote, C; Izquierdo-Pulido, M; Permanyer, J; Franch, A; Castell, M

2007-01-01

Previous studies have shown the down-regulating in vitro effect of cocoa flavonoids on lymphocyte and macrophage activation. In the present paper, we report the capacity of a long-term rich cocoa diet to modulate macrophage cytokine secretion and lymphocyte function in young rats. Weaned rats received natural cocoa (4% or 10% food intake), containing 32 mg flavonoids/g, for 3 weeks. Spleen immune function was then evaluated through the analysis of lymphocyte composition, their proliferative response and their ability to secrete cytokines and Ig. In addition, the status of activated peritoneal macrophages was established through tumour necrosis factor (TNF)-α secretion. The richest cocoa diet (10%) caused a reduction of TNF-α secretion by peritoneal macrophages showing anti-inflammatory activity. Similarly, although a 10% cocoa diet increased lymphocyte proliferation rate, it down-regulated T helper 2 (Th2)-related cytokines and decreased Ig secretion. These changes were accompanied by an increase in spleen B cell proportion and a decrease in Th cell percentage. In summary, these results demonstrate the functional activity of a cocoa-high dosage in down-regulating the immune response that might be beneficial in hypersensitivity and autoimmunity. PMID:17565606

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

PubMed

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

2018-04-01

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

Granulocyte-macrophage and macrophage colony-stimulating factors differentially regulate alpha v integrin expression on cultured human macrophages.

PubMed

De Nichilo, M O; Burns, G F

1993-03-15

The colony-stimulating factors (CSFs) greatly influence mature macrophage function in vitro: macrophage (M)-CSF induces maturation of monocytes and enhances differentiated cell function; granulocyte-macrophage (GM)-CSF stimulates a variety of antimicrobial functions. In vivo M-CSF is thought to promote differentiation, and GM-CSF is thought to potentiate the inflammatory response. One mechanism by which these differential effects may be achieved is through the receptor-mediated interaction of macrophages with their extracellular matrix. Here we show that M-CSF induces specifically the expression of the alpha v beta 5 integrin receptor, whereas GM-CSF rapidly induces mRNA and surface expression of the alpha v beta 3 integrin. The M-CSF-treated cells acquire a flattened epitheloid phenotype, and on vitronectin the alpha v beta 5 is located in adhesion plaques. These cells do not bind collagen or laminin. In contrast, cells treated with GM-CSF adopt an elongated phenotype on a number of substrates, including collagen and laminin, and express alpha v beta 3 at the leading edge of cells on vitronectin. These results suggest that a primary means by which the CSFs exert their individual effects on mature cells may be through regulating integrin expression.

Forkhead Box O1 Regulates Macrophage Polarization Following Staphylococcus aureus Infection: Experimental Murine Data and Review of the Literature.

PubMed

Wang, Yu-Chen; Ma, Hong-Di; Yin, Xue-Ying; Wang, Yin-Hu; Liu, Qing-Zhi; Yang, Jing-Bo; Shi, Qing-Hua; Sun, Baolin; Gershwin, M Eric; Lian, Zhe-Xiong

2016-12-01

The functions of macrophages that lead to effective host responses are critical for protection against Staphylococcus aureus. Deep tissue-invading S. aureus initially countered by macrophages trigger macrophage accumulation and induce inflammatory responses through surface receptors, especially toll-like receptor 2 (TLR2). Here, we found that macrophages formed sporadic aggregates in the liver during infection. Within those aggregates, macrophages co-localized with T cells and were indispensable for their infiltration. In addition, we have focused on the mechanisms underlying the polarization of macrophages in Forkhead box transcription factor O1 (FoxO1) conditional knockout Lys Cre/+ FoxO1 fl/fl mice following S. aureus infection and report herein that macrophage M1-M2 polarization via TLR2 is intrinsically regulated by FoxO1. Indeed, for effective FoxO1 activity, stimulation of TLR2 is essential. However, following S. aureus challenge, there was a decrease in macrophage FoxO1, with increased phosphorylation of FoxO1 because of TLR2-mediated activation of PI3K/Akt and c-Raf/MEK/ERK pathway. Following infection in Lys Cre/+ FoxO1 fl/fl mice, mice became more susceptible to S. aureus with reduced macrophage aggregation in the liver and attenuated Th1 and Th17 responses. FoxO1 abrogation reduced M1 pro-inflammatory responses triggered by S. aureus and enhanced M2 polarization in macrophages. In contrast, overexpression of FoxO1 in macrophages increased pro-inflammatory mediators and functional surface molecule expression. In conclusion, macrophage FoxO1 is critical to promote M1 polarization and maintain a competent T cell immune response against S. aureus infection in the liver. FoxO1 regulates macrophage M1-M2 polarization downstream of TLR2 dynamically through phosphorylation.

The macrophage marker translocator protein (TSPO) is down-regulated on pro-inflammatory 'M1' human macrophages.

PubMed

Narayan, Nehal; Mandhair, Harpreet; Smyth, Erica; Dakin, Stephanie Georgina; Kiriakidis, Serafim; Wells, Lisa; Owen, David; Sabokbar, Afsie; Taylor, Peter

2017-01-01

The translocator protein (TSPO) is a mitochondrial membrane protein, of as yet uncertain function. Its purported high expression on activated macrophages, has lent utility to TSPO targeted molecular imaging in the form of positron emission tomography (PET), as a means to detect and quantify inflammation in vivo. However, existing literature regarding TSPO expression on human activated macrophages is lacking, mostly deriving from brain tissue studies, including studies of brain malignancy, and inflammatory diseases such as multiple sclerosis. Here, we utilized three human sources of monocyte derived macrophages (MDM), from THP-1 monocytes, healthy peripheral blood monocytes and synovial fluid monocytes from patients with rheumatoid arthritis, to undertake a detailed investigation of TSPO expression in activated macrophages. In this work, we demonstrate a consistent down-regulation of TSPO mRNA and protein in macrophages activated to a pro-inflammatory, or 'M1' phenotype. Conversely, stimulation of macrophages to an M2 phenotype with IL-4, dexamethasone or TGF-β1 did not alter TSPO expression, regardless of MDM source. The reasons for this are uncertain, but our study findings add some supporting evidence for recent investigations concluding that TSPO may be involved in negative regulation of inflammatory responses in macrophages.

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

PubMed Central

Yang, Yanyan; Yu, Tao; Sung, Gi-Ho; Yoo, Byong Chul

2014-01-01

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. PMID:24771982

Galectin-3 in M2 macrophages plays a protective role in resolution of neuropathology in brain parasitic infection by regulating neutrophil turnover.

PubMed

Quenum Zangbede, Fredice O; Chauhan, Arun; Sharma, Jyotika; Mishra, Bibhuti B

2018-06-26

Macrophages/microglia with M2- activation phenotype are thought to play an important anti-inflammatory and tissue reparative functions in the brain, yet the molecular basis of their functions in the central nervous system (CNS) remain to be clearly defined. In a preclinical model of neurocysticercosis using brain infection with a parasite Mesocestoides corti , we previously reported the presence of large numbers of M2 cells in the CNS. In this study using female mice, we report that M2 macrophages in the parasite-infected brain display abundant galectin-3 expression. Disease severity was increased in Galectin-3 -/- mice correlating with increased neurological defects, augmented cell death and, importantly, massive accumulation of neutrophils and M2 macrophages in the CNS of these mice. Because neutrophil clearance by efferocytosis is an important function of M2 macrophages, we investigated a possible role of galectin-3 in this process. Indeed, galectin-3 deficient M2 macrophages exhibited a defect in efferocytic clearance of neutrophils in-vitro. Furthermore, adoptive transfer of M2 macrophages from Galectin-3 sufficient WT mice reduced neutrophilia in the CNS and ameliorated disease severity in parasite-infected Galectin-3 -/- mice. Together, these results demonstrate for the first time a novel role of galectin-3 in M2 macrophage function in neutrophil turnover and resolution of inflammatory pathology in the CNS. This likely will have implications in neurocysticercosis and neuro-inflammatory diseases. SIGNIFICANCE STATEMENT Macrophages/microglia with M1-activation phenotype are thought to promote CNS pathology, whereas M2-anti-inflammatory phenotype promote CNS repair. However, the mechanisms regulating M2 cell protective functions in the CNS microenvironment are undefined. Quenum Zangbede et. al., report that helminth infection of the brain induces an increased expression of galectin-3 in M2 macrophages accumulated in the CNS. Using multiple experimental models in vivo and in vitro , they show that galectin-3 in M2 macrophages functions to clear neutrophils accumulated in the CNS. Importantly, galectin-3 in M2 macrophages plays a central role in the containment of neuropathology and disease severity. These results provide a direct mechanistic evidence of the protective function of M2- macrophages in the CNS. Copyright © 2018 the authors.

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

PubMed Central

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

2014-01-01

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

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

PubMed

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.

Phagocytosis imprints heterogeneity in tissue-resident macrophages

PubMed Central

A-Gonzalez, Noelia; Quintana, Juan A.; Mazariegos, Marina; González de la Aleja, Arturo; Nicolás-Ávila, José A.; Crainiciuc, Georgiana; Rothlin, Carla V.; Peinado, Héctor; Castrillo, Antonio

2017-01-01

Tissue-resident macrophages display varying phenotypic and functional properties that are largely specified by their local environment. One of these functions, phagocytosis, mediates the natural disposal of billions of cells, but its mechanisms and consequences within living tissues are poorly defined. Using a parabiosis-based strategy, we identified and isolated macrophages from multiple tissues as they phagocytosed blood-borne cellular material. Phagocytosis was circadianally regulated and mediated by distinct repertoires of receptors, opsonins, and transcription factors in macrophages from each tissue. Although the tissue of residence defined the core signature of macrophages, phagocytosis imprinted a distinct antiinflammatory profile. Phagocytic macrophages expressed CD206, displayed blunted expression of Il1b, and supported tissue homeostasis. Thus, phagocytosis is a source of macrophage heterogeneity that acts together with tissue-derived factors to preserve homeostasis. PMID:28432199

Evidence for the opposing roles of different gamma delta T cell subsets in macrophage homeostasis.

PubMed

Tramonti, Daniela; Andrew, Elizabeth M; Rhodes, Kate; Newton, Darren J; Carding, Simon R

2006-07-01

To ensure invading pathogens are eliminated with minimal damage to host tissues it is essential that macrophage activation be tightly regulated. Previously we demonstrated that a subset of gammadelta T cells (Vgamma1(+)) contributes to resolving pathogen-induced immune responses by killing activated macrophages. However, the exaggerated macrophage response seen in infected Vgamma1(+) T cell-deficient mice suggests that gammadelta T cells play a broader role in macrophage homeostasis and other subsets might promote macrophage activation. Using a macrophage:gammadelta T cell co-culture system we have shown that gammadelta T cells increase the activity of macrophages activated in vivo by Listeria monocytogenes infection. In a dose-dependent manner, gammadelta T cells up-regulated production of cytokines (TNF-alpha, IL-6, IL-10) and chemokines (MIP-1alpha, MIP-1beta) by Listeria-elicited macrophages. The ability to increase macrophage cytokine production was prominent among Vgamma4(+) gammadelta T cells. Reciprocally, Vgamma4(+) gammadelta T cells were activated by Listeria-elicited macrophages, resulting in production of the anti-inflammatory cytokine, IL-10. gammadelta T cell adoptive transfer experiments showed that Vgamma4(+) T cells protected TCRdelta(-/-) mice against Listeria-induced liver injury and necrosis. These findings identify distinct and non-overlapping roles for gammadelta T cell subsets in regulating macrophage function during pathogen-induced immune responses.

Elevation in Body Temperature to Fever Range Enhances and Prolongs Subsequent Responsiveness of Macrophages to Endotoxin Challenge

PubMed Central

Lee, Chen-Ting; Zhong, Lingwen; Mace, Thomas A.; Repasky, Elizabeth A.

2012-01-01

Macrophages are often considered the sentries in innate immunity, sounding early immunological alarms, a function which speeds the response to infection. Compared to the large volume of studies on regulation of macrophage function by pathogens or cytokines, relatively little attention has been devoted to the role of physical parameters such as temperature. Given that temperature is elevated during fever, a long-recognized cardinal feature of inflammation, it is possible that macrophage function is responsive to thermal signals. To explore this idea, we used LPS to model an aseptic endotoxin-induced inflammatory response in BALB/c mice and found that raising mouse body temperature by mild external heat treatment significantly enhances subsequent LPS-induced release of TNF-α into the peritoneal fluid. It also reprograms macrophages, resulting in sustained subsequent responsiveness to LPS, i.e., this treatment reduces “endotoxin tolerance” in vitro and in vivo. At the molecular level, elevating body temperature of mice results in a increase in LPS-induced downstream signaling including enhanced phosphorylation of IKK and IκB, NF-κB nuclear translocation and binding to the TNF-α promoter in macrophages upon secondary stimulation. Mild heat treatment also induces expression of HSP70 and use of HSP70 inhibitors (KNK437 or Pifithrin-µ) largely abrogates the ability of the thermal treatment to enhance TNF-α, suggesting that the induction of HSP70 is important for mediation of thermal effects on macrophage function. Collectively, these results support the idea that there has been integration between the evolution of body temperature regulation and macrophage function that could help to explain the known survival benefits of fever in organisms following infection. PMID:22253887

CD68 on rat macrophages binds tightly to S100A8 and S100A9 and helps to regulate the cells' immune functions.

PubMed

Okada, Kohki; Arai, Satoshi; Itoh, Hiroshi; Adachi, Souichi; Hayashida, Masahiko; Nakase, Hiroshi; Ikemoto, Masaki

2016-11-01

S100A8 and S100A9 (S100 proteins) are regulators of immune cells of myeloid origin. Whereas S100 proteins are found at high concentrations in such cells, their immunologic roles remain unclear. We focused on cluster of differentiation 68 (CD68). The aim of this study is to investigate whether CD68 binds to extracellular S100A8 and/or S100A9 and subsequently participates in the regulation of the cells' immune functions. ELISA and affinity chromatography showed that both recombinant rat S100A8 (r-S100A8) and r-S100A9 bound to r-CD68, but not to r-CD14. Flow cytometry clearly showed evidences supporting above the 2 results. As analyzed by flow cytometry, a less amount of r-S100A8 or r-S100A9 bound to the macrophages treated with some deglycosylation enzymes. In an in vitro assay, the expression levels of S100A8 and S100A9 were significantly suppressed after the macrophages had been treated with an anti-CD68 antibody (ED1). As stimulated macrophages with r-S100A9, the expression of IL-1β mRNA in macrophages, which were treated with anti-TLR4 or -RAGE antibodies, was significantly suppressed. r-S100A8 up-regulated IL-6 and IL-10 mRNAs, while r-S100A9 did TNF-α and IL-6 mRNAs, although these regulations were not statistically significant. Small interfering CD68 also significantly suppressed activation of macrophages through an autocrine pathway by r-S100A8 or r-S100A9. In macrophages stimulated with LPS, fluorescent immunologic staining showed that most CD68 colocalized with S100A8 or S100A9 and that the levels of all 3 molecules were markedly increased. In conclusion, CD68 on macrophages binds to S100A8 and S100A9 and thereby, plays a role in the regulation of the cells' immune functions. © Society for Leukocyte Biology.

Cysteamine re-establishes the clearance of Pseudomonas aeruginosa by macrophages bearing the cystic fibrosis-relevant F508del-CFTR mutation.

PubMed

Ferrari, Eleonora; Monzani, Romina; Villella, Valeria R; Esposito, Speranza; Saluzzo, Francesca; Rossin, Federica; D'Eletto, Manuela; Tosco, Antonella; De Gregorio, Fabiola; Izzo, Valentina; Maiuri, Maria C; Kroemer, Guido; Raia, Valeria; Maiuri, Luigi

2017-01-12

Cystic fibrosis (CF), the most common lethal monogenic disease in Caucasians, is characterized by recurrent bacterial infections and colonization, mainly by Pseudomonas aeruginosa, resulting in unresolved airway inflammation. CF is caused by mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein, which functions as a chloride channel in epithelial cells, macrophages, and other cell types. Impaired bacterial handling by macrophages is a feature of CF airways, although it is still debated how defective CFTR impairs bacterial killing. Recent evidence indicates that a defective autophagy in CF macrophages leads to alterations of bacterial clearance upon infection. Here we use bone marrow-derived macrophages from transgenic mice to provide the genetic proof that defective CFTR compromises both uptake and clearance of internalized Pseudomonas aeruginosa. We demonstrate that the proteostasis regulator cysteamine, which rescues the function of the most common F508del-CFTR mutant and hence reduces lung inflammation in CF patients, can also repair the defects of CF macrophages, thus restoring both bacterial internalization and clearance through a process that involves upregulation of the pro-autophagic protein Beclin 1 and re-establishment of the autophagic pathway. Altogether these results indicate that cysteamine restores the function of several distinct cell types, including that of macrophages, which might contribute to its beneficial effects on CF.

Cysteamine re-establishes the clearance of Pseudomonas aeruginosa by macrophages bearing the cystic fibrosis-relevant F508del-CFTR mutation

PubMed Central

Ferrari, Eleonora; Monzani, Romina; Villella, Valeria R; Esposito, Speranza; Saluzzo, Francesca; Rossin, Federica; D'Eletto, Manuela; Tosco, Antonella; De Gregorio, Fabiola; Izzo, Valentina; Maiuri, Maria C; Kroemer, Guido; Raia, Valeria; Maiuri, Luigi

2017-01-01

Cystic fibrosis (CF), the most common lethal monogenic disease in Caucasians, is characterized by recurrent bacterial infections and colonization, mainly by Pseudomonas aeruginosa, resulting in unresolved airway inflammation. CF is caused by mutations in the gene coding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein, which functions as a chloride channel in epithelial cells, macrophages, and other cell types. Impaired bacterial handling by macrophages is a feature of CF airways, although it is still debated how defective CFTR impairs bacterial killing. Recent evidence indicates that a defective autophagy in CF macrophages leads to alterations of bacterial clearance upon infection. Here we use bone marrow-derived macrophages from transgenic mice to provide the genetic proof that defective CFTR compromises both uptake and clearance of internalized Pseudomonas aeruginosa. We demonstrate that the proteostasis regulator cysteamine, which rescues the function of the most common F508del-CFTR mutant and hence reduces lung inflammation in CF patients, can also repair the defects of CF macrophages, thus restoring both bacterial internalization and clearance through a process that involves upregulation of the pro-autophagic protein Beclin 1 and re-establishment of the autophagic pathway. Altogether these results indicate that cysteamine restores the function of several distinct cell types, including that of macrophages, which might contribute to its beneficial effects on CF. PMID:28079883

Macrophages in tissue repair, regeneration, and fibrosis

PubMed Central

Wynn, Thomas A.; Vannella, Kevin M.

2016-01-01

Inflammatory monocytes and resident tissue macrophages are key regulators of tissue repair, regeneration, and fibrosis. Following tissue injury, monocytes and macrophages undergo marked phenotypic and functional changes to play critical roles during the initiation, maintenance, and resolution phases of tissue repair. Disturbances in macrophage function can lead to aberrant repair, with uncontrolled inflammatory mediator and growth factor production, deficient generation of anti-inflammatory macrophages, or failed communication between macrophages and epithelial cells, endothelial cells, fibroblasts, and stem or tissue progenitor cells all contributing to a state of persistent injury, which may lead to the development of pathological fibrosis. In this review, we discuss the mechanisms that instruct macrophages to adopt pro-inflammatory, pro-wound healing, pro-fibrotic, anti-inflammatory, anti-fibrotic, pro-resolving, and tissue regenerating phenotypes following injury, and highlight how some of these mechanisms and macrophage activation states could be exploited therapeutically. PMID:26982353

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

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

Srivastava, Ritesh K.; Li, Changzhao; Chaudhary, Sandeep C.

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,more » 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.« less

The macrophage marker translocator protein (TSPO) is down-regulated on pro-inflammatory ‘M1’ human macrophages

PubMed Central

Mandhair, Harpreet; Smyth, Erica; Dakin, Stephanie Georgina; Kiriakidis, Serafim; Wells, Lisa; Owen, David; Sabokbar, Afsie; Taylor, Peter

2017-01-01

The translocator protein (TSPO) is a mitochondrial membrane protein, of as yet uncertain function. Its purported high expression on activated macrophages, has lent utility to TSPO targeted molecular imaging in the form of positron emission tomography (PET), as a means to detect and quantify inflammation in vivo. However, existing literature regarding TSPO expression on human activated macrophages is lacking, mostly deriving from brain tissue studies, including studies of brain malignancy, and inflammatory diseases such as multiple sclerosis. Here, we utilized three human sources of monocyte derived macrophages (MDM), from THP-1 monocytes, healthy peripheral blood monocytes and synovial fluid monocytes from patients with rheumatoid arthritis, to undertake a detailed investigation of TSPO expression in activated macrophages. In this work, we demonstrate a consistent down-regulation of TSPO mRNA and protein in macrophages activated to a pro-inflammatory, or ‘M1’ phenotype. Conversely, stimulation of macrophages to an M2 phenotype with IL-4, dexamethasone or TGF-β1 did not alter TSPO expression, regardless of MDM source. The reasons for this are uncertain, but our study findings add some supporting evidence for recent investigations concluding that TSPO may be involved in negative regulation of inflammatory responses in macrophages. PMID:28968465

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

PubMed

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.

Lipid homeostasis and inflammatory activation are disturbed in classically activated macrophages with peroxisomal β-oxidation deficiency.

PubMed

Geric, Ivana; Tyurina, Yulia Y; Krysko, Olga; Krysko, Dmitri V; De Schryver, Evelyn; Kagan, Valerian E; Van Veldhoven, Paul P; Baes, Myriam; Verheijden, Simon

2018-03-01

Macrophage activation is characterized by pronounced metabolic adaptation. Classically activated macrophages show decreased rates of mitochondrial fatty acid oxidation and oxidative phosphorylation and acquire a glycolytic state together with their pro-inflammatory phenotype. In contrast, alternatively activated macrophages require oxidative phosphorylation and mitochondrial fatty acid oxidation for their anti-inflammatory function. Although it is evident that mitochondrial metabolism is regulated during macrophage polarization and essential for macrophage function, little is known on the regulation and role of peroxisomal β-oxidation during macrophage activation. In this study, we show that peroxisomal β-oxidation is strongly decreased in classically activated bone-marrow-derived macrophages (BMDM) and mildly induced in alternatively activated BMDM. To examine the role of peroxisomal β-oxidation in macrophages, we used Mfp2 -/- BMDM lacking the key enzyme of this pathway. Impairment of peroxisomal β-oxidation in Mfp2 -/- BMDM did not cause lipid accumulation but rather an altered distribution of lipid species with very-long-chain fatty acids accumulating in the triglyceride and phospholipid fraction. These lipid alterations in Mfp2 -/- macrophages led to decreased inflammatory activation of Mfp2 -/- BMDM and peritoneal macrophages evidenced by impaired production of several inflammatory cytokines and chemokines, but did not affect anti-inflammatory polarization. The disturbed inflammatory responses of Mfp2 -/- macrophages did not affect immune cell infiltration, as mice with selective elimination of MFP2 from myeloid cells showed normal monocyte and neutrophil influx upon challenge with zymosan. Together, these data demonstrate that peroxisomal β-oxidation is involved in fine-tuning the phenotype of macrophages, probably by influencing the dynamic lipid profile during macrophage polarization. © 2017 John Wiley & Sons Ltd.

Differentiation and Glucocorticoid Regulated Apopto-Phagocytic Gene Expression Patterns in Human Macrophages. Role of Mertk in Enhanced Phagocytosis

PubMed Central

Zahuczky, Gábor; Kristóf, Endre; Majai, Gyöngyike; Fésüs, László

2011-01-01

The daily clearance of physiologically dying cells is performed safely mainly by cells in the mononuclear phagocyte system. They can recognize and engulf dying cells utilizing several cooperative mechanisms. In our study we show that the expression of a broad range of apopto-phagocytic genes is strongly up-regulated during differentiation of human monocytes to macrophages with different donor variability. The glucocorticoid dexamethasone has a profound effect on this process by selectively up-regulating six genes and down-regulating several others. The key role of the up-regulated mer tyrosine kinase (Mertk) in dexamethasone induced enhancement of phagocytosis could be demonstrated in human monocyte derived macrophages by gene silencing as well as blocking antibodies, and also in a monocyte-macrophage like cell line. However, the additional role of other glucocorticoid induced elements must be also considered since the presence of autologous serum during phagocytosis could almost completely compensate for the blocked function of Mertk. PMID:21731712

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

PubMed Central

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

Heparanase regulates the M1 polarization of renal macrophages and their crosstalk with renal epithelial tubular cells after ischemia/reperfusion injury.

PubMed

Masola, Valentina; Zaza, Gianluigi; Bellin, Gloria; Dall'Olmo, Luigi; Granata, Simona; Vischini, Gisella; Secchi, Maria Francesca; Lupo, Antonio; Gambaro, Giovanni; Onisto, Maurizio

2018-02-01

Heparanase (HPSE) is part of the biologic network triggered by ischemia/reperfusion (I/R) injury, a complication of renal transplantation and acute kidney injury. During this period, the kidney or graft undergoes a process of macrophages recruitment and activation. HPSE may therefore control these biologic effects. We measured the ability of HPSE and its inhibitor, SST0001, to regulate macrophage polarization and the crosstalk between macrophages and HK-2 renal tubular cells during in vitro hypoxia/reoxygenation (H/R). Furthermore, we evaluated in vivo renal inflammation, macrophage polarization, and histologic changes in mice subjected to monolateral I/R and treated with SST0001 for 2 or 7 d. The in vitro experiments showed that HPSE sustained M1 macrophage polarization and modulated apoptosis, the release of damage associated molecular patterns in post-H/R tubular cells, the synthesis of proinflammatory cytokines, and the up-regulation of TLRs on both epithelial cells and macrophages. HPSE also regulated M1 polarization induced by H/R-injured tubular cells and the partial epithelial-mesenchymal transition of these epithelial cells by M1 macrophages. All these effects were prevented by inhibiting HPSE. Furthermore, the inhibition of HPSE in vivo reduced inflammation and M1 polarization in mice undergoing I/R injury, partially restored renal function and normal histology, and reduced apoptosis. These results show for the first time that HPSE regulates macrophage polarization as well as renal damage and repair after I/R. HPSE inhibitors could therefore provide a new pharmacologic approach to minimize acute kidney injury and to prevent the chronic profibrotic damages induced by I/R.-Masola, V., Zaza, G., Bellin, G., Dall'Olmo, L., Granata, S., Vischini, G., Secchi, M. F., Lupo, A., Gambaro, G., Onisto, M. Heparanase regulates the M1 polarization of renal macrophages and their crosstalk with renal epithelial tubular cells after ischemia/reperfusion injury.

Differential regulation of macrophage inflammatory activation by fibrin and fibrinogen.

PubMed

Hsieh, Jessica Y; Smith, Tim D; Meli, Vijaykumar S; Tran, Thi N; Botvinick, Elliot L; Liu, Wendy F

2017-01-01

Fibrin is a major component of the provisional extracellular matrix formed during tissue repair following injury, and enables cell infiltration and anchoring at the wound site. Macrophages are dynamic regulators of this process, advancing and resolving inflammation in response to cues in their microenvironment. Although much is known about how soluble factors such as cytokines and chemokines regulate macrophage polarization, less is understood about how insoluble and adhesive cues, specifically the blood coagulation matrix fibrin, influence macrophage behavior. In this study, we observed that fibrin and its precursor fibrinogen elicit distinct macrophage functions. Culturing macrophages on fibrin gels fabricated by combining fibrinogen with thrombin stimulated secretion of the anti-inflammatory cytokine, interleukin-10 (IL-10). In contrast, exposure of macrophages to soluble fibrinogen stimulated high levels of inflammatory cytokine tumor necrosis factor alpha (TNF-α). Macrophages maintained their anti-inflammatory behavior when cultured on fibrin gels in the presence of soluble fibrinogen. In addition, adhesion to fibrin matrices inhibited TNF-α production in response to stimulation with LPS and IFN-γ, cytokines known to promote inflammatory macrophage polarization. Our data demonstrate that fibrin exerts a protective effect on macrophages, preventing inflammatory activation by stimuli including fibrinogen, LPS, and IFN-γ. Together, our study suggests that the presentation of fibrin(ogen) may be a key switch in regulating macrophage phenotype behavior, and this feature may provide a valuable immunomodulatory strategy for tissue healing and regeneration. Fibrin is a fibrous protein resulting from blood clotting and provides a provisional matrix into which cells migrate and to which they adhere during wound healing. Macrophages play an important role in this process, and are needed for both advancing and resolving inflammation. We demonstrate that culture of macrophages on fibrin matrices exerts an anti-inflammatory effect, whereas the soluble precursor fibrinogen stimulates inflammatory activation. Moreover, culture on fibrin completely abrogates inflammatory signaling caused by fibrinogen or known inflammatory stimuli including LPS and IFN-γ. Together, these studies show that the presentation of fibrin(ogen) is important for regulating a switch between macrophage pro- and anti-inflammatory behavior. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Leishmania donovani chaperonin 10 regulates parasite internalization and intracellular survival in human macrophages.

PubMed

Colineau, Lucie; Clos, Joachim; Moon, Kyung-Mee; Foster, Leonard J; Reiner, Neil E

2017-06-01

Protozoa of the genus Leishmania infect macrophages in their mammalian hosts causing a spectrum of diseases known as the leishmaniases. The search for leishmania effectors that support macrophage infection is a focus of significant interest. One such candidate is leishmania chaperonin 10 (CPN10) which is secreted in exosomes and may have immunosuppressive properties. Here, we report for the first time that leishmania CPN10 localizes to the cytosol of infected macrophages. Next, we generated two genetically modified strains of Leishmania donovani (Ld): one strain overexpressing CPN10 (CPN10+++) and the second, a CPN10 single allele knockdown (CPN10+/-), as the null mutant was lethal. When compared with the wild-type (WT) parental strain, CPN10+/- Ld showed higher infection rates and parasite loads in human macrophages after 24 h of infection. Conversely, CPN10+++ Ld was associated with lower initial infection rates. This unexpected apparent gain-of-function for the knockdown could have been explained either by enhanced parasite internalization or by enhanced intracellular survival. Paradoxically, we found that CPN10+/- leishmania were more readily internalized than WT Ld, but also displayed significantly impaired intracellular survival. This suggests that leishmania CPN10 negatively regulates the rate of parasite uptake by macrophages while being required for intracellular survival. Finally, quantitative proteomics identified an array of leishmania proteins whose expression was positively regulated by CPN10. In contrast, many macrophage proteins involved in innate immunity were negatively regulated by CPN10. Taken together, these findings identify leishmania CPN10 as a novel effector with broad based effects on macrophage cell regulation and parasite survival.

Macrophage Phenotypes Regulate Scar Formation and Chronic Wound Healing.

PubMed

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

2017-07-17

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

Transcription factor NFAT5 promotes macrophage survival in rheumatoid arthritis

PubMed Central

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

2017-01-01

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

Macrophages: An Inflammatory Link between Angiogenesis and Lymphangiogenesis

PubMed Central

Corliss, Bruce A.; Azimi, Mohammad S.; Munson, Jenny; Peirce, Shayn M.; Murfee, Walter Lee

2015-01-01

Angiogenesis and lymphangiogenesis often occur in response to tissue injury or in the presence of pathology (e.g. cancer), and it is these types of environments in which macrophages are activated and increased in number. Moreover, the blood vascular microcirculation and the lymphatic circulation serve as the conduits for entry and exit for monocyte-derived macrophages in nearly every tissue and organ. Macrophages both affect and are affected by the vessels through which they travel. Therefore, it is not surprising that examination of macrophage behaviors in both angiogenesis and lymphangiogenesis has yielded interesting observations that suggest macrophages may be key regulators of these complex growth and remodeling processes. In this review, we will take a closer look at macrophages through the lens of angiogenesis and lymphangiogenesis, examining how their dynamic behaviors may regulate vessel sprouting and function. We present macrophages as a cellular link that spatially and temporally connects angiogenesis with lymphangiogenesis, in both physiological growth and in pathological adaptations, such as tumorigenesis. As such, attempts to therapeutically target macrophages in order to affect these processes may be particularly effective, and studying macrophages in both settings will accelerate the field’s understanding of this important cell type in health and disease. PMID:26614117

Regulation of CRIg Expression and Phagocytosis in Human Macrophages by Arachidonate, Dexamethasone, and Cytokines

PubMed Central

Gorgani, Nick N.; Thathaisong, Umaporn; Mukaro, Violet R.S.; Poungpair, Ornnuthchar; Tirimacco, Amanda; Hii, Charles S.T.; Ferrante, Antonio

2011-01-01

Although the importance of the macrophage complement receptor immunoglobulin (CRIg) in the phagocytosis of complement opsonized bacteria and in inflammation has been established, the regulation of CRIg expression remains undefined. Because cellular activation during inflammation leads to the release of arachidonate, a stimulator of leukocyte function, we sought to determine whether arachidonate regulates CRIg expression. Adding arachidonate to maturing human macrophages and to prematured CRIg+ macrophages caused a significant decrease in the expression of cell-surface CRIg and CRIg mRNA. This effect was independent of the metabolism of arachidonate via the cyclooxygenase and lipoxygenase pathways, because it was not inhibited by the nonsteroidal anti-inflammatory drugs indomethacin and nordihydroguaiaretic acid. Studies with specific pharmacological inhibitors of arachidonate-mediated signaling pathways showed that protein kinase C was involved. Administration of dexamethasone to macrophages caused an increase in CRIg expression. Studies with proinflammatory and immunosuppressive cytokines showed that IL-10 increased, but interferon-γ, IL-4, and transforming growth factor-β1 decreased CRIg expression on macrophages. This down- and up-regulation of CRIg expression was reflected in a decrease and increase, respectively, in the phagocytosis of complement opsonized Candida albicans. These data suggest that a unique inflammatory mediator network regulates CRIg expression and point to a mechanism by which arachidonate and dexamethasone have reciprocal effects on inflammation. PMID:21741936

Protein Thiol Redox Signaling in Monocytes and Macrophages.

PubMed

Short, John D; Downs, Kevin; Tavakoli, Sina; Asmis, Reto

2016-11-20

Monocyte and macrophage dysfunction plays a critical role in a wide range of inflammatory disease processes, including obesity, impaired wound healing diabetic complications, and atherosclerosis. Emerging evidence suggests that the earliest events in monocyte or macrophage dysregulation include elevated reactive oxygen species production, thiol modifications, and disruption of redox-sensitive signaling pathways. This review focuses on the current state of research in thiol redox signaling in monocytes and macrophages, including (i) the molecular mechanisms by which reversible protein-S-glutathionylation occurs, (ii) the identification of bona fide S-glutathionylated proteins that occur under physiological conditions, and (iii) how disruptions of thiol redox signaling affect monocyte and macrophage functions and contribute to atherosclerosis. Recent Advances: Recent advances in redox biochemistry and biology as well as redox proteomic techniques have led to the identification of many new thiol redox-regulated proteins and pathways. In addition, major advances have been made in expanding the list of S-glutathionylated proteins and assessing the role that protein-S-glutathionylation and S-glutathionylation-regulating enzymes play in monocyte and macrophage functions, including monocyte transmigration, macrophage polarization, foam cell formation, and macrophage cell death. Protein-S-glutathionylation/deglutathionylation in monocytes and macrophages has emerged as a new and important signaling paradigm, which provides a molecular basis for the well-established relationship between metabolic disorders, oxidative stress, and cardiovascular diseases. The identification of specific S-glutathionylated proteins as well as the mechanisms that control this post-translational protein modification in monocytes and macrophages will facilitate the development of new preventive and therapeutic strategies to combat atherosclerosis and other metabolic diseases. Antioxid. Redox Signal. 25, 816-835.

Leishmania infection inhibits macrophage motility by altering F-actin dynamics and the expression of adhesion complex proteins

PubMed Central

de Menezes, Juliana Perrone Bezerra; Koushik, Amrita; Das, Satarupa; Guven, Can; Siegel, Ariel; Laranjeira-Silva, Maria Fernanda; Losert, Wolfgang; Andrews, Norma W.

2016-01-01

Leishmania is an intracellular protozoan parasite that causes a broad spectrum of clinical manifestations, ranging from self-healing skin lesions to fatal visceralizing disease. As the host cells of choice for all species of Leishmania, macrophages are critical for the establishment of infections. How macrophages contribute to parasite homing to specific tissues and how parasites modulate macrophage function is still poorly understood. In this study we show that L. amazonensis infection inhibits macrophage roaming motility. The reduction in macrophage speed is not dependent on particle load or on factors released by infected macrophages. L. amazonensis-infected macrophages also show reduced directional migration in response to the chemokine MCP-1. We found that infected macrophages have lower levels of total paxillin, phosphorylated paxillin and phosphorylated FAK when compared to non-infected macrophages, indicating abnormalities in the formation of signaling adhesion complexes that regulate motility. Analysis of the dynamics of actin polymerization at peripheral sites also revealed a markedly enhanced F-actin turnover frequency in L. amazonensis-infected macrophages. Thus, Leishmania infection inhibits macrophage motility by altering actin dynamics and impairing the expression of proteins that function in plasma membrane-extracellular matrix interactions. PMID:27641840

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

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

Tian, Ping-Ge; Jiang, Zhi-Xin; Li, Jian-Hua

Macrophage autophagy plays an important role in the development of atherosclerosis, but the precise mechanism mediating this process is unclear. The potential role of the X-box binding protein 1 (XBP1), a crucial transduction factor that is involved in endoplasmic reticulum stress and the unfolded protein response, in bone marrow-derived macrophage autophagy is unknown. This study mainly explores the roles of XBP1 mRNA splicing in bone marrow-derived macrophage autophagy. The present study shows that the transient overexpression of spliced XBP1 via adenovirus-mediated gene transfer induces autophagy and promotes proliferation in bone marrow-derived macrophages via the down-regulation of Beclin-1, but that themore » sustained overexpression of spliced XBP1 leads to apoptosis. When XBP1 is down-regulated in bone marrow-derived macrophages using siRNA, rapamycin-induced autophagosome formation is ablated. Furthermore, we have detected the overexpression of XBP1 in areas of atherosclerotic plaques in the arteries of ApoE−/− mice. These results demonstrate that XBP1 mRNA splicing plays an important role in maintaining the function of bone marrow-derived macrophages and provide new insight into the study and treatment of atherosclerosis. - Highlights: • XBP1 was up-regulated in atherosclerotic plaques of ApoE−/− mice. • Transient spliced XBP1 overexpression induced macrophages autophagy via Beclin-1. • Sustained spliced XBP1 overexpression triggered macrophages apoptosis. • Spliced XBP1 plays a key role in maintaining the macrophages survival.« less

Arginine methylation regulates c-Myc-dependent transcription by altering promoter recruitment of the acetyltransferase p300.

PubMed

Tikhanovich, Irina; Zhao, Jie; Bridges, Brian; Kumer, Sean; Roberts, Ben; Weinman, Steven A

2017-08-11

Protein arginine methyltransferase 1 (PRMT1) is an essential enzyme controlling about 85% of the total cellular arginine methylation in proteins. We have shown previously that PRMT1 is an important regulator of innate immune responses and that it is required for M2 macrophage differentiation. c-Myc is a transcription factor that is critical in regulating cell proliferation and also regulates the M2 transcriptional program in macrophages. Here, we sought to determine whether c-Myc in myeloid cells is regulated by PRMT1-dependent arginine methylation. We found that PRMT1 activity was necessary for c-Myc binding to the acetyltransferase p300. PRMT1 inhibition decreased p300 recruitment to c-Myc target promoters and increased histone deacetylase 1 (HDAC1) recruitment, thereby decreasing transcription at these sites. Moreover, PRMT1 inhibition blocked c-Myc-mediated induction of several of its target genes, including peroxisome proliferator-activated receptor γ ( PPARG ) and mannose receptor C-type 1 ( MRC1 ), suggesting that PRMT1 is necessary for c-Myc function in M2 macrophage differentiation. Of note, in primary human blood monocytes, p300-c-Myc binding was strongly correlated with PRMT1 expression, and in liver sections, PRMT1, c-Myc, and M2 macrophage levels were strongly correlated with each other. Both PRMT1 levels and M2 macrophage numbers were significantly lower in livers from individuals with a history of spontaneous bacterial peritonitis, known to have defective cellular immunity. In conclusion, our findings demonstrate that PRMT1 is an important regulator of c-Myc function in myeloid cells. PRMT1 loss in individuals with cirrhosis may contribute to their immune defects. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

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

PubMed

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.

Human macrophage ATP7A is localized in the trans-Golgi apparatus, controls intracellular copper levels, and mediates macrophage responses to dermal wounds.

PubMed

Kim, Ha Won; Chan, Qilin; Afton, Scott E; Caruso, Joseph A; Lai, Barry; Weintraub, Neal L; Qin, Zhenyu

2012-02-01

The copper transporter ATP7A has attracted significant attention since the discovery of its gene mutation leading to human Menkes disease. We previously reported that ATP7A is highly expressed in the human vasculature and identified a novel vascular function of ATP7A in modulation of the expression and activity of extracellular superoxide dismutase. We recently identified that ATP7A expression in THP-1 cells (a monocyte/macrophage model cell line) plays a role in the oxidation of low density lipoproteins, indicating that it is necessary to further investigate its expression and function in monocytes/macrophages. In the current study, we demonstrated the protein and mRNA expression of ATP7A in human peripheral blood mononuclear cell (PBMC)-derived macrophages and alveolar macrophages. ATP7A was strongly co-localized with the trans-Golgi apparatus in PBMC-derived macrophages. Intracellular copper, detected by synchrotron X-ray fluorescence microscopy, was found to be distributed to the nucleus and cytoplasm in human THP-1 cells. To confirm the role of endogenous ATP7A in macrophage copper homeostasis, we performed inductively coupled plasma mass spectrometry in murine peritoneal macrophages, which showed markedly increased intracellular copper levels in macrophages isolated from ATP7A-deficient mice versus control mice. Moreover, the role of ATP7A in regulating macrophage responses to dermal wounds was studied by introduction of control and ATP7A-downregulated THP-1 cells into dermal wounds of nude mice. Infiltration of THP-1 cells into the wounded area (detected by expression of human macrophage markers MAC2 and CD68) was reduced in response to downregulation of ATP7A, hinting decreased macrophage accumulation subsequent to dermal wounds. In summary, alongside our previous studies, these findings indicate that human macrophage ATP7A is localized in the trans-Golgi apparatus, regulates intracellular copper levels, and mediates macrophage responses to a dermal wound.

The regulation of TNFα production after heat and endotoxin stimulation is dependent on Annexin-A1 and HSP70.

PubMed

Nair, Sunitha; Arora, Suruchi; Lim, Jyue Yuan; Lee, Lay Hoon; Lim, Lina H K

2015-07-01

Febrile temperatures can induce stress responses which protect cells from damage and can reduce inflammation during infections and sepsis. However, the mechanisms behind the protective functions of heat in response to the bacterial endotoxin LPS are unclear. We have recently shown that Annexin-1 (ANXA1)-deficient macrophages exhibited higher TNFα levels after LPS stimulation. Moreover, we have previously reported that ANXA1 can function as a stress protein. Therefore in this study, we determined if ANXA1 is involved in the protective effects of heat on cytokine levels in macrophages after heat and LPS. Exposure of macrophages to 42 °C for 1 h prior to LPS results in an inhibition of TNFα production, which was not evident in ANXA1(-/-) macrophages. We show that this regulation involves primarily MYD88-independent pathways. ANXA1 regulates TNFα mRNA stability after heat and LPS, and this is dependent on endogenous ANXA1 expression and not exogenously secreted factors. Further mechanistic studies revealed the possible involvement of the heat shock protein HSP70 and JNK in the heat and inflammatory stress response regulated by ANXA1. This study shows that ANXA1, an immunomodulatory protein, is critical in the heat stress response induced after heat and endotoxin stimulation.

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

PubMed

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.

How does temperature affect the function of tissue macrophages?

NASA Astrophysics Data System (ADS)

Lee, Chen-Ting; Repasky, Elizabeth A.

2011-03-01

Macrophages create a major danger signal following injury or infection and upon activation release pro-inflammatory cytokines, which in turn help to generate febrile conditions. Thus, like other cells of the body, tissue macrophages are often exposed to naturally occurring elevations in tissue temperature during inflammation and fever. However, whether macrophages sense and respond to temperature changes in a specific manner which modulates their function is still not clear. In this brief review, we highlight recent studies which have analyzed the effects of temperatures on macrophage function, and summarize the possible underlying molecular mechanisms which have been identified. Mild, physiological range hyperthermia has been shown to have both pro- and anti-inflammatory roles in regulating macrophage inflammatory cytokine production and at the meeting presentation, we will show new data demonstrating that hyperthermia can indeed exert both positive and negative signals to macrophages. While some thermal effects are correlated with the induction of heat shock factors/heat shock proteins, overall it is not clear how mild hyperthermia can exert both pro- and anti-inflammatory functions. We also summarize data which shows that hyperthermia can affect other macrophage effector functions, including the anti-tumor cytotoxicity. Overall, these studies may help us to better understand the immunological role of tissue temperature and may provide important information needed to maximize the application of heat in the treatment of various diseases including cancer.

Sonic hedgehog-Dependent Induction of MicroRNA 31 and MicroRNA 150 Regulates Mycobacterium bovis BCG-Driven Toll-Like Receptor 2 Signaling

PubMed Central

Ghorpade, Devram Sampat; Holla, Sahana; Kaveri, Srini V.; Bayry, Jagadeesh; Patil, Shripad A.

2013-01-01

Hedgehog (HH) signaling is a significant regulator of cell fate decisions during embryogenesis, development, and perpetuation of various disease conditions. Testing whether pathogen-specific HH signaling promotes unique innate recognition of intracellular bacteria, we demonstrate that among diverse Gram-positive or Gram-negative microbes, Mycobacterium bovis BCG, a vaccine strain, elicits a robust activation of Sonic HH (SHH) signaling in macrophages. Interestingly, sustained tumor necrosis factor alpha (TNF-α) secretion by macrophages was essential for robust SHH activation, as TNF-α−/− macrophages exhibited compromised ability to activate SHH signaling. Neutralization of TNF-α or blockade of TNF-α receptor signaling significantly reduced the infection-induced SHH signaling activation both in vitro and in vivo. Intriguingly, activated SHH signaling downregulated M. bovis BCG-mediated Toll-like receptor 2 (TLR2) signaling events to regulate a battery of genes associated with divergent functions of M1/M2 macrophages. Genome-wide expression profiling as well as conventional gain-of-function or loss-of-function analysis showed that SHH signaling-responsive microRNA 31 (miR-31) and miR-150 target MyD88, an adaptor protein of TLR2 signaling, thus leading to suppression of TLR2 responses. SHH signaling signatures could be detected in vivo in tuberculosis patients and M. bovis BCG-challenged mice. Collectively, these investigations identify SHH signaling to be what we believe is one of the significant regulators of host-pathogen interactions. PMID:23166298

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

PubMed Central

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

2016-01-01

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

Genome-Wide Survey and Expression Profiling of CCCH-Zinc Finger Family Reveals a Functional Module in Macrophage Activation

PubMed Central

Liang, Jian; Song, Wenjun; Tromp, Gail; Kolattukudy, Pappachan E.; Fu, Mingui

2008-01-01

Previously, we have identified a novel CCCH zinc finger protein family as negative regulators of macrophage activation. To gain an overall insight into the entire CCCH zinc finger gene family and to evaluate their potential role in macrophage activation, here we performed a genome-wide survey of CCCH zinc finger genes in mouse and human. Totally 58 CCCH zinc finger genes in mouse and 55 in human were identified and most of them have not been reported previously. Phylogenetic analysis revealed that the mouse CCCH family was divided into 6 groups. Meanwhile, we employed quantitative real-time PCR to profile their tissue expression patterns in adult mice. Clustering analysis showed that most of CCCH genes were broadly expressed in all of tissues examined with various levels. Interestingly, several CCCH genes Mbnl3, Zfp36l2, Zfp36, Zc3h12a, Zc3h12d, Zc3h7a and Leng9 were enriched in macrophage-related organs such as thymus, spleen, lung, intestine and adipose. Consistently, a comprehensive assessment of changes in expression of the 58 members of the mouse CCCH family during macrophage activation also revealed that these CCCH zinc finger genes were associated with the activation of bone marrow-derived macrophages by lipopolysaccharide. Taken together, this study not only identified a functional module of CCCH zinc finger genes in the regulation of macrophage activation but also provided the framework for future studies to dissect the function of this emerging gene family. PMID:18682727

β-Catenin–regulated myeloid cell adhesion and migration determine wound healing

PubMed Central

Amini-Nik, Saeid; Cambridge, Elizabeth; Yu, Winston; Guo, Anne; Whetstone, Heather; Nadesan, Puviindran; Poon, Raymond; Hinz, Boris; Alman, Benjamin A.

2014-01-01

A β-catenin/T cell factor–dependent transcriptional program is critical during cutaneous wound repair for the regulation of scar size; however, the relative contribution of β-catenin activity and function in specific cell types in the granulation tissue during the healing process is unknown. Here, cell lineage tracing revealed that cells in which β-catenin is transcriptionally active express a gene profile that is characteristic of the myeloid lineage. Mice harboring a macrophage-specific deletion of the gene encoding β-catenin exhibited insufficient skin wound healing due to macrophage-specific defects in migration, adhesion to fibroblasts, and ability to produce TGF-β1. In irradiated mice, only macrophages expressing β-catenin were able to rescue wound-healing deficiency. Evaluation of scar tissue collected from patients with hypertrophic and normal scars revealed a correlation between the number of macrophages within the wound, β-catenin levels, and cellularity. Our data indicate that β-catenin regulates myeloid cell motility and adhesion and that β-catenin–mediated macrophage motility contributes to the number of mesenchymal cells and ultimate scar size following cutaneous injury. PMID:24837430

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

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

Matsumura, Takayuki; Department of Life Science and Medical Bio-Science, Waseda University, Shinjuku-ku, Tokyo 162-8480; Oyama, Masaaki

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 analysismore » 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.« less

Cavin1; a Regulator of Lung Function and Macrophage Phenotype

PubMed Central

Govender, Praveen; Romero, Freddy; Shah, Dilip; Paez, Jesus; Ding, Shi-Ying; Liu, Libin; Gower, Adam; Baez, Elizabeth; Aly, Sherif Shawky; Pilch, Paul; Summer, Ross

2013-01-01

Caveolae are cell membrane invaginations that are highly abundant in adipose tissue, endothelial cells and the lung. The formation of caveolae is dependent on the expression of various structural proteins that serve as scaffolding for these membrane invaginations. Cavin1 is a newly identified structural protein whose deficiency in mice leads to loss of caveolae formation and to development of a lipodystrophic phenotype. In this study, we sought to investigate the functional role of Cavin1 in the lung. Cavin1 deficient mice possessed dramatically altered distal lung morphology and exhibited significant physiological alterations, notably, increased lung elastance. The changes in distal lung architecture were associated with hypercellularity and the accumulation of lung macrophages. The increases in lung macrophages occurred without changes to circulating numbers of mononuclear cells and without evidence for increased proliferation. However, the increases in lung macrophages were associated with higher levels of macrophage chemotactic factors CXCL2 and CCL2 in BAL fluid from Cavin1−/− mice suggesting a possible mechanism by which these cells accumulate. In addition, lung macrophages from Cavin1−/− mice were larger and displayed measurable differences in gene expression when compared to macrophages from wild-type mice. Interestingly, macrophages were also increased in adipose tissue but not in liver, kidney or skeletal muscle from Cavin1−/− mice, and similar tissue specificity for macrophage accumulation was observed in lungs and adipose tissue from Caveolin1−/− mice. In conclusion, this study demonstrates an important role for Cavin1 in lung homeostasis and suggests that caveolae structural proteins are necessary for regulating macrophage number and phenotype in the lung. PMID:23634221

Embryonic Stem Cells Promoting Macrophage Survival and Function are Crucial for Teratoma Development

PubMed Central

Chen, Tianxiang; Wang, Xi; Guo, Lei; Wu, Mingmei; Duan, Zhaoxia; Lv, Jing; Tai, Wenjiao; Renganathan, Hemamalini; Didier, Ruth; Li, Jinhua; Sun, Dongming; Chen, Xiaoming; He, Xijing; Fan, Jianqing; Young, Wise; Ren, Yi

2014-01-01

Stem cell therapies have had tremendous potential application for many diseases in recent years. However, the tumorigenic properties of stem cells restrict their potential clinical application; therefore, strategies for reducing the tumorigenic potential of stem cells must be established prior to transplantation. We have demonstrated that syngeneic transplantation of embryonic stem cells (ESCs) provokes an inflammatory response that involves the rapid recruitment of bone marrow-derived macrophages (BMDMs). ESCs are able to prevent mature macrophages from macrophage colony-stimulating factor (M-CSF) withdrawal-induced apoptosis, and thus prolong macrophage lifespan significantly by blocking various apoptotic pathways in an M-CSF-independent manner. ESCs express and secrete IL-34, which may be responsible for ESC-promoted macrophage survival. This anti-apoptotic effect of ESCs involves activation of extracellular signal-regulated kinase (ERK)1/2 and PI3K/Akt pathways and thus, inhibition of ERK1/2 and PI3K/AKT activation decreases ESC-induced macrophage survival. Functionally, ESC-treated macrophages also showed a higher level of phagocytic activity. ESCs further serve to polarize BMDMs into M2-like macrophages that exhibit most tumor-associated macrophage phenotypic and functional features. ESC-educated macrophages produce high levels of arginase-1, Tie-2, and TNF-α, which participate in angiogenesis and contribute to teratoma progression. Our study suggests that induction of M2-like macrophage activation is an important mechanism for teratoma development. Strategies targeting macrophages to inhibit teratoma development would increase the safety of ESC-based therapies, inasmuch as the depletion of macrophages completely inhibits ESC-induced angiogenesis and teratoma development. PMID:25071759

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

PubMed

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

2016-02-01

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

Leishmania infection inhibits macrophage motility by altering F-actin dynamics and the expression of adhesion complex proteins.

PubMed

de Menezes, Juliana Perrone Bezerra; Koushik, Amrita; Das, Satarupa; Guven, Can; Siegel, Ariel; Laranjeira-Silva, Maria Fernanda; Losert, Wolfgang; Andrews, Norma W

2017-03-01

Leishmania is an intracellular protozoan parasite that causes a broad spectrum of clinical manifestations, ranging from self-healing skin lesions to fatal visceralizing disease. As the host cells of choice for all species of Leishmania, macrophages are critical for the establishment of infections. How macrophages contribute to parasite homing to specific tissues and how parasites modulate macrophage function are still poorly understood. In this study, we show that Leishmania amazonensis infection inhibits macrophage roaming motility. The reduction in macrophage speed is not dependent on particle load or on factors released by infected macrophages. L. amazonensis-infected macrophages also show reduced directional migration in response to the chemokine MCP-1. We found that infected macrophages have lower levels of total paxillin, phosphorylated paxillin, and phosphorylated focal adhesion kinase when compared to noninfected macrophages, indicating abnormalities in the formation of signaling adhesion complexes that regulate motility. Analysis of the dynamics of actin polymerization at peripheral sites also revealed a markedly enhanced F-actin turnover frequency in L. amazonensis-infected macrophages. Thus, Leishmania infection inhibits macrophage motility by altering actin dynamics and impairing the expression of proteins that function in plasma membrane-extracellular matrix interactions. © 2016 John Wiley & Sons Ltd.

Differential Transcriptional Response in Macrophages Infected with Cell Wall Deficient versus Normal Mycobacterium Tuberculosis

PubMed Central

Fu, Yu-Rong; Gao, Kun-Shan; Ji, Rui; Yi, Zheng-Jun

2015-01-01

Host-pathogen interactions determine the outcome following infection by mycobacterium tuberculosis (Mtb). Under adverse circumstances, normal Mtb can form cell-wall deficient (CWD) variants within macrophages, which have been considered an adaptive strategy for facilitating bacterial survival inside macrophages. However, the molecular mechanism by which infection of macrophages with different phenotypic Mtb elicits distinct responses of macrophages is not fully understood. To explore the molecular events triggered upon Mtb infection of macrophages, differential transcriptional responses of RAW264.7 cells infected with two forms of Mtb, CWD-Mtb and normal Mtb, were studied by microarray analysis. Some of the differentially regulated genes were confirmed by RT-qPCR in both RAW264.7 cells and primary macrophages. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway was used to analyze functions of differentially expressed genes. Distinct gene expression patterns were observed between CWD-Mtb and normal Mtb group. Mapt was up-regulated, while NOS2 and IL-11 were down-regulated in CWD-Mtb infected RAW264.7 cells and primary macrophages compared with normal Mtb infected ones. Many deregulated genes were found to be related to macrophages activation, immune response, phagosome maturation, autophagy and lipid metabolism. KEGG analysis showed that the differentially expressed genes were mainly involved in MAPK signaling pathway, nitrogen metabolism, cytokine-cytokine receptor interaction and focal adhesion. Taken together, the present study showed that differential macrophage responses were induced by intracellular CWD-Mtb an normal Mtb infection, which suggested that interactions between macrophages and different phenotypic Mtb are very complex. The results provide evidence for further understanding of pathogenesis of CWD-Mtb and may help in improving strategies to eliminate intracellular CWD-Mtb. PMID:25552926

Downmodulation of the Inflammatory Response to Bacterial Infection by γδ T Cells Cytotoxic for Activated Macrophages

PubMed Central

Egan, Paul J.; Carding, Simon R.

2000-01-01

Although γδ T cells are involved in the regulation of inflammation after infection, their precise function is not known. Intraperitoneal infection of T cell receptor (TCR)-δ−/− mice with the intracellular bacterium Listeria monocytogenes resulted in the development of necrotic foci in the livers. In contrast, the peritoneal cavities of infected TCR-δ−/− mice contained an accumulation of low density activated macrophages and a reduced percentage of macrophages undergoing apoptosis. γδ T cell hybridomas derived from mice infected with Listeria were preferentially stimulated by low density macrophages from peritoneal exudates of infected mice. Furthermore, primary splenic γδ T cells isolated from Listeria-infected mice were cytotoxic for low density macrophages in vitro, and cytotoxicity was inhibited in the presence of antibodies to the γδ TCR. These results demonstrate a novel interaction between γδ T cells and activated macrophages in which γδ T cells are stimulated by terminally differentiated macrophages to acquire cytotoxic activity and which, in turn, induce macrophage cell death. This interaction suggests that γδ T cells regulate the inflammatory response to infection with intracellular pathogens by eliminating activated macrophages at the termination of the response. PMID:10859339

Infection-induced regulation of NK cells by macrophages and collagen at the lymph node subcapsular sinus

PubMed Central

Coombes, Janine L.; Han, Seong-Ji; van Rooijen, Nico; Raulet, David H.; Robey, Ellen A.

2012-01-01

Summary Infection leads to heightened activation of natural killer (NK) cells, a process that likely involves direct cell-to-cell contact, but how this occurs in vivo is poorly understood. We have used two-photon laser-scanning microscopy in conjunction with Toxoplasma gondii-mouse infection models to address this question. We found that NK cells accumulated in the subcapsular region of the lymph node following infection where they formed low motility contacts with collagen fibers and CD169+ macrophages. We provide evidence that interactions with collagen regulate NK cell migration, whereas CD169+ macrophages increase the activation state of NK cells. Interestingly, a subset of CD169+ macrophages that co-express the inflammatory monocyte marker Ly6C had the most potent ability to activate NK cells. Our data reveal pathways through which NK cell migration and function are regulated following infection, and identify an important accessory cell population for activation of NK cell responses in lymph nodes. PMID:22840403

Epigenomics of macrophages

PubMed Central

Gosselin, David; Glass, Christopher K

2014-01-01

Summary Macrophages play essential roles in tissue homeostasis, pathogen elimination, and tissue repair. A defining characteristic of these cells is their ability to efficiently adapt to a variety of abruptly changing and complex environments. This ability is intrinsically linked to a capacity to quickly alter their transcriptome, and this is tightly associated with the epigenomic organization of these cells and, in particular, their enhancer repertoire. Indeed, enhancers are genomic sites that serve as platforms for the integration of signaling pathways with the mechanisms that regulate mRNA transcription. Notably, transcription is pervasive at active enhancers and enhancer RNAs (eRNAs) are tightly coupled to regulated transcription of protein-coding genes. Furthermore, given that each cell type possesses a defining enhancer repertoire, studies on enhancers provide a powerful method to study how specialization of functions among the diverse macrophage subtypes may arise. Here, we review recent studies providing insights into the distinct mechanisms that contribute to the establishment of enhancers and their role in the regulation of transcription in macrophages. PMID:25319330

MicroRNA-9 regulates the expression of peroxisome proliferator-activated receptor δ in human monocytes during the inflammatory response

PubMed Central

THULIN, PETRA; WEI, TIANLING; WERNGREN, OLIVERA; CHEUNG, LOUISA; FISHER, RACHEL M.; GRANDÉR, DAN; CORCORAN, MARTIN; EHRENBORG, EWA

2013-01-01

PPARδ is involved in the inflammatory response and its expression is induced by cytokines, however, limited knowledge has been produced regarding its regulation. Since recent findings have shown that microRNAs, which are small non-coding RNAs that regulate gene expression, are involved in the immune response, we set out to investigate whether PPARδ can be regulated by microRNAs expressed in monocytes. Bioinformatic analysis identified a putative miR-9 target site within the 3′-UTR of PPARδ that was subsequently verified to be functional using reporter constructs. Primary human monocytes stimulated with LPS showed a downregulation of PPARδ and its target genes after 4 h while the expression of miR-9 was induced. Analysis of pro-inflammatory (M1) and anti-inflammatory (M2) macrophages showed that human PPARδ mRNA as well as miR-9 expression was higher in M1 compared to M2 macrophages. Furthermore, treatment with the PPARδ agonist, GW501516, induced the expression of PPARδ target genes in the pro-inflammatory M1 macrophages while no change was observed in the anti-inflammatory M2 macrophages. Taken together, these data suggest that PPARδ is regulated by miR-9 in monocytes and that activation of PPARδ may be of importance in M1 pro-inflammatory but not in M2 anti-inflammatory macrophages in humans. PMID:23525285

Reprogramming mitochondrial metabolism in macrophages as an anti-inflammatory signal.

PubMed

Mills, Evanna L; O'Neill, Luke A

2016-01-01

Mitochondria are master regulators of metabolism. Mitochondria generate ATP by oxidative phosphorylation using pyruvate (derived from glucose and glycolysis) and fatty acids (FAs), both of which are oxidized in the Krebs cycle, as fuel sources. Mitochondria are also an important source of reactive oxygen species (ROS), creating oxidative stress in various contexts, including in the response to bacterial infection. Recently, complex changes in mitochondrial metabolism have been characterized in mouse macrophages in response to varying stimuli in vitro. In LPS and IFN-γ-activated macrophages (M1 macrophages), there is decreased respiration and a broken Krebs cycle, leading to accumulation of succinate and citrate, which act as signals to alter immune function. In IL-4-activated macrophages (M2 macrophages), the Krebs cycle and oxidative phosphorylation are intact and fatty acid oxidation (FAO) is also utilized. These metabolic alterations in response to the nature of the stimulus are proving to be determinants of the effector functions of M1 and M2 macrophages. Furthermore, reprogramming of macrophages from M1 to M2 can be achieved by targeting metabolic events. Here, we describe the role that metabolism plays in macrophage function in infection and immunity, and propose that reprogramming with metabolic inhibitors might be a novel therapeutic approach for the treatment of inflammatory diseases. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

SOCS3 promotes TLR4 response in macrophages by feedback inhibiting TGF-beta1/Smad3 signaling.

PubMed

Liu, Xia; Zhang, Yongliang; Yu, Yizhi; Yang, Xiao; Cao, Xuetao

2008-03-01

Endogenous transforming growth factor-beta1 (TGF-beta1) plays an important role in the negative regulation of toll-like receptor (TLR) signaling in a feedback manner. Suppressors of cytokine signaling 3 (SOCS3) has been shown to be induced by TGF-beta1 in osteoclast/macrophage, while the reports on the role of SOCS3 in regulating TLR4 signaling were controversial. The functional relationship between SOCS3 and TGF-beta1/Smad3 pathway in TLR4 response also remains unclear. In this study, we demonstrate that LPS-induced endogenous TGF-beta1 contributes to the inducible SOCS3 expression in macrophages. SOCS3 silencing could markedly decrease the LPS-induced production of TNF-alpha and IL-6 in macrophages. Interestingly, less decrease of LPS-induced TNF-alpha, IL-6 by SOCS3 silencing was observed in Smad3 null macrophages. Furthermore, we found SOCS3 could interact with Smad3, and inhibit Smad3 nuclear translocation and transcriptional activity. Therefore, our data demonstrate that SOCS3 is a positive regulator of TLR4 response by feedback inhibiting endogenous TGF-beta1/Smad3 signaling, thus outlining a new feedback regulatory manner for TLR4 response in macrophages.

Suppression of the lipopolysaccharide-induced expression of MARCKS-related protein (MRP) affects transmigration in activated RAW264.7 cells.

PubMed

Chun, Kwang-Rok; Bae, Eun Mi; Kim, Jae-Kwan; Suk, Kyoungho; Lee, Won-Ha

2009-01-01

The molecular action mechanism of MRP, one of the protein kinase C (PKC) substrates, has been under intense investigation, but reports on its role in macrophage function remain controversial. The treatment of macrophage cell lines with bacterial lipopolysaccharide (LPS) induced a high level of MRP expression suggesting that MRP plays a role in the function of activated macrophages. In order to investigate the role of MRP in activated RAW264.7 cells, we stably transfected MRP-specific shRNA expression constructs and tested for alterations in macrophage-related functions. The down-regulation of MRP expression resulted in a marked reduction in chemotaxis toward MCP-1 or extracellular matrix proteins. Furthermore, pharmacological inhibitors of PKC significantly inhibited the chemotaxis in RAW264.7 cells. These data reveals the pivotal role of MRP in the transmigration of activated RAW264.7 cells.

Apoptosis is an innate defense function of macrophages against Mycobacterium tuberculosis

PubMed Central

Behar, SM; Martin, CJ; Booty, MG; Nishimura, T; Zhao, X; Gan, H; Divangahi, M; Remold, HG

2011-01-01

Two different forms of death are commonly observed when Mycobacterium tuberculosis (Mtb)-infected macrophages die: (i) necrosis, a death modality defined by cell lysis and (ii) apoptosis, a form of death that maintains an intact plasma membrane. Necrosis is a mechanism used by bacteria to exit the macrophage, evade host defenses, and spread. In contrast, apoptosis of infected macrophages is associated with diminished pathogen viability. Apoptosis occurs when tumor necrosis factor activates the extrinsic death domain pathway, leading to caspase-8 activation. In addition, mitochondrial outer membrane permeabilization leading to activation of the intrinsic apoptotic pathway is required. Both pathways lead to caspase-3 activation, which results in apoptosis. We have recently demonstrated that during mycobacterial infection, cell death is regulated by the eicosanoids, prostaglandin E2 (proapoptotic) and lipoxin (LX)A4 (pronecrotic). Although PGE2 protects against necrosis, virulent Mtb induces LXA4 and inhibits PGE2 production. Under such conditions, mitochondrial inner membrane damage leads to macrophage necrosis. Thus, virulent Mtb subverts eicosanoid regulation of cell death to foil innate defense mechanisms of the macrophage. PMID:21307848

Apoptosis is an innate defense function of macrophages against Mycobacterium tuberculosis.

PubMed

Behar, S M; Martin, C J; Booty, M G; Nishimura, T; Zhao, X; Gan, H-X; Divangahi, M; Remold, H G

2011-05-01

Two different forms of death are commonly observed when Mycobacterium tuberculosis (Mtb)-infected macrophages die: (i) necrosis, a death modality defined by cell lysis and (ii) apoptosis, a form of death that maintains an intact plasma membrane. Necrosis is a mechanism used by bacteria to exit the macrophage, evade host defenses, and spread. In contrast, apoptosis of infected macrophages is associated with diminished pathogen viability. Apoptosis occurs when tumor necrosis factor activates the extrinsic death domain pathway, leading to caspase-8 activation. In addition, mitochondrial outer membrane permeabilization leading to activation of the intrinsic apoptotic pathway is required. Both pathways lead to caspase-3 activation, which results in apoptosis. We have recently demonstrated that during mycobacterial infection, cell death is regulated by the eicosanoids, prostaglandin E(2) (proapoptotic) and lipoxin (LX)A(4) (pronecrotic). Although PGE(2) protects against necrosis, virulent Mtb induces LXA(4) and inhibits PGE(2) production. Under such conditions, mitochondrial inner membrane damage leads to macrophage necrosis. Thus, virulent Mtb subverts eicosanoid regulation of cell death to foil innate defense mechanisms of the macrophage.

Nontransformed, GM-CSF-dependent macrophage lines are a unique model to study tissue macrophage functions.

PubMed

Fejer, György; Wegner, Mareike Dorothee; Györy, Ildiko; Cohen, Idan; Engelhard, Peggy; Voronov, Elena; Manke, Thomas; Ruzsics, Zsolt; Dölken, Lars; Prazeres da Costa, Olivia; Branzk, Nora; Huber, Michael; Prasse, Antje; Schneider, Robert; Apte, Ron N; Galanos, Chris; Freudenberg, Marina A

2013-06-11

Macrophages are diverse cell types in the first line of antimicrobial defense. Only a limited number of primary mouse models exist to study their function. Bone marrow-derived, macrophage-CSF-induced cells with a limited life span are the most common source. We report here a simple method yielding self-renewing, nontransformed, GM-CSF/signal transducer and activator of transcription 5-dependent macrophages (Max Planck Institute cells) from mouse fetal liver, which reflect the innate immune characteristics of alveolar macrophages. Max Planck Institute cells are exquisitely sensitive to selected microbial agents, including bacterial LPS, lipopeptide, Mycobacterium tuberculosis, cord factor, and adenovirus and mount highly proinflammatory but no anti-inflammatory IL-10 responses. They show a unique pattern of innate responses not yet observed in other mononuclear phagocytes. This includes differential LPS sensing and an unprecedented regulation of IL-1α production upon LPS exposure, which likely plays a key role in lung inflammation in vivo. In conclusion, Max Planck Institute cells offer an useful tool to study macrophage biology and for biomedical science.

Regulation of Macrophage Recognition through the Interplay of Nanoparticle Surface Functionality and Protein Corona.

PubMed

Saha, Krishnendu; Rahimi, Mehran; Yazdani, Mahdieh; Kim, Sung Tae; Moyano, Daniel F; Hou, Singyuk; Das, Ridhha; Mout, Rubul; Rezaee, Farhad; Mahmoudi, Morteza; Rotello, Vincent M

2016-04-26

Using a family of cationic gold nanoparticles (NPs) with similar size and charge, we demonstrate that proper surface engineering can control the nature and identity of protein corona in physiological serum conditions. The protein coronas were highly dependent on the hydrophobicity and arrangement of chemical motifs on NP surface. The NPs were uptaken in macrophages in a corona-dependent manner, predominantly through recognition of specific complement proteins in the NP corona. Taken together, this study shows that surface functionality can be used to tune the protein corona formed on NP surface, dictating the interaction of NPs with macrophages.

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

PubMed

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

2015-09-01

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

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.

PubMed

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.

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

Kodali, Vamsi; Littke, Matthew H.; Tilton, Susan C.

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 responsemore » 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 strategies should therefore consider how exposure to these materials alters susceptibility to other environmental exposures.« less

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

PubMed

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

2015-01-01

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

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

PubMed Central

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

2015-01-01

Cells of the immune system are highly sensitive to altered gravity, and the monocyte as well as the macrophage function is proven to be impaired under microgravity conditions. In our study, we investigated the surface expression of ICAM-1 protein and expression of ICAM-1 mRNA in cells of the monocyte/macrophage system in microgravity during clinostat, parabolic flight, sounding rocket, and orbital experiments. In murine BV-2 microglial cells, we detected a downregulation of ICAM-1 expression in clinorotation experiments and a rapid and reversible downregulation in the microgravity phase of parabolic flight experiments. In contrast, ICAM-1 expression increased in macrophage-like differentiated human U937 cells during the microgravity phase of parabolic flights and in long-term microgravity provided by a 2D clinostat or during the orbital SIMBOX/Shenzhou-8 mission. In nondifferentiated U937 cells, no effect of microgravity on ICAM-1 expression could be observed during parabolic flight experiments. We conclude that disturbed immune function in microgravity could be a consequence of ICAM-1 modulation in the monocyte/macrophage system, which in turn could have a strong impact on the interaction with T lymphocytes and cell migration. Thus, ICAM-1 can be considered as a rapid-reacting and sustained gravity-regulated molecule in mammalian cells. PMID:25654110

IL-25 Elicits Innate Lymphoid Cells and Multipotent Progenitor Type 2 Cells That Reduce Renal Ischemic/Reperfusion Injury

PubMed Central

Huang, Qingsong; Niu, Zhiguo; Tan, Jing; Yang, Jun; Liu, Yun; Ma, Haijun; Lee, Vincent W.S.; Sun, Shuming; Song, Xiangfeng; Guo, Minghao; Wang, Yiping

2015-01-01

IL-25 is an important immune regulator that can promote Th2 immune response-dependent immunity, inflammation, and tissue repair in asthma, intestinal infection, and autoimmune diseases. In this study, we examined the effects of IL-25 in renal ischemic/reperfusion injury (IRI). Treating IRI mice with IL-25 significantly improved renal function and reduced renal injury. Furthermore, IL-25 treatment increased the levels of IL-4, IL-5, and IL-13 in serum and kidney and promoted induction of alternatively activated (M2) macrophages in kidney. Notably, IL-25 treatment also increased the frequency of type 2 innate lymphoid cells (ILC2s) and multipotent progenitor type 2 (MPPtype2) cells in kidney. IL-25–responsive ILC2 and MPPtype2 cells produced greater amounts of Th2 cytokines that associated with the induction of M2 macrophages and suppression of classically activated (M1) macrophages in vitro. Finally, adoptive transfer of ILC2s or MPPtype2 cells not only reduced renal functional and histologic injury in IRI mice but also induced M2 macrophages in kidney. In conclusion, our data identify a mechanism whereby IL-25-elicited ILC2 and MPPtype2 cells regulate macrophage phenotype in kidney and prevent renal IRI. PMID:25556172

Src family kinase expression and subcellular localization in macrophages: implications for their role in CSF-1-induced macrophage migration.

PubMed

Dwyer, Amy R; Mouchemore, Kellie A; Steer, James H; Sunderland, Andrew J; Sampaio, Natalia G; Greenland, Eloise L; Joyce, David A; Pixley, Fiona J

2016-07-01

A major role of colony-stimulating factor-1 is to stimulate the differentiation of mononuclear phagocytic lineage cells into adherent, motile, mature macrophages. The colony-stimulating factor-1 receptor transduces colony-stimulating factor-1 signaling, and we have shown previously that phosphatidylinositol 3-kinase p110δ is a critical mediator of colony-stimulating factor-1-stimulated motility through the colony-stimulating factor-1 receptor pY721 motif. Src family kinases are also implicated in the regulation of macrophage motility and in colony-stimulating factor-1 receptor signaling, although functional redundancy of the multiple SFKs expressed in macrophages makes it challenging to delineate their specific functions. We report a comprehensive analysis of individual Src family kinase expression in macrophage cell lines and primary macrophages and demonstrate colony-stimulating factor-1-induced changes in Src family kinase subcellular localization, which provides clues to their distinct and redundant functions in macrophages. Moreover, expression of individual Src family kinases is both species specific and dependent on colony-stimulating factor-1-induced macrophage differentiation. Hck associated with the activated colony-stimulating factor-1 receptor, whereas Lyn associated with the receptor in a constitutive manner. Consistent with this, inhibitor studies revealed that Src family kinases were important for both colony-stimulating factor-1 receptor activation and colony-stimulating factor-1-induced macrophage spreading, motility, and invasion. Distinct colony-stimulating factor-1-induced changes in the subcellular localization of individual SFKs suggest specific roles for these Src family kinases in the macrophage response to colony-stimulating factor-1. © Society for Leukocyte Biology.

The influence of aging and estradiol to progesterone ratio on rat macrophage phenotypic profile and NO and TNF-α production.

PubMed

Dimitrijević, Mirjana; Stanojević, Stanislava; Kuštrimović, Nataša; Mitić, Katarina; Vujić, Vesna; Aleksić, Iva; Radojević, Katarina; Leposavić, Gordana

2013-11-01

The phenotype and function of tissue macrophages substantially depend on the cellular milieu and biological effector molecules, such as steroid hormones, to which they are exposed. Furthermore, in female rats, aging is associated with the altered macrophage functioning and the increased estrogen level is followed by a decrease in that of progesterone. Therefore, the present study aimed to investigate the influence of estradiol/progesterone balance on rat macrophage function and phenotype throughout whole adult lifespan. We ovariectomized rats at the late prepubertal age or at the very end of reproductive lifespan, and examined the expression of ED2 (CD163, a marker of mature resident macrophages related to secretion of inflammatory mediators) on peritoneal macrophages and their ability to produce TNF-α and NO upon LPS-stimulation at different age points. In addition, to delineate direct and indirect effects of estrogen, we assessed the in vitro influence of different concentrations of 17β-estradiol on LPS-induced macrophage TNF-α and NO production. Results showed that: (a) the low frequency of ED2(high) cells amongst peritoneal macrophages of aged rats was accompanied with the reduced TNF-α, but not NO production; (b) estradiol level gradually increased following ovariectomy; (c) macrophage ED2 expression and TNF-α production were dependent on estradiol/progesterone balance and they changed in the same direction; (d) changes in estradiol/progesterone balance differentially affected macrophages TNF-α and NO production; and (e) estradiol exerted pro-inflammatory and anti-inflammatory effects on macrophages in vivo and in vitro, respectively. Overall, our study discloses that estradiol/progesterone balance contributes to the fine-tuning of rat macrophage secretory capacity, and adds to a better understanding of the ovarian steroid hormone role in the regulation of macrophage function, and its significance for the age-associated changes in innate immunity. © 2013.

Delayed growth of EL4 lymphoma in SR-A-deficient mice is due to upregulation of nitric oxide and interferon-gamma production by tumor-associated macrophages.

PubMed

Komohara, Yoshihiro; Takemura, Kenichi; Lei, Xiao Feng; Sakashita, Naomi; Harada, Mamoru; Suzuki, Hiroshi; Kodama, Tatsuhiko; Takeya, Motohiro

2009-11-01

Class A scavenger receptors (SR-A, CD204) are highly expressed in tumor-associated macrophages (TAM). To investigate the function of SR-A in TAM, wild-type and SR-A-deficient (SR-A(-/-)) mice were injected with EL4 cells. Although these groups of mice did not differ in the numbers of infiltrating macrophages and lymphocytes and in neovascularization, SR-A(-/-) mice had delayed growth of EL4 tumors. Expression of inducible nitric oxide (NO) synthase and interferon (IFN)-gamma mRNA increased significantly in tumor tissues from SR-A(-/-) mice. Engulfment of necrotic EL4 cells induced upregulation of NO and IFN-gamma production by cultured macrophages, and production of NO and IFN-gamma increased in SR-A(-/-) macrophages in vitro. IFN-beta production by cultured macrophages was also elevated in SR-A(-/-) macrophages in vitro. These results suggested that the antitumor activity of macrophages increased in SR-A(-/-) mice because of upregulation of NO and IFN-gamma production. These data indicate an important role of SR-A in regulating TAM function by inhibiting toll-like receptor (TLR)4-IFN-beta signaling.

Hypoxia regulates macrophage functions in inflammation.

PubMed

Murdoch, Craig; Muthana, Munitta; Lewis, Claire E

2005-11-15

The presence of areas of hypoxia is a prominent feature of various inflamed, diseased tissues, including malignant tumors, atherosclerotic plaques, myocardial infarcts, the synovia of joints with rheumatoid arthritis, healing wounds, and sites of bacterial infection. These areas form when the blood supply is occluded and/or unable to keep pace with the growth and/or infiltration of inflammatory cells in a given area. Macrophages are present in all tissues of the body where they normally assist in guarding against invading pathogens and regulate normal cell turnover and tissue remodeling. However, they are also known to accumulate in large numbers in such ischemic/hypoxic sites. Recent studies show that macrophages then respond rapidly to the hypoxia present by altering their expression of a wide array of genes. In the present study, we outline and compare the phenotypic responses of macrophages to hypoxia in different diseased states and the implications of these for their progression and treatment.

Increased expression of T cell immunoglobulin and mucin domain 3 aggravates brain inflammation via regulation of the function of microglia/macrophages after intracerebral hemorrhage in mice.

PubMed

Xu, ChangJun; Wang, Tao; Cheng, Si; Liu, YuGuang

2013-12-01

Microglia/macrophages are known to play important roles in initiating brain inflammation after spontaneous intracerebral hemorrhage (ICH). T cell immunoglobulin and mucin domain-3 (Tim-3) have been proven to play a critical part in several inflammatory diseases through regulation of both adaptive and innate immune responses. Tim-3 can be expressed by microglia/macrophages and regulates their function in the innate immune response. However, the effect of Tim-3 on inflammatory responses following ICH is unclear. In this study, we investigated Tim-3 expression, the inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), and brain water content in peri-hematomal brain tissue at 12 hours and at 1, 3, 5, and 7 days post-ICH in wild type (WT) ICH and Tim-3-/- ICH mice. The numbers of Tim-3 positive cells,astrocytes, neutrophils and microglia/macrophages were detected using immunofluorescence staining. Cytokines were measured by ELISA. Double immunofluorescence labeling was performed to identify the cellular source of Tim-3 expression. Mouse neurological deficit scores were assessed through animal behavior. Expression of Tim-3 increased early in mouse peri-hematomal brain tissue after autologous blood injection, peaked at day 1, and was positively correlated with the concentrations of TNF-α, IL-1β, and brain water content. Tim-3 was predominantly expressed in microglia/macrophages. Compared with WT mice, Tim-3-/- mice had reduced ICH-induced brain inflammation with decreased TNF-α and IL-1β, cerebral edema and neurological deficit scores. Moreover, Tim-/- inhibited activation of microglia/macrophages. The number of activated microglia/macrophages in Tim-3-/- ICH mice was much lower than that in WT ICH mice. Our findings demonstrate that Tim-3 plays an important role in brain inflammation after ICH, and may be a potential treatment target.

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

PubMed

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.

[Advances in nanoparticle-targeting tumor associated macrophages for cancer imaging and therapy].

PubMed

Fengliang, Guo; Guping, Tang; Qinglian, H U

2017-03-25

Tumor tissues are composed of tumor cells and complicate microenvironment. Tumor associated macrophages (TAMs) as an important component in tumor microenvironment, play fundamental roles in tumor progression, metastasis and microenvironment regulation. Recently, studies have found that nanotechnology, as an emerging platform, provides unique potential for cancer imaging and therapy. With the nanotechnology, TAMs imaging presents direct evidence for cancer development, progression, and the effectiveness of cancer treatments; it also can regulate the immunosuppression of tumor microenvironment and improve therapeutic efficiency through TAMs targeted killing or phenotypic transformation. In this article, we illustrate the function of TAMs and review the latest development in nano-carriers and their applications in tumor associated macrophage targeting cancer imaging and therapy.

Regulation of Embryonic and Postnatal Development by the CSF-1 Receptor

PubMed Central

Chitu, Violeta; Stanley, E. Richard

2017-01-01

Macrophages are found in all tissues and regulate tissue morphogenesis during development through trophic and scavenger functions. The colony stimulating factor-1 (CSF-1) receptor (CSF-1R) is the major regulator of tissue macrophage development and maintenance. In combination with receptor activator of nuclear factor κB (RANK), the CSF-1R also regulates the differentiation of the bone-resorbing osteoclast and controls bone remodeling during embryonic and early postnatal development. CSF-1R-regulated macrophages play trophic and remodeling roles in development. Outside the mononuclear phagocytic system, the CSF-1R directly regulates neuronal survival and differentiation, the development of intestinal Paneth cells and of preimplantation embryos, as well as trophoblast innate immune function. Consistent with the pleiotropic roles of the receptor during development, CSF-1R deficiency in most mouse strains causes embryonic or perinatal death and the surviving mice exhibit multiple developmental and functional deficits. The CSF-1R is activated by two dimeric glycoprotein ligands, CSF-1, and interleukin-34 (IL-34). Homozygous Csf1-null mutations phenocopy most of the deficits of Csf1r-null mice. In contrast, Il34-null mice have no gross phenotype, except for decreased numbers of Langerhans cells and microglia, indicating that CSF-1 plays the major developmental role. Homozygous inactivating mutations of the Csf1r or its ligands have not been reported in man. However, heterozygous inactivating mutations in the Csf1r lead to a dominantly inherited adult-onset progressive dementia, highlighting the importance of CSF-1R signaling in the brain. PMID:28236968

Regulation of Embryonic and Postnatal Development by the CSF-1 Receptor.

PubMed

Chitu, Violeta; Stanley, E Richard

2017-01-01

Macrophages are found in all tissues and regulate tissue morphogenesis during development through trophic and scavenger functions. The colony stimulating factor-1 (CSF-1) receptor (CSF-1R) is the major regulator of tissue macrophage development and maintenance. In combination with receptor activator of nuclear factor κB (RANK), the CSF-1R also regulates the differentiation of the bone-resorbing osteoclast and controls bone remodeling during embryonic and early postnatal development. CSF-1R-regulated macrophages play trophic and remodeling roles in development. Outside the mononuclear phagocytic system, the CSF-1R directly regulates neuronal survival and differentiation, the development of intestinal Paneth cells and of preimplantation embryos, as well as trophoblast innate immune function. Consistent with the pleiotropic roles of the receptor during development, CSF-1R deficiency in most mouse strains causes embryonic or perinatal death and the surviving mice exhibit multiple developmental and functional deficits. The CSF-1R is activated by two dimeric glycoprotein ligands, CSF-1, and interleukin-34 (IL-34). Homozygous Csf1-null mutations phenocopy most of the deficits of Csf1r-null mice. In contrast, Il34-null mice have no gross phenotype, except for decreased numbers of Langerhans cells and microglia, indicating that CSF-1 plays the major developmental role. Homozygous inactivating mutations of the Csf1r or its ligands have not been reported in man. However, heterozygous inactivating mutations in the Csf1r lead to a dominantly inherited adult-onset progressive dementia, highlighting the importance of CSF-1R signaling in the brain. © 2017 Elsevier Inc. All rights reserved.

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

PubMed Central

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

1996-01-01

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

Infection-induced regulation of natural killer cells by macrophages and collagen at the lymph node subcapsular sinus.

PubMed

Coombes, Janine L; Han, Seong-Ji; van Rooijen, Nico; Raulet, David H; Robey, Ellen A

2012-07-26

Infection leads to heightened activation of natural killer (NK) cells, a process that likely involves direct cell-to-cell contact, but how this occurs in vivo is poorly understood. We have used two-photon laser-scanning microscopy in conjunction with Toxoplasma gondii mouse infection models to address this question. We found that after infection, NK cells accumulated in the subcapsular region of the lymph node, where they formed low-motility contacts with collagen fibers and CD169(+) macrophages. We provide evidence that interactions with collagen regulate NK cell migration, whereas CD169(+) macrophages increase the activation state of NK cells. Interestingly, a subset of CD169(+) macrophages that coexpress the inflammatory monocyte marker Ly6C had the most potent ability to activate NK cells. Our data reveal pathways through which NK cell migration and function are regulated after infection and identify an important accessory cell population for activation of NK cell responses in lymph nodes. Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.

Granulocyte-Macrophage Colony Stimulatory Factor Enhances the Pro-Inflammatory Response of Interferon-γ-Treated Macrophages to Pseudomonas aeruginosa Infection

PubMed Central

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 infected with P. aeruginosa: untreated < treated with GM-CSF < treated with IFN-γ < treated with GM-CSF and IFN-γ. PMID:25706389

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

PubMed

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 infected with P. aeruginosa: untreated < treated with GM-CSF < treated with IFN-γ < treated with GM-CSF and IFN-γ.

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

PubMed

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.

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

PubMed Central

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

2016-01-01

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

The Role of AIRE in the Immunity Against Candida Albicans in a Model of Human Macrophages.

PubMed

de Albuquerque, Jose Antonio Tavares; Banerjee, Pinaki Prosad; Castoldi, Angela; Ma, Royce; Zurro, Nuria Bengala; Ynoue, Leandro Hideki; Arslanian, Christina; Barbosa-Carvalho, Marina Uchoa Wall; Correia-Deur, Joya Emilie de Menezes; Weiler, Fernanda Guimarães; Dias-da-Silva, Magnus Regios; Lazaretti-Castro, Marise; Pedroza, Luis Alberto; Câmara, Niels Olsen Saraiva; Mace, Emily; Orange, Jordan Scott; Condino-Neto, Antonio

2018-01-01

Autoimmune-polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a primary immunodeficiency caused by mutations in the autoimmune regulator gene ( AIRE ). Patients with AIRE mutations are susceptible to Candida albicans infection and present with autoimmune disorders. We previously demonstrated that cytoplasmic AIRE regulates the Syk-dependent Dectin-1 pathway. In this study, we further evaluated direct contact with fungal elements, synapse formation, and the response of macrophage-like THP-1 cells to C. albicans hyphae to determine the role of AIRE upon Dectin receptors function and signaling. We examined the fungal synapse (FS) formation in wild-type and AIRE-knockdown THP-1 cells differentiated to macrophages, as well as monocyte-derived macrophages from APECED patients. We evaluated Dectin-2 receptor signaling, phagocytosis, and cytokine secretion upon hyphal stimulation. AIRE co-localized with Dectin-2 and Syk at the FS upon hyphal stimulation of macrophage-like THP-1 cells. AIRE-knockdown macrophage-like THP-1 cells exhibited less Dectin-1 and Dectin-2 receptors accumulation, decreased signaling pathway activity at the FS, lower C. albicans phagocytosis, and less lysosome formation. Furthermore, IL-1β, IL-6, or TNF-α secretion by AIRE-knockdown macrophage-like THP-1 cells and AIRE-deficient patient macrophages was decreased compared to control cells. Our results suggest that AIRE modulates the FS formation and hyphal recognition and help to orchestrate an effective immune response against C. albicans .

Therapeutic potential of carbohydrates as regulators of macrophage activation.

PubMed

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.

MiR-155-regulated molecular network orchestrates cell fate in the innate and adaptive immune response to Mycobacterium tuberculosis.

PubMed

Rothchild, Alissa C; Sissons, James R; Shafiani, Shahin; Plaisier, Christopher; Min, Deborah; Mai, Dat; Gilchrist, Mark; Peschon, Jacques; Larson, Ryan P; Bergthaler, Andreas; Baliga, Nitin S; Urdahl, Kevin B; Aderem, Alan

2016-10-11

The regulation of host-pathogen interactions during Mycobacterium tuberculosis (Mtb) infection remains unresolved. MicroRNAs (miRNAs) are important regulators of the immune system, and so we used a systems biology approach to construct an miRNA regulatory network activated in macrophages during Mtb infection. Our network comprises 77 putative miRNAs that are associated with temporal gene expression signatures in macrophages early after Mtb infection. In this study, we demonstrate a dual role for one of these regulators, miR-155. On the one hand, miR-155 maintains the survival of Mtb-infected macrophages, thereby providing a niche favoring bacterial replication; on the other hand, miR-155 promotes the survival and function of Mtb-specific T cells, enabling an effective adaptive immune response. MiR-155-induced cell survival is mediated through the SH2 domain-containing inositol 5-phosphatase 1 (SHIP1)/protein kinase B (Akt) pathway. Thus, dual regulation of the same cell survival pathway in innate and adaptive immune cells leads to vastly different outcomes with respect to bacterial containment.

Profiling post-translational modifications of histones in human monocyte-derived macrophages.

PubMed

Olszowy, Pawel; Donnelly, Maire Rose; Lee, Chanho; Ciborowski, Pawel

2015-01-01

Histones and their post-translational modifications impact cellular function by acting as key regulators in the maintenance and remodeling of chromatin, thus affecting transcription regulation either positively (activation) or negatively (repression). In this study we describe a comprehensive, bottom-up proteomics approach to profiling post-translational modifications (acetylation, mono-, di- and tri-methylation, phosphorylation, biotinylation, ubiquitination, citrullination and ADP-ribosylation) in human macrophages, which are primary cells of the innate immune system. As our knowledge expands, it becomes more evident that macrophages are a heterogeneous population with potentially subtle differences in their responses to various stimuli driven by highly complex epigenetic regulatory mechanisms. To profile post-translational modifications (PTMs) of histones in macrophages we used two platforms of liquid chromatography and mass spectrometry. One platform was based on Sciex5600 TripleTof and the second one was based on VelosPro Orbitrap Elite ETD mass spectrometers. We provide side-by-side comparison of profiling using two mass spectrometric platforms, ion trap and qTOF, coupled with the application of collisional induced and electron transfer dissociation. We show for the first time methylation of a His residue in macrophages and demonstrate differences in histone PTMs between those currently reported for macrophage cell lines and what we identified in primary cells. We have found a relatively low level of histone PTMs in differentiated but resting human primary monocyte derived macrophages. This study is the first comprehensive profiling of histone PTMs in primary human MDM. Our study implies that epigenetic regulatory mechanisms operative in transformed cell lines and primary cells are overlapping to a limited extent. Our mass spectrometric approach provides groundwork for the investigation of how histone PTMs contribute to epigenetic regulation in primary human macrophages.

Deregulation of PPARβ/δ target genes in tumor-associated macrophages by fatty acid ligands in the ovarian cancer microenvironment.

PubMed

Schumann, Tim; Adhikary, Till; Wortmann, Annika; Finkernagel, Florian; Lieber, Sonja; Schnitzer, Evelyn; Legrand, Nathalie; Schober, Yvonne; Nockher, W Andreas; Toth, Philipp M; Diederich, Wibke E; Nist, Andrea; Stiewe, Thorsten; Wagner, Uwe; Reinartz, Silke; Müller-Brüsselbach, Sabine; Müller, Rolf

2015-05-30

The nuclear receptor peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a lipid ligand-inducible transcription factor associated with macrophage polarization. However, its function in tumor-associated macrophages (TAMs) has not been investigated to date. Here, we report the PPARβ/δ-regulated transcriptome and cistrome for TAMs from ovarian carcinoma patients. Comparison with monocyte-derived macrophages shows that the vast majority of direct PPARβ/δ target genes are upregulated in TAMs and largely refractory to synthetic agonists, but repressible by inverse agonists. Besides genes with metabolic functions, these include cell type-selective genes associated with immune regulation and tumor progression, e.g., LRP5, CD300A, MAP3K8 and ANGPTL4. This deregulation is not due to increased expression of PPARβ/δ or its enhanced recruitment to target genes. Instead, lipidomic analysis of malignancy-associated ascites revealed high concentrations of polyunsaturated fatty acids, in particular linoleic acid, acting as potent PPARβ/δ agonists in macrophages. These fatty acid ligands accumulate in lipid droplets in TAMs, thereby providing a reservoir of PPARβ/δ ligands. These observations suggest that the deregulation of PPARβ/δ target genes by ligands of the tumor microenvironment contributes to the pro-tumorigenic polarization of ovarian carcinoma TAMs. This conclusion is supported by the association of high ANGPTL4 expression with a shorter relapse-free survival in serous ovarian carcinoma.

Deregulation of PPARβ/δ target genes in tumor-associated macrophages by fatty acid ligands in the ovarian cancer microenvironment

PubMed Central

Finkernagel, Florian; Lieber, Sonja; Schnitzer, Evelyn; Legrand, Nathalie; Schober, Yvonne; Nockher, W. Andreas; Toth, Philipp M.; Diederich, Wibke E.; Nist, Andrea; Stiewe, Thorsten; Wagner, Uwe; Reinartz, Silke; Müller-Brüsselbach, Sabine; Müller, Rolf

2015-01-01

The nuclear receptor peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a lipid ligand-inducible transcription factor associated with macrophage polarization. However, its function in tumor-associated macrophages (TAMs) has not been investigated to date. Here, we report the PPARβ/δ-regulated transcriptome and cistrome for TAMs from ovarian carcinoma patients. Comparison with monocyte-derived macrophages shows that the vast majority of direct PPARβ/δ target genes are upregulated in TAMs and largely refractory to synthetic agonists, but repressible by inverse agonists. Besides genes with metabolic functions, these include cell type-selective genes associated with immune regulation and tumor progression, e.g., LRP5, CD300A, MAP3K8 and ANGPTL4. This deregulation is not due to increased expression of PPARβ/δ or its enhanced recruitment to target genes. Instead, lipidomic analysis of malignancy-associated ascites revealed high concentrations of polyunsaturated fatty acids, in particular linoleic acid, acting as potent PPARβ/δ agonists in macrophages. These fatty acid ligands accumulate in lipid droplets in TAMs, thereby providing a reservoir of PPARβ/δ ligands. These observations suggest that the deregulation of PPARβ/δ target genes by ligands of the tumor microenvironment contributes to the pro-tumorigenic polarization of ovarian carcinoma TAMs. This conclusion is supported by the association of high ANGPTL4 expression with a shorter relapse-free survival in serous ovarian carcinoma. PMID:25968567

Oxygen tension limits nitric oxide synthesis by activated macrophages.

PubMed Central

McCormick, C C; Li, W P; Calero, M

2000-01-01

Previous studies have established that constitutive calcium-dependent ('low-output') nitric oxide synthase (NOS) is regulated by oxygen tension. We have investigated the role of oxygen tension in the synthesis of NO by the 'high-output' calcium-independent NOS in activated macrophages. Hypoxia increased macrophage NOS gene expression in the presence of one additional activator, such as lipopolysaccharide or interferon-gamma, but not in the presence of both. Hypoxia markedly reduced the synthesis of NO by activated macrophages (as measured by accumulation of nitrite and citrulline), such that, at 1% oxygen tension, NO accumulation was reduced by 80-90%. The apparent K(m) for oxygen calculated from cells exposed to a range of oxygen tensions was found to be 10.8%, or 137 microM, O(2) This value is considerably higher than the oxygen tension in tissues, and is virtually identical to that reported recently for purified recombinant macrophage NOS. The decrease in NO synthesis did not appear to be due to diminished arginine or cofactor availability, since arginine transport and NO synthesis during recovery in normoxia were normal. Analysis of NO synthesis during hypoxia as a function of extracellular arginine indicated that an altered V(max), but not K(m)(Arg), accounted for the observed decrease in NO synthesis. We conclude that oxygen tension regulates the synthesis of NO in macrophages by a mechanism similar to that described previously for the calcium-dependent low-output NOS. Our data suggest that oxygen tension may be an important physiological regulator of macrophage NO synthesis in vivo. PMID:10970783

Brown adipose tissue macrophages control tissue innervation and homeostatic energy expenditure

PubMed Central

Cortese, Nina; Haimon, Zhana; Sar Shalom, Hadas; Kuperman, Yael; Kalchenko, Vyacheslav; Brandis, Alexander; David, Eyal; Segal-Hayoun, Yifat; Chappell-Maor, Louise; Yaron, Avraham; Jung, Steffen

2017-01-01

Tissue macrophages provide immune defense and contribute to establishment and maintenance of tissue homeostasis. Here we used constitutive and inducible mutagenesis to delete the nuclear transcription regulator methyl-CpG binding protein 2 (Mecp2) in defined tissue macrophages. Animals lacking the Rett syndrome-associated gene in macrophages did not show signs of neurodevelopmental disorder, but displayed spontaneous obesity, which could be linked to impaired brown adipose tissue (BAT) function. Specifically, mutagenesis of a BAT-resident CX3CR1+ macrophage subpopulation compromised homeostatic, though not acute cold-induced thermogenesis. Mechanistically, BAT malfunction of pre-obese mice harboring mutant macrophages was associated with decreased sympathetic innervation and local norepinephrine titers, resulting in reduced adipocyte expression of thermogenic factors. Mutant macrophages over-expressed PlexinA4, which might contribute to the phenotype by repulsion of Sema6A-expressing sympathetic axons. Collectively, we report a previously unappreciated homeostatic role of macrophages in the control of tissue innervation, disruption of which in BAT results in metabolic imbalance. PMID:28459435

Developmental origin of lung macrophage diversity

PubMed Central

Tan, Serena Y. S.; Krasnow, Mark A.

2016-01-01

Macrophages are specialized phagocytic cells, present in all tissues, which engulf and digest pathogens, infected and dying cells, and debris, and can recruit and regulate other immune cells and the inflammatory response and aid in tissue repair. Macrophage subpopulations play distinct roles in these processes and in disease, and are typically recognized by differences in marker expression, immune function, or tissue of residency. Although macrophage subpopulations in the brain have been found to have distinct developmental origins, the extent to which development contributes to macrophage diversity between tissues and within tissues is not well understood. Here, we investigate the development and maintenance of mouse lung macrophages by marker expression patterns, genetic lineage tracing and parabiosis. We show that macrophages populate the lung in three developmental waves, each giving rise to a distinct lineage. These lineages express different markers, reside in different locations, renew in different ways, and show little or no interconversion. Thus, development contributes significantly to lung macrophage diversity and targets each lineage to a different anatomical domain. PMID:26952982

The transcriptional PPARβ/δ network in human macrophages defines a unique agonist-induced activation state

PubMed Central

Adhikary, Till; Wortmann, Annika; Schumann, Tim; Finkernagel, Florian; Lieber, Sonja; Roth, Katrin; Toth, Philipp M.; Diederich, Wibke E.; Nist, Andrea; Stiewe, Thorsten; Kleinesudeik, Lara; Reinartz, Silke; Müller-Brüsselbach, Sabine; Müller, Rolf

2015-01-01

Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a lipid ligand-inducible transcription factor with established metabolic functions, whereas its anti-inflammatory function is poorly understood. To address this issue, we determined the global PPARβ/δ-regulated signaling network in human monocyte-derived macrophages. Besides cell type-independent, canonical target genes with metabolic and immune regulatory functions we identified a large number of inflammation-associated NFκB and STAT1 target genes that are repressed by agonists. Accordingly, PPARβ/δ agonists inhibited the expression of multiple pro-inflammatory mediators and induced an anti-inflammatory, IL-4-like morphological phenotype. Surprisingly, bioinformatic analyses also identified immune stimulatory effects. Consistent with this prediction, PPARβ/δ agonists enhanced macrophage survival under hypoxic stress and stimulated CD8+ T cell activation, concomitantly with the repression of immune suppressive target genes and their encoded products CD274 (PD-1 ligand), CD32B (inhibitory Fcγ receptor IIB) and indoleamine 2,3-dioxygenase 1 (IDO-1), as well as a diminished release of the immune suppressive IDO-1 metabolite kynurenine. Comparison with published data revealed a significant overlap of the PPARβ/δ transcriptome with coexpression modules characteristic of both anti-inflammatory and pro-inflammatory cytokines. Our findings indicate that PPARβ/δ agonists induce a unique macrophage activation state with strong anti-inflammatory but also specific immune stimulatory components, pointing to a context-dependent function of PPARβ/δ in immune regulation. PMID:25934804

The immunosuppressive effects of a novel recombinant LipQ (Rv2485c) protein of Mycobacterium tuberculosis on human macrophage cell lines.

PubMed

Kumar, Anjani; Manisha; Sangha, Gurkamaljit Kaur; Shrivastava, Anju; Kaur, Jagdeep

2017-06-01

Mycobacterium tuberculosis (MTB), an intracellular pathogen, still represents a major global health challenge. A number of mycobacterial macromolecules have been shown to target biological processes within host macrophages; however, the exact mechanism for the majority of these host pathogen interactions is still poorly understood. Moreover, the lipid metabolic pathway is one of the most important physiologic pathways that plays a vital role in the survival and infection of Mycobacterium tuberculosis. In present study, we investigated the effect of rLipQ from Mycobacterium tuberculosis H37Rv on macrophage functions in vitro.Our results demonstrate that rLipQ significantly lowers the expression level of pro-inflammatory cytokines (TNF-α& IFN-γ) and augments the level of anti inflammatory cytokines such as IL-4 & IL-10as compared to LPS stimulated macrophages. An up-regulation of anti-inflammatory and down-regulation of pro-inflammatory cytokines levels in rLipQ pretreated macrophages implies immuno-modulatory functions in TB patients. Interestingly, rLipQ also inhibited the expression of iNOS, TLR-2 and transcription factor NF-kB in LPS stimulated macrophages whereas the expression of TLR-4 remains unchanged. The inhibition in the expression of these signaling molecules has been correlated to the inhibition of NO production in macrophages. Taken together, these studies demonstrate that rLipQ is a novel lipase that is highly immunogenic and may play an important role in the virulence and pathogenesis of M. tuberculosis infection, by altering the balance of cytokines, which might help to assess prognosis and contribute to a better understanding against host-pathogen interactions. Copyright © 2017 Elsevier Ltd. All rights reserved.

Protein-tyrosine phosphatase Shp2 positively regulates macrophage oxidative burst.

PubMed

Li, Xing Jun; Goodwin, Charles B; Nabinger, Sarah C; Richine, Briana M; Yang, Zhenyun; Hanenberg, Helmut; Ohnishi, Hiroshi; Matozaki, Takashi; Feng, Gen-Sheng; Chan, Rebecca J

2015-02-13

Macrophages are vital to innate immunity and express pattern recognition receptors and integrins for the rapid detection of invading pathogens. Stimulation of Dectin-1 and complement receptor 3 (CR3) activates Erk- and Akt-dependent production of reactive oxygen species (ROS). Shp2, a protein-tyrosine phosphatase encoded by Ptpn11, promotes activation of Ras-Erk and PI3K-Akt and is crucial for hematopoietic cell function; however, no studies have examined Shp2 function in particulate-stimulated ROS production. Maximal Dectin-1-stimulated ROS production corresponded kinetically to maximal Shp2 and Erk phosphorylation. Bone marrow-derived macrophages (BMMs) from mice with a conditionally deleted allele of Ptpn11 (Shp2(flox/flox);Mx1Cre+) produced significantly lower ROS levels compared with control BMMs. Although YFP-tagged phosphatase dead Shp2-C463A was strongly recruited to the early phagosome, its expression inhibited Dectin-1- and CR3-stimulated phospho-Erk and ROS levels, placing Shp2 phosphatase function and Erk activation upstream of ROS production. Further, BMMs expressing gain of function Shp2-D61Y or Shp2-E76K and peritoneal exudate macrophages from Shp2D61Y/+;Mx1Cre+ mice produced significantly elevated levels of Dectin-1- and CR3-stimulated ROS, which was reduced by pharmacologic inhibition of Erk. SIRPα (signal regulatory protein α) is a myeloid inhibitory immunoreceptor that requires tyrosine phosphorylation to exert its inhibitory effect. YFP-Shp2C463A-expressing cells have elevated phospho-SIRPα levels and an increased Shp2-SIRPα interaction compared with YFP-WT Shp2-expressing cells. Collectively, these findings indicate that Shp2 phosphatase function positively regulates Dectin-1- and CR3-stimulated ROS production in macrophages by dephosphorylating and thus mitigating the inhibitory function of SIRPα and by promoting Erk activation. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Autocrine IL-10 functions as a rheostat for M1 macrophage glycolytic commitment by tuning nitric oxide production.

PubMed

Baseler, Walter A; Davies, Luke C; Quigley, Laura; Ridnour, Lisa A; Weiss, Jonathan M; Hussain, S Perwez; Wink, David A; McVicar, Daniel W

2016-12-01

Inflammatory maturation of M1 macrophages by proinflammatory stimuli such as toll like receptor ligands results in profound metabolic reprogramming resulting in commitment to aerobic glycolysis as evidenced by repression of mitochondrial oxidative phosphorylation (OXPHOS) and enhanced glucose utilization. In contrast, "alternatively activated" macrophages adopt a metabolic program dominated by fatty acid-fueled OXPHOS. Despite the known importance of these developmental stages on the qualitative aspects of an inflammatory response, relatively little is know regarding the regulation of these metabolic adjustments. Here we provide evidence that the immunosuppressive cytokine IL-10 defines a metabolic regulatory loop. Our data show for the first time that lipopolysaccharide (LPS)-induced glycolytic flux controls IL-10-production via regulation of mammalian target of rapamycin (mTOR) and that autocrine IL-10 in turn regulates macrophage nitric oxide (NO) production. Genetic and pharmacological manipulation of IL-10 and nitric oxide (NO) establish that metabolically regulated autocrine IL-10 controls glycolytic commitment by limiting NO-mediated suppression of OXPHOS. Together these data support a model where autocine IL-10 production is controlled by glycolytic flux in turn regulating glycolytic commitment by preserving OXPHOS via suppression of NO. We propose that this IL-10-driven metabolic rheostat maintains metabolic equilibrium during M1 macrophage differentiation and that perturbation of this regulatory loop, either directly by exogenous cellular sources of IL-10 or indirectly via limitations in glucose availability, skews the cellular metabolic program altering the balance between inflammatory and immunosuppressive phenotypes. Copyright © 2016. Published by Elsevier B.V.

Restraint stress alters neutrophil and macrophage phenotypes during wound healing

PubMed Central

Tymen, Stéphanie D.; Rojas, Isolde G.; Zhou, Xiaofeng; Fang, Zong Juan; Zhao, Yan; Marucha, Phillip T.

2013-01-01

Previous studies reported that stress delays wound healing, impairs bacterial clearance, and elevates the risk for opportunistic infection. Neutrophils and macrophages are responsible for the removal of bacteria present at the wound site. The appropriate recruitment and functions of these cells are necessary for efficient bacterial clearance. In our current study we found that restraint stress induced an excessive recruitment of neutrophils extending the inflammatory phase of healing, and the gene expression of neutrophil attracting chemokines MIP-2 and KC. However, restraint stress did not affect macrophage infiltration. Stress decreased the phagocytic abilities of phagocytic cells ex vivo, yet it did not affect superoxide production. The cell surface expression of adhesion molecules CD11b and TLR4 were decreased in peripheral blood monocytes in stressed mice. The phenotype of macrophages present at the wound site was also altered. Gene expression of markers of pro-inflammatory classically activated macrophages, CXCL10 and CCL5, were down-regulated; as were markers associated with wound healing macrophages, CCL22, IGF-1, RELMα; and the regulatory macrophage marker, chemokine CCL1. Restraint stress also induced up-regulation of IL10 gene expression. In summary, our study has shown that restraint stress suppresses the phenotype shift of the macrophage population, as compared to the changes observed during normal wound healing, while the number of macrophages remains constant. We also observed a general suppression of chemokine gene expression. Modulation of the macrophage phenotype could provide a new therapeutic approach in the treatment of wounds under stress conditions in the clinical setting. PMID:22884902

Novel high throughput pooled shRNA screening identifies NQO1 as a potential drug target for host directed therapy for tuberculosis

PubMed Central

Li, Qing; Karim, Ahmad F.; Ding, Xuedong; Das, Biswajit; Dobrowolski, Curtis; Gibson, Richard M.; Quiñones-Mateu, Miguel E.; Karn, Jonathan; Rojas, Roxana E.

2016-01-01

Chemical regulation of macrophage function is one key strategy for developing host-directed adjuvant therapies for tuberculosis (TB). A critical step to develop these therapies is the identification and characterization of specific macrophage molecules and pathways with a high potential to serve as drug targets. Using a barcoded lentivirus-based pooled short-hairpin RNA (shRNA) library combined with next generation sequencing, we identified 205 silenced host genes highly enriched in mycobacteria-resistant macrophages. Twenty-one of these “hits” belonged to the oxidoreductase functional category. NAD(P)H:quinone oxidoreductase 1 (NQO1) was the top oxidoreductase “hit”. NQO1 expression was increased after mycobacterial infection, and NQO1 knockdown increased macrophage differentiation, NF-κB activation, and the secretion of pro-inflammatory cytokines TNF-α and IL-1β in response to infection. This suggests that mycobacteria hijacks NQO1 to down-regulate pro-inflammatory and anti-bacterial functions. The competitive inhibitor of NQO1 dicoumarol synergized with rifampin to promote intracellular killing of mycobacteria. Thus, NQO1 is a new host target in mycobacterial infection that could potentially be exploited to increase antibiotic efficacy in vivo. Our findings also suggest that pooled shRNA libraries could be valuable tools for genome-wide screening in the search for novel druggable host targets for adjunctive TB therapies. PMID:27297123

NOX2 drives M1-like microglial/macrophage activation and neurodegeneration following experimental traumatic brain injury

PubMed Central

Kumar, Alok; Barrett, James P.; Alvarez-Croda, Dulce-Mariely; Stoica, Bogdan A.; Faden, Alan I.; Loane, David J.

2016-01-01

Following traumatic brain injury (TBI), activation of microglia and peripherally derived inflammatory macrophages occurs in association with tissue damage. This neuroinflammatory response may have beneficial or detrimental effects on neuronal survival, depending on the functional polarization of these cells along a continuum from M1-like to M2-like activation states. The mechanisms that regulate M1-like and M2-like activation after TBI are not well understood, but appear in part to reflect the redox state of the lesion microenvironment. NADPH oxidase (NOX2) is a critical enzyme system that generates reactive oxygen species in microglia/macrophages. After TBI, NOX2 is strongly up-regulated in M1-like, but not in M2-like polarized cells. Therefore, we hypothesized that NOX2 drives M1-like neuroinflammation and contributes to neurodegeneration and loss of neurological function after TBI. In the present studies we inhibited NOX2 activity using NOX2-knockout mice or the selective peptide inhibitor gp91ds-tat. We show that NOX2 is highly up-regulated in infiltrating macrophages after injury, and that NOX2 deficiency reduces markers of M1-like activation, limits tissue loss and neurodegeneration, and improves motor recovery after moderate-level control cortical injury (CCI). NOX2 deficiency also promotes M2-like activation after CCI, through increased IL-4Rα signaling in infiltrating macrophages, suggesting that NOX2 acts as a critical switch between M1- and M2-like activation states after TBI. Administration of gp91ds-tat to wild-type CCI mice starting at 24 hours post-injury reduces deficits in cognitive function and increased M2-like activation in the hippocampus. Collectively, our data indicate that increased NOX2 activity after TBI drives M1-like activation that contributes to inflammatory-mediated neurodegeneration, and that inhibiting this pathway provides neuroprotection, in part by altering M1-/M2-like balance towards the M2-like neuroinflammatory response. PMID:27477920

Dual Roles for Ikaros in Regulation of Macrophage Chromatin State and Inflammatory Gene Expression.

PubMed

Oh, Kyu-Seon; Gottschalk, Rachel A; Lounsbury, Nicolas W; Sun, Jing; Dorrington, Michael G; Baek, Songjoon; Sun, Guangping; Wang, Ze; Krauss, Kathleen S; Milner, Joshua D; Dutta, Bhaskar; Hager, Gordon L; Sung, Myong-Hee; Fraser, Iain D C

2018-06-13

Macrophage activation by bacterial LPS leads to induction of a complex inflammatory gene program dependent on numerous transcription factor families. The transcription factor Ikaros has been shown to play a critical role in lymphoid cell development and differentiation; however, its function in myeloid cells and innate immune responses is less appreciated. Using comprehensive genomic analysis of Ikaros-dependent transcription, DNA binding, and chromatin accessibility, we describe unexpected dual repressor and activator functions for Ikaros in the LPS response of murine macrophages. Consistent with the described function of Ikaros as transcriptional repressor, Ikzf1 -/- macrophages showed enhanced induction for select responses. In contrast, we observed a dramatic defect in expression of many delayed LPS response genes, and chromatin immunoprecipitation sequencing analyses support a key role for Ikaros in sustained NF-κB chromatin binding. Decreased Ikaros expression in Ikzf1 +/- mice and human cells dampens these Ikaros-enhanced inflammatory responses, highlighting the importance of quantitative control of Ikaros protein level for its activator function. In the absence of Ikaros, a constitutively open chromatin state was coincident with dysregulation of LPS-induced chromatin remodeling, gene expression, and cytokine responses. Together, our data suggest a central role for Ikaros in coordinating the complex macrophage transcriptional program in response to pathogen challenge.

Counter-regulatory paracrine actions of FGF-23 and 1,25(OH)2D in macrophages

PubMed Central

Han, Xiaobin; Li, Linqiang; Yang, Jiancheng; King, Gwendalyn; Xiao, Zhousheng; Quarles, Leigh Darryl

2016-01-01

Mechanisms underlying the association between fibroblastic growth factor 23 (FGF-23) and inflammation are uncertain. We found that FGF-23 was markedly up-regulated in LPS/INF-γ-induced proinflammatory M1 macrophages and Hyp mouse-derived peritoneal macrophages, but not in IL-4-induced M2 anti-inflammatory macrophages. NF-κB and JAK/STAT1 pathways mediated the increased transcription of FGF-23 in response to M1 polarization. FGF-23 stimulated TNF-α, but not IL-6, expression in M0 macrophages and suppressed Arginase-1 expression in M2 macrophages through FGFR-mediated mechanisms. 1,25(OH)2D stimulated Arginase-1 expression and inhibited FGF-23 stimulation of TNF-α. FGF-23 has proinflammatory paracrine functions and counter-regulatory actions to 1,25(OH)2D on innate immune responses. PMID:26762170

Role of selenium-containing proteins in T cell and macrophage function

PubMed Central

Carlson, Bradley A.; Yoo, Min-Hyuk; Shrimali, Rajeev K.; Irons, Robert; Gladyshev, Vadim N.; Hatfield, Dolph L.; Park, Jin Mo

2011-01-01

Synopsis Selenium has been known for many years to have a role in boosting immune function, but the manner in which this element acts at the molecular level in host defense and inflammatory diseases is poorly understood. To elucidate the role of selenium-containing proteins in immune function, we knocked out the expression of this protein class in T cells or macrophages of mice by targeting the removal of the selenocysteine tRNA gene using loxP-Cre technology. Mice with selenoprotein-less T cells manifested reduced pools of mature and functional T cells in lymphoid tissues and an impairment in T cell-dependent antibody responses. Furthermore, selenoprotein deficiency in T cells led to an inability of these cells to suppress reactive oxygen species (ROS) production, which in turn affected their ability to proliferate in response to T cell receptor stimulation. Selenoprotein-less macrophages, on the other hand, manifested mostly normal inflammatory responses, but this deficiency resulted in an altered regulation in extracellular matrix-related gene expression and a diminished migration of macrophages in a protein gel matrix. These observations provided novel insights into the role of selenoproteins in immune function and tissue homeostasis. PMID:20576203

Lentivirus-ABCG1 instillation reduces lipid accumulation and improves lung compliance in GM-CSF knock-out mice

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

Malur, Anagha; Huizar, Isham; Wells, Greg

2011-11-18

Highlights: Black-Right-Pointing-Pointer Lentivirus-ABCG1 reduces lipid accumulation in lungs of GM-CSF knock-out mice. Black-Right-Pointing-Pointer Up-regulation of ABCG1 improves lung function. Black-Right-Pointing-Pointer Upregulation of ABCG1 improves surfactant metabolism. -- Abstract: We have shown decreased expression of the nuclear transcription factor, peroxisome proliferator-activated receptor-gamma (PPAR{gamma}) and the PPAR{gamma}-regulated ATP-binding cassette transporter G1 (ABCG1) in alveolar macrophages from patients with pulmonary alveolar proteinosis (PAP). PAP patients also exhibit neutralizing antibodies to granulocyte-macrophage colony stimulating factor (GM-CSF), an upregulator of PPAR{gamma}. In association with functional GM-CSF deficiency, PAP lung is characterized by surfactant-filled alveolar spaces and lipid-filled alveolar macrophages. Similar pathology characterizes GM-CSF knock-out (KO)more » mice. We reported previously that intratracheal instillation of a lentivirus (lenti)-PPAR{gamma} plasmid into GM-CSF KO animals elevated ABCG1 and reduced alveolar macrophage lipid accumulation. Here, we hypothesized that instillation of lenti-ABCG1 might be sufficient to decrease lipid accumulation and improve pulmonary function in GM-CSF KO mice. Animals received intratracheal instillation of lenti-ABCG1 or control lenti-enhanced Green Fluorescent Protein (eGFP) plasmids and alveolar macrophages were harvested 10 days later. Alveolar macrophage transduction efficiency was 79% as shown by lenti-eGFP fluorescence. Quantitative PCR analyses indicated a threefold (p = 0.0005) increase in ABCG1 expression with no change of PPAR{gamma} or ABCA1 in alveolar macrophages of lenti-ABCG1 treated mice. ABCG1 was unchanged in control lenti-eGFP and PBS-instilled groups. Oil Red O staining detected reduced intracellular neutral lipid in alveolar macrophages from lenti-ABCG1 treated mice. Extracellular cholesterol and phospholipids were also decreased as shown by analysis of bronchoalveolar lavage fluid. Lung compliance was diminished in untreated GMCSF KO mice but improved significantly after lenti-ABCG1 treatment. Data demonstrate that in vivo instillation of lenti-ABCG1 in GM-CSF KO mice is sufficient to restore pulmonary homeostasis by: (1) upregulating ABCG1; (2) reducing intra and extracellular lipids; and (3) improving lung function. Results suggest that the ABCG1 lipid transporter is the key downstream target of GM-CSF-induced PPAR{gamma} necessary for surfactant catabolism.« less

Macrophages of multiple sclerosis patients display deficient SHP-1 expression and enhanced inflammatory phenotype.

PubMed

Christophi, George P; Panos, Michael; Hudson, Chad A; Christophi, Rebecca L; Gruber, Ross C; Mersich, Akos T; Blystone, Scott D; Jubelt, Burk; Massa, Paul T

2009-07-01

Recent studies in mice have demonstrated that the protein tyrosine phosphatase SHP-1 is a crucial negative regulator of proinflammatory cytokine signaling, TLR signaling, and inflammatory gene expression. Furthermore, mice genetically lacking SHP-1 (me/me) display a profound susceptibility to inflammatory CNS demyelination relative to wild-type mice. In particular, SHP-1 deficiency may act predominantly in inflammatory macrophages to increase CNS demyelination as SHP-1-deficient macrophages display coexpression of inflammatory effector molecules and increased demyelinating activity in me/me mice. Recently, we reported that PBMCs of multiple sclerosis (MS) patients have a deficiency in SHP-1 expression relative to normal control subjects indicating that SHP-1 deficiency may play a similar role in MS as to that seen in mice. Therefore, it became essential to examine the specific expression and function of SHP-1 in macrophages from MS patients. Herein, we document that macrophages of MS patients have deficient SHP-1 protein and mRNA expression relative to those of normal control subjects. To examine functional consequences of the lower SHP-1, the activation of STAT6, STAT1, and NF-kappaB was quantified and macrophages of MS patients showed increased activation of these transcription factors. In accordance with this observation, several STAT6-, STAT1-, and NF-kappaB-responsive genes that mediate inflammatory demyelination were increased in macrophages of MS patients following cytokine and TLR agonist stimulation. Supporting a direct role of SHP-1 deficiency in altered macrophage function, experimental depletion of SHP-1 in normal subject macrophages resulted in an increased STAT/NF-kappaB activation and increased inflammatory gene expression to levels seen in macrophages of MS patients. In conclusion, macrophages of MS patients display a deficiency of SHP-1 expression, heightened activation of STAT6, STAT1, and NF-kappaB and a corresponding inflammatory profile that may be important in controlling macrophage-mediated demyelination in MS.

Osteomacs interact with megakaryocytes and osteoblasts to regulate murine hematopoietic stem cell function.

PubMed

Mohamad, Safa F; Xu, Linlin; Ghosh, Joydeep; Childress, Paul J; Abeysekera, Irushi; Himes, Evan R; Wu, Hao; Alvarez, Marta B; Davis, Korbin M; Aguilar-Perez, Alexandra; Hong, Jung Min; Bruzzaniti, Angela; Kacena, Melissa A; Srour, Edward F

2017-12-12

Networking between hematopoietic stem cells (HSCs) and cells of the hematopoietic niche is critical for stem cell function and maintenance of the stem cell pool. We characterized calvariae-resident osteomacs (OMs) and their interaction with megakaryocytes to sustain HSC function and identified distinguishing properties between OMs and bone marrow (BM)-derived macrophages. OMs, identified as CD45 + F4/80 + cells, were easily detectable (3%-5%) in neonatal calvarial cells. Coculture of neonatal calvarial cells with megakaryocytes for 7 days increased OM three- to sixfold, demonstrating that megakaryocytes regulate OM proliferation. OMs were required for the hematopoiesis-enhancing activity of osteoblasts, and this activity was augmented by megakaryocytes. Serial transplantation demonstrated that HSC repopulating potential was best maintained by in vitro cultures containing osteoblasts, OMs, and megakaryocytes. With or without megakaryocytes, BM-derived macrophages were unable to functionally substitute for neonatal calvarial cell-associated OMs. In addition, OMs differentiated into multinucleated, tartrate resistant acid phosphatase-positive osteoclasts capable of bone resorption. Nine-color flow cytometric analysis revealed that although BM-derived macrophages and OMs share many cell surface phenotypic similarities (CD45, F4/80, CD68, CD11b, Mac2, and Gr-1), only a subgroup of OMs coexpressed M-CSFR and CD166, thus providing a unique profile for OMs. CD169 was expressed by both OMs and BM-derived macrophages and therefore was not a distinguishing marker between these 2 cell types. These results demonstrate that OMs support HSC function and illustrate that megakaryocytes significantly augment the synergistic activity of osteoblasts and OMs. Furthermore, this report establishes for the first time that the crosstalk between OMs, osteoblasts, and megakaryocytes is a novel network supporting HSC function.

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

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

Li, Weina, E-mail: liweina228@163.com; He, Fei, E-mail: hesili1027@163.com

2014-07-18

Highlights: • Expression of MMD is increased in lung cancer tissues. • Knockdown of MMD inhibits growth of A549 and LLC cells in vitro and in vivo. • MMD is a direct functional target of miR-140-5p. • MiR-140-5p/MMD axis regulates Erk1/2 signaling. - Abstract: Monocyte to macrophage differentiation-associated (MMD) is identified in macrophages as a gene associated with the differentiation from monocytes to macrophages. Recent microarray analysis for non-small cell lung cancer (NSCLC) suggests that MMD is an important signature associated with relapse and survival among patients with NSCLC. Therefore, we speculate that MMD likely plays a role in lungmore » 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.« less

Macrophage Control of Phagocytosed Mycobacteria Is Increased by Factors Secreted by Alveolar Epithelial Cells through Nitric Oxide Independent Mechanisms

PubMed Central

Freidl, Raphaela; Fernández, Carmen

2014-01-01

Tissue-resident macrophages are heterogeneous with tissue-specific and niche-specific functions. Thus, simplified models of macrophage activation do not explain the extent of heterogeneity seen in vivo. We focus here on the respiratory tract and ask whether factors secreted by alveolar epithelial cells (AEC) can influence the functionality of resident pulmonary macrophages (PuM). We have previously reported that factors secreted by AEC increase control of intracellular growth of BCG in macrophages. In the current study, we also aimed to investigate possible mechanisms by which AEC-derived factors increase intracellular control of BCG in both primary murine interstitial macrophages, and bone marrow-derived macrophages and characterize further the effect of these factors on macrophage differentiation. We show that; a) in contrast to other macrophage types, IFN-γ did not increase intracellular growth control of Mycobacterium bovis, Bacillus Calmette-Guérin (BCG) by interstitial pulmonary macrophages although the same macrophages could be activated by factors secreted by AEC; b) the lack of response of pulmonary macrophages to IFN-γ was apparently regulated by suppressor of cytokine signaling (SOCS)1; c) AEC-derived factors did not induce pro-inflammatory pathways induced by IFN-γ e.g. expression of inducible nitric oxide synthase (iNOS), secretion of nitric oxide (NO), or IL-12, d) in contrast to IFN-γ, intracellular bacterial destruction induced by AEC-derived factors was not dependent on iNOS transcription and NO production. Collectively, our data show that PuM were restricted in inflammatory responses mediated by IFN-γ through SOCS1 and that factors secreted by AEC- enhanced the microbicidal capacities of macrophages by iNOS independent mechanisms. PMID:25089618

Macrophage heterogeneity in liver injury and fibrosis.

PubMed

Tacke, Frank; Zimmermann, Henning W

2014-05-01

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

A small molecule deubiquitinase inhibitor increases localization of inducible nitric oxide synthase to the macrophage phagosome and enhances bacterial killing.

PubMed

Burkholder, Kristin M; Perry, Jeffrey W; Wobus, Christiane E; Donato, Nicholas J; Showalter, Hollis D; Kapuria, Vaibhav; O'Riordan, Mary X D

2011-12-01

Macrophages are key mediators of antimicrobial defense and innate immunity. Innate intracellular defense mechanisms can be rapidly regulated at the posttranslational level by the coordinated addition and removal of ubiquitin by ubiquitin ligases and deubiquitinases (DUBs). While ubiquitin ligases have been extensively studied, the contribution of DUBs to macrophage innate immune function is incompletely defined. We therefore employed a small molecule DUB inhibitor, WP1130, to probe the role of DUBs in the macrophage response to bacterial infection. Treatment of activated bone marrow-derived macrophages (BMM) with WP1130 significantly augmented killing of the intracellular bacterial pathogen Listeria monocytogenes. WP1130 also induced killing of phagosome-restricted bacteria, implicating a bactericidal mechanism associated with the phagosome, such as the inducible nitric oxide synthase (iNOS). WP1130 had a minimal antimicrobial effect in macrophages lacking iNOS, indicating that iNOS is an effector mechanism for WP1130-mediated bacterial killing. Although overall iNOS levels were not notably different, we found that WP1130 significantly increased colocalization of iNOS with the Listeria-containing phagosome during infection. Taken together, our data indicate that the deubiquitinase inhibitor WP1130 increases bacterial killing in macrophages by enhancing iNOS localization to the phagosome and suggest a potential role for ubiquitin regulation in iNOS trafficking.

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

Umikawa, Masato, E-mail: umikawa@med.u-ryukyu.ac.jp; Umikawa, Asako; Asato, Tsuyoshi

Monocytes and macrophages are important effectors and regulators of inflammation, and both their differentiation and activation are regulated strictly in response to environmental cues. Angiopoietin-like protein 2 (Angptl2) is a multifaceted protein, displaying many physiological and pathological functions in inflammation, angiogenesis, hematopoiesis, and tumor development. Although recent studies implicate Angptl2 in chronic inflammation, the mechanisms of inflammation caused by Angptl2 remain unclear. The purpose of the present study was to elucidate the role of Angptl2 in inflammation by understanding the effects of Angptl2 on monocytes/macrophages. We showed that Angptl2 directly activates resident murine peritoneal monocytes and macrophages and induces amore » drastic upregulation of the transcription of several inflammatory genes including nitric oxide synthase 2 and prostaglandin-endoperoxide synthase 2, and several proinflammatory cytokine genes such as interleukin (IL)-1β, IL-6, TNFα, and CSF2, along with activation of ERK, JNK, p38, and nuclear factor kappa B signaling pathways. Concordantly, proinflammatory cytokines IL-1β, IL-6, TNFα, and GM-CSF, were rapidly elevated from murine peritoneal monocytes and macrophages. These results demonstrate a novel role for Angptl2 in inflammation via the direct activation of peritoneal monocytes and macrophages. - Highlights: • Angptl2 directly activates resident murine peritoneal monocytes and macrophages. • Angptl2 induces a drastic upregulation of expression of inflammatory genes. • Angptl2 induces activation of ERK, JNK, p38, and nuclear factor kappa B signaling pathways. • Angptl2 does not activate bone marrow derived macrophages or macrophage cell lines.« less

Regulation of the macrophage oxytocin receptor in response to inflammation

PubMed Central

Szeto, Angela; Sun-Suslow, Ni; Mendez, Armando J.; Hernandez, Rosa I.; Wagner, Klaus V.

2017-01-01

It has been demonstrated that the neuropeptide oxytocin (OT) attenuates oxidative stress and inflammation in macrophages. In the current study, we examined the role of inflammation on the expression of the oxytocin receptor (OXTR). We hypothesized that OXTR expression is increased during the inflammation through a nuclear factor-κB (NF-κB)-mediated pathway, thus responding as an acute-phase protein. Inflammation was induced by treating macrophages (human primary, THP-1, and murine) with lipopolysaccharide (LPS) and monitored by expression of IL-6. Expression of OXTR and vasopressin receptors was assessed by qPCR, and OXTR expression was confirmed by immunoblotting. Inflammation upregulated OXTR transcription 10- to 250-fold relative to control in THP-1 and human primary macrophages and increased OXTR protein expression. In contrast, vasopressin receptor-2 mRNA expression was reduced following LPS treatment. Blocking NF-κB activation prevented the increase in OXTR transcription. OT treatment of control cells and LPS-treated cells increased ERK1/2 phosphorylation, demonstrating activation of the OXTR/Gαq/11 signaling pathway. OT activation of OXTR reduced secretion of IL-6 in LPS-activated macrophages. Collectively, these findings suggest that OXTR is an acute-phase protein and that its increased expression is regulated by NF-κB and functions to attenuate cellular inflammatory responses in macrophages. PMID:28049625

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

PubMed

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.

Development and maintenance of the brain's immune toolkit: Microglia and non-parenchymal brain macrophages.

PubMed

Lopez-Atalaya, Jose P; Askew, Katharine E; Sierra, Amanda; Gomez-Nicola, Diego

2018-06-01

Microglia and non-parenchymal macrophages located in the perivascular space, the meninges and the choroid plexus are independent immune populations that play vital roles in brain development, homeostasis, and tissue healing. Resident macrophages account for a significant proportion of cells in the brain and their density remains stable throughout the lifespan thanks to constant turnover. Microglia develop from yolk sac progenitors, later evolving through intermediate progenitors in a fine-tuned process in which intrinsic factors and external stimuli combine to progressively sculpt their cell type-specific transcriptional profiles. Recent evidence demonstrates that non-parenchymal macrophages are also generated during early embryonic development. In recent years, the development of powerful fate mapping approaches combined with novel genomic and transcriptomic methodologies have greatly expanded our understanding of how brain macrophages develop and acquire specialized functions, and how cell population dynamics are regulated. Here, we review the transcription factors, epigenetic remodeling, and signaling pathways orchestrating the embryonic development of microglia and non-parenchymal macrophages. Next, we describe the dynamics of the macrophage populations of the brain and discuss the role of progenitor cells, to gain a better understanding of their functions in the healthy and diseased brain. © 2017 Wiley Periodicals, Inc. Develop Neurobiol 78: 561-579, 2018. © 2017 The Authors Developmental Neurobiology Published by Wiley Periodicals, Inc.

Functional relevance of protein glycosylation to the pro-inflammatory effects of extracellular matrix metalloproteinase inducer (EMMPRIN) on monocytes/macrophages.

PubMed

Ge, Heng; Yuan, Wei; Liu, Jidong; He, Qing; Ding, Song; Pu, Jun; He, Ben

2015-01-01

Extracellular matrix metalloproteinase inducer (EMMPRIN) is an important pro-inflammatory protein involved in the cellular functions of monocytes/macrophages. We have hypothesized that high-level heterogeneousness of protein glycosylation of EMMPRIN may have functional relevance to its biological effects and affect the inflammatory activity of monocytes/macrophages. The glycosylation patterns of EMMPRIN expressed by monocytes/macrophages (THP-1 cells) in response to different extracellular stimuli were observed, and the structures of different glycosylation forms were identified. After the purification of highly- and less-glycosylated proteins respectively, the impacts of different glycosylation forms on the pro-inflammatory effects of EMMPRIN were examined in various aspects, such as cell adhesion to endothelial cells, cell migrations, cytokine expression, and activation of inflammatory signalling pathway. 1) It was mainly the highly-glycosylated form of EMMPRIN (HG-EMMPRIN) that increased after being exposed to inflammatory signals (PMA and H2O2). 2) Glycosylation of EMMPRIN in monocytes/macrophages led to N-linked-glycans being added to the protein, with the HG form containing complex-type glycans and the less-glycosylated form (LG) the simple type. 3) Only the HG-EMMPRIN but not the LG-EMMPRIN exhibited pro-inflammatory effects and stimulated inflammatory activities of the monocytes/macrophages (i.e., activation of ERK1/2 and NF-κB pathway, enhanced monocyte-endothelium adhesion, cell migration and matrix metalloproteinase -9 expression). Post-transcriptional glycosylation represents an important mechanism that determines the biological effects of EMMPRIN in monocytes/macrophages. Glycosylation of EMMPRIN may serve as a potential target for regulating the inflammatory activities of monocytes/macrophages.

The role of HFE genotype in macrophage phenotype.

PubMed

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

2018-02-01

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

The phosphoproteome of toll-like receptor-activated macrophages

PubMed Central

Weintz, Gabriele; Olsen, Jesper V; Frühauf, Katja; Niedzielska, Magdalena; Amit, Ido; Jantsch, Jonathan; Mages, Jörg; Frech, Cornelie; Dölken, Lars; Mann, Matthias; Lang, Roland

2010-01-01

Recognition of microbial danger signals by toll-like receptors (TLR) causes re-programming of macrophages. To investigate kinase cascades triggered by the TLR4 ligand lipopolysaccharide (LPS) on systems level, we performed a global, quantitative and kinetic analysis of the phosphoproteome of primary macrophages using stable isotope labelling with amino acids in cell culture, phosphopeptide enrichment and high-resolution mass spectrometry. In parallel, nascent RNA was profiled to link transcription factor (TF) phosphorylation to TLR4-induced transcriptional activation. We reproducibly identified 1850 phosphoproteins with 6956 phosphorylation sites, two thirds of which were not reported earlier. LPS caused major dynamic changes in the phosphoproteome (24% up-regulation and 9% down-regulation). Functional bioinformatic analyses confirmed canonical players of the TLR pathway and highlighted other signalling modules (e.g. mTOR, ATM/ATR kinases) and the cytoskeleton as hotspots of LPS-regulated phosphorylation. Finally, weaving together phosphoproteome and nascent transcriptome data by in silico promoter analysis, we implicated several phosphorylated TFs in primary LPS-controlled gene expression. PMID:20531401

Low Necroptosis Process Predicts Poor Treatment Outcome of Human Papillomavirus Positive Cervical Cancers by Decreasing Tumor-Associated Macrophages M1 Polarization.

PubMed

Li, Lin; Yu, Song; Zang, Chunyi

2018-01-01

The aim of this study was to assess the functions of the necroptosis process on the prognosis of high-risk human papillomavirus (HR-HPV)-related cervical cancer. PCR and western blotting were used to demonstrate the expression of the necroptosis marker, mixed lineage kinase domain-like protein (MLKL), in whole blood and peripheral blood mononuclears (PBMCs) of 89 cervical cancer patients and 15 healthy volunteers. Necroptosis levels and M1 polarization were determined in tumor co-cultured macrophages. We found that MLKL expressions were significantly increased in cervical cancer patients in both whole blood and PBMC samples compared to the expressions in the healthy controls. Low MLKL expression was significantly associated with decreased survival rate in overall survival and disease-free survival. Co-culture cervical cancer cells decrease the necroptosis process of macrophage, together with the proinflammatory factors (M1 markers) downregulation, and this negative regulation was exacerbated in HPV-positive cases. Necroptosis enhancer RIPK3 overexpression showed reversed regulation of these M1 markers, suggesting that co-culture cervical cancer cells decrease the macrophage M1 polarization partly through necroptosis downregulation. Our study revealed that necroptosis process could be a relevant marker for the determination of the prognosis in cervical cancer patients, which might be because of its role in regulating macrophage polarization. © 2018 S. Karger AG, Basel.

miR-155 Deletion in Mice Overcomes Neuron-Intrinsic and Neuron-Extrinsic Barriers to Spinal Cord Repair.

PubMed

Gaudet, Andrew D; Mandrekar-Colucci, Shweta; Hall, Jodie C E; Sweet, David R; Schmitt, Philipp J; Xu, Xinyang; Guan, Zhen; Mo, Xiaokui; Guerau-de-Arellano, Mireia; Popovich, Phillip G

2016-08-10

Axon regeneration after spinal cord injury (SCI) fails due to neuron-intrinsic mechanisms and extracellular barriers including inflammation. microRNA (miR)-155-5p is a small, noncoding RNA that negatively regulates mRNA translation. In macrophages, miR-155-5p is induced by inflammatory stimuli and elicits a response that could be toxic after SCI. miR-155 may also independently alter expression of genes that regulate axon growth in neurons. Here, we hypothesized that miR-155 deletion would simultaneously improve axon growth and reduce neuroinflammation after SCI by acting on both neurons and macrophages. New data show that miR-155 deletion attenuates inflammatory signaling in macrophages, reduces macrophage-mediated neuron toxicity, and increases macrophage-elicited axon growth by ∼40% relative to control conditions. In addition, miR-155 deletion increases spontaneous axon growth from neurons; adult miR-155 KO dorsal root ganglion (DRG) neurons extend 44% longer neurites than WT neurons. In vivo, miR-155 deletion augments conditioning lesion-induced intraneuronal expression of SPRR1A, a regeneration-associated gene; ∼50% more injured KO DRG neurons expressed SPRR1A versus WT neurons. After dorsal column SCI, miR-155 KO mouse spinal cord has reduced neuroinflammation and increased peripheral conditioning-lesion-enhanced axon regeneration beyond the epicenter. Finally, in a model of spinal contusion injury, miR-155 deletion improves locomotor function at postinjury times corresponding with the arrival and maximal appearance of activated intraspinal macrophages. In miR-155 KO mice, improved locomotor function is associated with smaller contusion lesions and decreased accumulation of inflammatory macrophages. Collectively, these data indicate that miR-155 is a novel therapeutic target capable of simultaneously overcoming neuron-intrinsic and neuron-extrinsic barriers to repair after SCI. Axon regeneration after spinal cord injury (SCI) fails due to neuron-intrinsic mechanisms and extracellular barriers, including inflammation. Here, new data show that deleting microRNA-155 (miR-155) affects both mechanisms and improves repair and functional recovery after SCI. Macrophages lacking miR-155 have altered inflammatory capacity, which enhances neuron survival and axon growth of cocultured neurons. In addition, independent of macrophages, adult miR-155 KO neurons show enhanced spontaneous axon growth. Using either spinal cord dorsal column crush or contusion injury models, miR-155 deletion improves indices of repair and recovery. Therefore, miR-155 has a dual role in regulating spinal cord repair and may be a novel therapeutic target for SCI and other CNS pathologies. Copyright © 2016 the authors 0270-6474/16/368516-17$15.00/0.

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.

PubMed

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 proline/serine-rich putative SH3 binding domain.

Recent Advances in Obesity-Induced Inflammation and Insulin Resistance

PubMed Central

Tateya, Sanshiro; Kim, Francis; Tamori, Yoshikazu

2013-01-01

It has been demonstrated in rodents and humans that chronic inflammation characterized by macrophage infiltration occurs mainly in adipose tissue or liver during obesity, in which activation of immune cells is closely associated with insulin sensitivity. Macrophages can be classified as classically activated (M1) macrophages that support microbicidal activity or alternatively activated (M2) macrophages that support allergic and antiparasitic responses. In the context of insulin action, M2 macrophages sustain insulin sensitivity by secreting IL-4 and IL-10, while M1 macrophages induce insulin resistance through the secretion of proinflammatory cytokines, such as TNFα. Polarization of M1/M2 is controlled by various dynamic functions of other immune cells. It has been demonstrated that, in a lean state, TH2 cells, Treg cells, natural killer T cells, or eosinophils contribute to the M2 activation of macrophages by secreting IL-4 or IL-10. In contrast, obesity causes alteration of the constituent immune cells, in which TH1 cells, B cells, neutrophils, or mast cells induce M1 activation of macrophages by the elevated secretion of TNFα and IFNγ. Increased secretion of TNFα and free fatty acids from hypertrophied adipocytes also contributes to the M1 activation of macrophages. Since obesity-induced insulin resistance is established by macrophage infiltration and the activation of immune cells inside tissues, identification of the factors that regulate accumulation and the intracellular signaling cascades that define polarization of M1/M2 would be indispensable. Regulation of these factors would lead to the pharmacological inhibition of obesity-induced insulin resistance. In this review, we introduce molecular mechanisms relevant to the pathophysiology and review the most recent studies of clinical applications targeting chronic inflammation. PMID:23964268

Modulating macrophage polarization with divalent cations in nanostructured titanium implant surfaces

NASA Astrophysics Data System (ADS)

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

2016-02-01

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

Cytoprotective function of heme oxygenase 1 induced by a nitrated cyclic nucleotide formed during murine salmonellosis.

PubMed

Zaki, Mohammad Hasan; Fujii, Shigemoto; Okamoto, Tatsuya; Islam, Sabrina; Khan, Shahzada; Ahmed, Khandaker Ahtesham; Sawa, Tomohiro; Akaike, Takaaki

2009-03-15

Signaling mechanisms of NO-mediated host defense are yet to be elucidated. In this study, we report a unique signal pathway for cytoprotection during Salmonella infection that involves heme oxygenase 1 (HO-1) induced by a nitrated cyclic nucleotide, 8-nitroguanosine 3',5'-cyclic monophosphate (8-nitro-cGMP). Wild-type C57BL/6 mice and C57BL/6 mice lacking inducible NO synthase (iNOS) were infected with Salmonella enterica serovar Typhimurium LT2. HO-1 was markedly up-regulated during the infection, the level being significantly higher in wild-type mice than in iNOS-deficient mice. HO-1 up-regulation was associated with 8-nitro-cGMP formation detected immunohistochemically in Salmonella-infected mouse liver and peritoneal macrophages. 8-Nitro-cGMP either exogenously added or formed endogenously induced HO-1 in cultured macrophages infected with Salmonella. HO-1 inhibition by polyethylene glycol-conjugated zinc-protoporphyrin IX impaired intracellular killing of bacteria in mouse liver and in both RAW 264 cells and peritoneal macrophages. Infection-associated apoptosis was also markedly increased in polyethylene glycol-conjugated zinc-protoporphyrin IX-treated mouse liver cells and cultured macrophages. This effect of HO-1 inhibition was further confirmed by using HO-1 short interfering RNA in peritoneal macrophages. Our results suggest that HO-1 induced by NO-mediated 8-nitro-cGMP formation contributes, via its potent cytoprotective function, to host defense during murine salmonellosis.

Inhibition of nuclear factor of activated T cells (NFAT) c3 activation attenuates acute lung injury and pulmonary edema in murine models of sepsis

PubMed Central

Karpurapu, Manjula; Lee, Yong Gyu; Qian, Ziqing; Wen, Jin; Ballinger, Megan N.; Rusu, Luiza; Chung, Sangwoon; Deng, Jing; Qian, Feng; Reader, Brenda F.; Nirujogi, Teja Srinivas; Park, Gye Young; Pei, Dehua; Christman, John W.

2018-01-01

Specific therapies targeting cellular and molecular events of sepsis induced Acute Lung Injury (ALI) pathogenesis are lacking. We have reported a pivotal role for Nuclear Factors of Activated T cells (NFATc3) in regulating macrophage phenotype during sepsis induced ALI and subsequent studies demonstrate that NFATc3 transcriptionally regulates macrophage CCR2 and TNFα gene expression. Mouse pulmonary microvascular endothelial cell monolayer maintained a tighter barrier function when co-cultured with LPS stimulated NFATc3 deficient macrophages whereas wild type macrophages caused leaky monolayer barrier. More importantly, NFATc3 deficient mice showed decreased neutrophilic lung inflammation, improved alveolar capillary barrier function, arterial oxygen saturation and survival benefit in lethal CLP sepsis mouse models. In addition, survival of wild type mice subjected to the lethal CLP sepsis was not improved with broad-spectrum antibiotics, whereas the survival of NFATc3 deficient mice was improved to 40–60% when treated with imipenem. Passive adoptive transfer of NFATc3 deficient macrophages conferred protection against LPS induced ALI in wild type mice. Furthermore, CP9-ZIZIT, a highly potent, cell-permeable peptide inhibitor of Calcineurin inhibited NFATc3 activation. CP9-ZIZIT effectively reduced sepsis induced inflammatory cytokines and pulmonary edema in mice. Thus, this study demonstrates that inhibition of NFATc3 activation by CP9-ZIZIT provides a potential therapeutic option for attenuating sepsis induced ALI/pulmonary edema. PMID:29535830

Reticulon 4B (Nogo-B) is necessary for macrophage infiltration and tissue repair.

PubMed

Yu, Jun; Fernández-Hernando, Carlos; Suarez, Yajaira; Schleicher, Michael; Hao, Zhengrong; Wright, Paulette L; DiLorenzo, Annarita; Kyriakides, Themis R; Sessa, William C

2009-10-13

Blood vessel formation during ischemia and wound healing requires coordination of the inflammatory response with genes that regulate blood vessel assembly. Here we show that the reticulon family member 4B, aka Nogo-B, is upregulated in response to ischemia and is necessary for blood flow recovery secondary to ischemia and wound healing. Mice lacking Nogo-B exhibit reduced arteriogenesis and angiogenesis that are linked to a decrease in macrophage infiltration and inflammatory gene expression in vivo. Bone marrow-derived macrophages isolated from Nogo knock-out mice have reduced spreading and chemotaxis due to impaired Rac activation. Bone marrow reconstitution experiments show that Nogo in myeloid cells is necessary to promote macrophage homing and functional recovery after limb ischemia. Thus, endogenous Nogo coordinates macrophage-mediated inflammation with arteriogenesis, wound healing, and blood flow control.

Proteomic Analysis of ABCA1-Null Macrophages Reveals a Role for Stomatin-Like Protein-2 in Raft Composition and Toll-Like Receptor Signaling.

PubMed

Chowdhury, Saiful M; Zhu, Xuewei; Aloor, Jim J; Azzam, Kathleen M; Gabor, Kristin A; Ge, William; Addo, Kezia A; Tomer, Kenneth B; Parks, John S; Fessler, Michael B

2015-07-01

Lipid raft membrane microdomains organize signaling by many prototypical receptors, including the Toll-like receptors (TLRs) of the innate immune system. Raft-localization of proteins is widely thought to be regulated by raft cholesterol levels, but this is largely on the basis of studies that have manipulated cell cholesterol using crude and poorly specific chemical tools, such as β-cyclodextrins. To date, there has been no proteome-scale investigation of whether endogenous regulators of intracellular cholesterol trafficking, such as the ATP binding cassette (ABC)A1 lipid efflux transporter, regulate targeting of proteins to rafts. Abca1(-/-) macrophages have cholesterol-laden rafts that have been reported to contain increased levels of select proteins, including TLR4, the lipopolysaccharide receptor. Here, using quantitative proteomic profiling, we identified 383 proteins in raft isolates from Abca1(+/+) and Abca1(-/-) macrophages. ABCA1 deletion induced wide-ranging changes to the raft proteome. Remarkably, many of these changes were similar to those seen in Abca1(+/+) macrophages after lipopolysaccharide exposure. Stomatin-like protein (SLP)-2, a member of the stomatin-prohibitin-flotillin-HflK/C family of membrane scaffolding proteins, was robustly and specifically increased in Abca1(-/-) rafts. Pursuing SLP-2 function, we found that rafts of SLP-2-silenced macrophages had markedly abnormal composition. SLP-2 silencing did not compromise ABCA1-dependent cholesterol efflux but reduced macrophage responsiveness to multiple TLR ligands. This was associated with reduced raft levels of the TLR co-receptor, CD14, and defective lipopolysaccharide-induced recruitment of the common TLR adaptor, MyD88, to rafts. Taken together, we show that the lipid transporter ABCA1 regulates the protein repertoire of rafts and identify SLP-2 as an ABCA1-dependent regulator of raft composition and of the innate immune response. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Proteomic Analysis of ABCA1-Null Macrophages Reveals a Role for Stomatin-Like Protein-2 in Raft Composition and Toll-Like Receptor Signaling*

PubMed Central

Chowdhury, Saiful M.; Zhu, Xuewei; Aloor, Jim J.; Azzam, Kathleen M.; Gabor, Kristin A.; Ge, William; Addo, Kezia A.; Tomer, Kenneth B.; Parks, John S.; Fessler, Michael B.

2015-01-01

Lipid raft membrane microdomains organize signaling by many prototypical receptors, including the Toll-like receptors (TLRs) of the innate immune system. Raft-localization of proteins is widely thought to be regulated by raft cholesterol levels, but this is largely on the basis of studies that have manipulated cell cholesterol using crude and poorly specific chemical tools, such as β-cyclodextrins. To date, there has been no proteome-scale investigation of whether endogenous regulators of intracellular cholesterol trafficking, such as the ATP binding cassette (ABC)A1 lipid efflux transporter, regulate targeting of proteins to rafts. Abca1−/− macrophages have cholesterol-laden rafts that have been reported to contain increased levels of select proteins, including TLR4, the lipopolysaccharide receptor. Here, using quantitative proteomic profiling, we identified 383 proteins in raft isolates from Abca1+/+ and Abca1−/− macrophages. ABCA1 deletion induced wide-ranging changes to the raft proteome. Remarkably, many of these changes were similar to those seen in Abca1+/+ macrophages after lipopolysaccharide exposure. Stomatin-like protein (SLP)-2, a member of the stomatin-prohibitin-flotillin-HflK/C family of membrane scaffolding proteins, was robustly and specifically increased in Abca1−/− rafts. Pursuing SLP-2 function, we found that rafts of SLP-2-silenced macrophages had markedly abnormal composition. SLP-2 silencing did not compromise ABCA1-dependent cholesterol efflux but reduced macrophage responsiveness to multiple TLR ligands. This was associated with reduced raft levels of the TLR co-receptor, CD14, and defective lipopolysaccharide-induced recruitment of the common TLR adaptor, MyD88, to rafts. Taken together, we show that the lipid transporter ABCA1 regulates the protein repertoire of rafts and identify SLP-2 as an ABCA1-dependent regulator of raft composition and of the innate immune response. PMID:25910759

The function of cancer-shed gangliosides in macrophage phenotype: involvement with angiogenesis.

PubMed

Chung, Tae-Wook; Choi, Hee-Jung; Park, Mi-Ju; Choi, Hee-Jin; Lee, Syng-Ook; Kim, Keuk-Jun; Kim, Cheorl-Ho; Hong, Changwan; Kim, Kyun-Ha; Joo, Myungsoo; Ha, Ki-Tae

2017-01-17

Tumor-derived gangliosides in the tumor microenvironment are involved in the malignant progression of cancer. However, the molecular mechanisms underlying the effects of gangliosides shed from tumors on macrophage phenotype remain unknown. Here, we showed that ganglioside GM1 highly induced the activity and expression of arginase-1 (Arg-1), a major M2 macrophage marker, compared to various gangliosides in bone marrow-derived macrophages (BMDM), peritoneal macrophages and Raw264.7 macrophage cells. We found that GM1 bound to macrophage mannose receptor (MMR/CD206) and common gamma chain (γc). In addition, GM1 increased Arg-1 expression through CD206 and γc-mediated activation of Janus kinase 3 (JAK3) and signal transducer and activator of transcription- 6 (STAT-6). Interestingly, GM1-stimulated macrophages secreted monocyte chemoattractant protein-1 (MCP-1/CCL2) through a CD206/γc/STAT6-mediated signaling pathway and induced angiogenesis. Moreover, the angiogenic effect of GM1-treated macrophages was diminished by RS102895, an MCP-1 receptor (CCR2) antagonist. From these results we suggest that tumor-shed ganglioside is a secretory factor regulating the phenotype of macrophages and consequently enhancing angiogenesis.

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

PubMed Central

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

2011-01-01

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

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

PubMed

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

2011-12-01

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

Synergistic activation of Arg1 gene by retinoic acid and IL-4 involves chromatin remodeling for transcription initiation and elongation coupling

PubMed Central

Lee, Bomi; Wu, Cheng-Ying; Lin, Yi-Wei; Park, Sung Wook; Wei, Li-Na

2016-01-01

All-trans Retinoic acid (RA) and its derivatives are potent therapeutics for immunological functions including wound repair. However, the molecular mechanism of RA modulation in innate immunity is poorly understood, especially in macrophages. We found that topical application of RA significantly improves wound healing and that RA and IL-4 synergistically activate Arg1, a critical gene for tissue repair, in M2 polarized macrophages. This involves feed forward regulation of Raldh2, a rate-limiting enzyme for RA biosynthesis, and requires Med25 to coordinate RAR, STAT6 and chromatin remodeler, Brg1 to remodel the +1 nucleosome of Arg1 for transcription initiation. By recruiting elongation factor TFIIS, Med25 also facilitates transcriptional initiation-elongation coupling. This study uncovers synergistic activation of Arg1 by RA and IL-4 in M2 macrophages that involves feed forward regulation of RA synthesis and dual functions of Med25 in nucleosome remodeling and transcription initiation-elongation coupling that underlies robust modulatory activity of RA in innate immunity. PMID:27166374

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

PubMed

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.

Macrophage-expressed perforins mpeg1 and mpeg1.2 have an anti-bacterial function in zebrafish.

PubMed

Benard, Erica L; Racz, Peter I; Rougeot, Julien; Nezhinsky, Alexander E; Verbeek, Fons J; Spaink, Herman P; Meijer, Annemarie H

2015-01-01

Macrophage-expressed gene 1 (MPEG1) encodes an evolutionarily conserved protein with a predicted membrane attack complex/perforin domain associated with host defence against invading pathogens. In vertebrates, MPEG1/perforin-2 is an integral membrane protein of macrophages, suspected to be involved in the killing of intracellular bacteria by pore-forming activity. Zebrafish have 3 copies of MPEG1; 2 are expressed in macrophages, whereas the third could be a pseudogene. The mpeg1 and mpeg1.2 genes show differential regulation during infection of zebrafish embryos with the bacterial pathogens Mycobacterium marinum and Salmonella typhimurium. While mpeg1 is downregulated during infection with both pathogens, mpeg1.2 is infection inducible. Upregulation of mpeg1.2 is partially dependent on the presence of functional Mpeg1 and requires the Toll-like receptor adaptor molecule MyD88 and the transcription factor NFκB. Knockdown of mpeg1 alters the immune response to M. marinum infection and results in an increased bacterial burden. In Salmonella typhimurium infection, both mpeg1 and mpeg1.2 knockdown increase the bacterial burdens, but mpeg1 morphants show increased survival times. The combined results of these two in vivo infection models support the anti-bacterial function of the MPEG1/perforin-2 family and indicate that the intricate cross-regulation of the two mpeg1 copies aids the zebrafish host in combatting infection of various pathogens. © 2014 S. Karger AG, Basel.

Functional evidence for the inflammatory reflex in teleosts: A novel α7 nicotinic acetylcholine receptor modulates the macrophage response to dsRNA.

PubMed

Torrealba, Débora; Balasch, Joan Carles; Criado, Manuel; Tort, Lluís; Mackenzie, Simon; Roher, Nerea

2018-07-01

The inflammatory reflex modulates the innate immune system, keeping in check the detrimental consequences of overstimulation. A key player controlling the inflammatory reflex is the alpha 7 acetylcholine receptor (α7nAChR). This receptor is one of the signalling molecules regulating cytokine expression in macrophages. In this study, we characterize a novel teleost α7nAChR. Protein sequence analysis shows a high degree of conservation with mammalian orthologs and trout α7nAChR has all the features and essential amino acids to form a fully functional receptor. We demonstrate that trout macrophages can bind α-bungarotoxin (α-BTX), a competitive antagonist for α7nAChRs. Moreover, nicotine stimulation produces a decrease in pro-inflammatory cytokine expression after stimulation with poly(I:C). These results suggest the presence of a functional α7nAChR in the macrophage plasma membrane. Further, in vivo injection of poly(I:C) induced an increase in serum ACh levels in rainbow trout. Our results manifest for the first time the functional conservation of the inflammatory reflex in teleosts. Copyright © 2018 Elsevier Ltd. All rights reserved.

Suppression of CYP1 members of the AHR response by pathogen-associated molecular patterns.

PubMed

Peres, Adam G; Zamboni, Robert; King, Irah L; Madrenas, Joaquín

2017-12-01

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that triggers a broad response, which includes the regulation of proinflammatory cytokine production by monocytes and macrophages. AHR is negatively regulated by a set of genes that it transcriptionally activates, including the AHR repressor ( Ahrr ) and the cytochrome P450 1 ( Cyp1 ) family, which are critical for preventing exacerbated AHR activity. An imbalance in these regulatory mechanisms has been shown to cause severe defects in lymphoid cells. Therefore, we wanted to assess how AHR activation is regulated in monocytes and macrophages in the context of innate immune responses induced by pathogen-associated molecular patterns (PAMPs). We found that concomitant stimulation of primary human monocytes with PAMPs and the AHR agonist 6-formylindolo(3,2-b)carbazole (FICZ) led to a selective dose-dependent inhibition of Cyp1 family members induction. Two other AHR-dependent genes [ Ahrr and NADPH quinone dehydrogenase 1 ( Nqo1 )] were not affected under these conditions, suggesting a split in the AHR regulation by PAMPs. This down-regulation of Cyp1 family members did not require de novo protein production nor signaling through p38, ERK, or PI3K-Akt-mammalian target of rapamycin (mTOR) pathways. Furthermore, such a split regulation of the AHR response was more apparent in GM-CSF-derived macrophages, a finding corroborated at the functional level by decreased CYP1 activity and decreased proinflammatory cytokine production in response to FICZ and LPS. Collectively, our findings identify a role for pattern recognition receptor (PRR) signaling in regulating the AHR response through selective down-regulation of Cyp1 expression in human monocytes and macrophages. © Society for Leukocyte Biology.

VIP impairs acquisition of the macrophage proinflammatory polarization profile.

PubMed

Carrión, Mar; Pérez-García, Selene; Martínez, Carmen; Juarranz, Yasmina; Estrada-Capetillo, Lizbeth; Puig-Kröger, Amaya; Gomariz, Rosa P; Gutiérrez-Cañas, Irene

2016-12-01

This study tested the hypothesis that vasoactive intestinal peptide (VIP) is able to modify the macrophage inflammatory profile, thus supporting its therapeutic role in autoimmune diseases. Macrophages are innate immune cells that display a variety of functions and inflammatory profiles in response to the environment that critically controls their polarization. Deregulation between the pro- and anti-inflammatory phenotypes has been involved in different pathologies. Rheumatoid arthritis (RA) is an autoimmune disease, in which macrophages are considered central effectors of synovial inflammation, displaying a proinflammatory profile. VIP is a pleiotropic neuropeptide with proven anti-inflammatory actions. As modulation of the macrophage phenotype has been implicated in the resolution of inflammatory diseases, we evaluated whether VIP is able to modulate human macrophage polarization. In vitro-polarized macrophages by GM-CSF (GM-MØ), with a proinflammatory profile, expressed higher levels of VIP receptors, vasoactive intestinal polypeptide receptors 1 and 2 (VPAC1 and VPAC2, respectively), than macrophages polarized by M-CSF (M-MØ) with anti-inflammatory activities. RA synovial macrophages, according to their GM-CSF-like polarization state, expressed both VPAC1 and VPAC2. In vitro-generated GM-MØ exposed to VIP exhibited an up-regulation of M-MØ gene marker expression, whereas their proinflammatory cytokine profile was reduced in favor of an anti-inflammatory function. Likewise, in GM-MØ, generated in the presence of VIP, VIP somehow changes the macrophages physiology profile to a less-damaging phenotype. Therefore, these results add new value to VIP as an immunomodulatory agent on inflammatory diseases. © Society for Leukocyte Biology.

Anti-apoptotic Bcl-XL but not Mcl-1 contributes to protection against virus-induced apoptosis.

PubMed

Ohmer, Michaela; Weber, Arnim; Sutter, Gerd; Ehrhardt, Katrin; Zimmermann, Albert; Häcker, Georg

2016-08-18

Infection of mammalian cells with viruses often induces apoptosis. How the recognition of viruses leads to apoptosis of the infected cell and which host cell factors regulate this cell death is incompletely understood. In this study, we focussed on two major anti-apoptotic proteins of the host cell, whose abundance and activity are important for cell survival, the Bcl-2-like proteins Mcl-1 and Bcl-XL. During infection of epithelial cells and fibroblasts with modified vaccinia virus Ankara (MVA), Mcl-1 protein levels dropped but the MVA Bcl-2-like protein F1L could replace Mcl-1 functionally; a similar activity was found in vaccinia virus (VACV)-infected cells. During infection with murine cytomegalovirus (MCMV), Mcl-1-levels were not reduced but a viral Mcl-1-like activity was also generated. Infection of mouse macrophages with any of these viruses, on the other hand, induced apoptosis. Virus-induced macrophage apoptosis was unaltered in the absence of Mcl-1. However, apoptosis was substantially increased in infected Bcl-XL-deficient macrophages or macrophages treated with the Bcl-2/Bcl-XL-inhibitor ABT-737. Genetic loss of Bcl-XL or treatment of macrophages with ABT-737 reduced the generation of infectious VACV. These data show that Mcl-1 is dispensable for the regulation of apoptosis during infection with different large DNA viruses, either because the viruses replace its function (in fibroblasts and epithelial cells) or because the pro-apoptotic activity generated by the infection appears not to be blocked by it (in macrophages). Bcl-XL, on the other hand, can be important to maintain survival of virus-infected cells, and its activity can determine outcome of the infection.

Anti-apoptotic Bcl-XL but not Mcl-1 contributes to protection against virus-induced apoptosis

PubMed Central

Ohmer, Michaela; Weber, Arnim; Sutter, Gerd; Ehrhardt, Katrin; Zimmermann, Albert; Häcker, Georg

2016-01-01

Infection of mammalian cells with viruses often induces apoptosis. How the recognition of viruses leads to apoptosis of the infected cell and which host cell factors regulate this cell death is incompletely understood. In this study, we focussed on two major anti-apoptotic proteins of the host cell, whose abundance and activity are important for cell survival, the Bcl-2-like proteins Mcl-1 and Bcl-XL. During infection of epithelial cells and fibroblasts with modified vaccinia virus Ankara (MVA), Mcl-1 protein levels dropped but the MVA Bcl-2-like protein F1L could replace Mcl-1 functionally; a similar activity was found in vaccinia virus (VACV)-infected cells. During infection with murine cytomegalovirus (MCMV), Mcl-1-levels were not reduced but a viral Mcl-1-like activity was also generated. Infection of mouse macrophages with any of these viruses, on the other hand, induced apoptosis. Virus-induced macrophage apoptosis was unaltered in the absence of Mcl-1. However, apoptosis was substantially increased in infected Bcl-XL-deficient macrophages or macrophages treated with the Bcl-2/Bcl-XL-inhibitor ABT-737. Genetic loss of Bcl-XL or treatment of macrophages with ABT-737 reduced the generation of infectious VACV. These data show that Mcl-1 is dispensable for the regulation of apoptosis during infection with different large DNA viruses, either because the viruses replace its function (in fibroblasts and epithelial cells) or because the pro-apoptotic activity generated by the infection appears not to be blocked by it (in macrophages). Bcl-XL, on the other hand, can be important to maintain survival of virus-infected cells, and its activity can determine outcome of the infection. PMID:27537523

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

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

Li, Bin; Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208; Abdalrahman, Akram

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 promisesmore » 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 of Nrf2 independently of Keap1 and NF-κB, suggesting a unique therapeutic potential of dh404 for specific targeting a Nrf2-mediated resolution of inflammation.« less

Conditioned medium from persistently RSV-infected macrophages alters transcriptional profile and inflammatory response of non-infected macrophages.

PubMed

Rivera-Toledo, Evelyn; Salido-Guadarrama, Iván; Rodríguez-Dorantes, Mauricio; Torres-González, Laura; Santiago-Olivares, Carlos; Gómez, Beatriz

2017-02-15

Cells susceptible to persistent viral infections undergo important changes in their biological functions as a consequence of the expression of viral gene products that are capable of altering the gene expression profile of the host cell. Previously, we reported that persistence of the RSV genome in a mouse macrophage cell line induces important alterations in cell homeostasis, including constitutive expression of IFN-β and other pro-inflammatory cytokines. Here, we postulated that changes in the homeostasis of non-infected macrophages could be induced by soluble factors secreted by persistently RSV- infected macrophages. To test this hypothesis, non-infected mouse macrophages were treated with conditioned medium (CM) collected from cultures of persistently RSV-infected macrophages. Total RNA was extracted and a microarray-based gene expression analysis was performed. Non-infected macrophages, treated under similar conditions with CM obtained from cultures of non-infected macrophages, were used as a control to establish differential gene expression between the two conditions. Results showed that CM from the persistently RSV-infected cultures altered expression of a total of 95 genes in non-infected macrophages, resulting in an antiviral gene-transcription profile along with inhibition of the inflammatory response, since some inflammatory genes were down-regulated, including Nlrp3 and Il-1 β, both related to the inflammasome pathway. However, down-regulation of Nlrp3 and Il-1 β was reversible upon acute RSV infection. Additionally, we observed that the inflammatory response, evaluated by secreted IL-1 β, a final product of the inflammasome activity, was enhanced during acute RSV infection in macrophages treated with CM from persistently RSV-infected cultures, compared to that in macrophages treated with the control CM. This suggests that soluble factors secreted during RSV persistence may induce an exacerbated inflammatory response in non-infected cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Carboxyl-functionalized polyurethane nanoparticles with immunosuppressive properties as a new type of anti-inflammatory platform

NASA Astrophysics Data System (ADS)

Huang, Yen-Jang; Hung, Kun-Che; Hsieh, Fu-Yu; Hsu, Shan-Hui

2015-12-01

The interaction of nanoparticles (NPs) with the body immune system is critically important for their biomedical applications. Most NPs stimulate the immune response of macrophages. Here we show that synthetic polyurethane nanoparticles (PU NPs, diameter 34-64 nm) with rich surface COO- functional groups (zeta potential -70 to -50 mV) can suppress the immune response of macrophages. The specially-designed PU NPs reduce the gene expression levels of proinflammatory cytokines (IL-1β, IL-6, and TNF-α) for endotoxin-treated macrophages. The PU NPs increase the intracellular calcium of macrophages (4.5-6.5 fold) and activate autophagy. This is in contrast to the autophagy dysfunction generally observed upon NP exposure. These PU NPs may further decrease the nuclear factor-κB-related inflammation via autophagy pathways. The immunosuppressive activities of PU NPs can prevent animal death by inhibiting the macrophage recruitment and proinflammatory responses, confirmed by an in vivo zebrafish model. Therefore, the novel biodegradable PU NPs demonstrate COO- dependent immunosuppressive properties without carrying any anti-inflammatory agents. This study suggests that NP surface chemistry may regulate the immune response, which provides a new paradigm for potential applications of NPs in anti-inflammation and immunomodulation.The interaction of nanoparticles (NPs) with the body immune system is critically important for their biomedical applications. Most NPs stimulate the immune response of macrophages. Here we show that synthetic polyurethane nanoparticles (PU NPs, diameter 34-64 nm) with rich surface COO- functional groups (zeta potential -70 to -50 mV) can suppress the immune response of macrophages. The specially-designed PU NPs reduce the gene expression levels of proinflammatory cytokines (IL-1β, IL-6, and TNF-α) for endotoxin-treated macrophages. The PU NPs increase the intracellular calcium of macrophages (4.5-6.5 fold) and activate autophagy. This is in contrast to the autophagy dysfunction generally observed upon NP exposure. These PU NPs may further decrease the nuclear factor-κB-related inflammation via autophagy pathways. The immunosuppressive activities of PU NPs can prevent animal death by inhibiting the macrophage recruitment and proinflammatory responses, confirmed by an in vivo zebrafish model. Therefore, the novel biodegradable PU NPs demonstrate COO- dependent immunosuppressive properties without carrying any anti-inflammatory agents. This study suggests that NP surface chemistry may regulate the immune response, which provides a new paradigm for potential applications of NPs in anti-inflammation and immunomodulation. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr06379e

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

PubMed

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 selective hair cell lesion. Because these afferent neurons carry sound information from the cochlea to the auditory brainstem, their survival is a key determinant of the success of cochlear prosthetics. Our data suggest that fractalkine signaling in the cochlea is neuroprotective, and reveal a previously uncharacterized interaction between cells of the cochlea and the innate immune system. Copyright © 2015 the authors 0270-6474/15/3515050-12$15.00/0.

Macrophages: Their Emerging Roles in Bone

PubMed Central

Sinder, Benjamin P; Pettit, Allison R; McCauley, Laurie K

2016-01-01

Macrophages are present in nearly all tissues and are critical for development, homeostasis, and regeneration. Resident tissue macrophages of bone, termed osteal macrophages, are recently classified myeloid cells that are distinct from osteoclasts. Osteal macrophages are located immediately adjacent to osteoblasts, regulate bone formation, and play diverse roles in skeletal homeostasis. Genetic or pharmacological modulation of macrophages in vivo results in significant bone phenotypes, and these phenotypes depend on which macrophage subsets are altered. Macrophages are also key mediators of osseous wound healing and fracture repair, with distinct roles at various stages of the repair process. A central function of macrophages is their phagocytic ability. Each day, billions of cells die in the body and efferocytosis (phagocytosis of apoptotic cells) is a critical process in both clearing dead cells and recruitment of replacement progenitor cells to maintain homeostasis. Recent data suggest a role for efferocytosis in bone biology and these new mechanisms are outlined. Finally, although macrophages have an established role in primary tumors, emerging evidence suggests that macrophages in bone support cancers which preferentially metastasize to the skeleton. Collectively, this developing area of osteoimmunology raises new questions and promises to provide novel insights into pathophysiologic conditions as well as therapeutic and regenerative approaches vital for skeletal health. PMID:26531055

The suppression of inflammatory macrophage-mediated cytotoxicity and proinflammatory cytokine production by transgenic expression of HLA-E.

PubMed

Maeda, Akira; Kawamura, Takuji; Ueno, Takehisa; Usui, Noriaki; Eguchi, Hiroshi; Miyagawa, Shuji

2013-12-01

Macrophages participate in xenogenic rejection and represent a major biological obstacle to successful xenotransplantation. The signal inhibitory regulatory protein α (SIRPα) receptor was reported to be a negative regulator of macrophage phagocytic activity via interaction with CD47, its ligand. Because a majority of human macrophages express the inhibitory receptor CD94/NKG2A, which binds specifically to the human leukocyte antigen (HLA)-E and contains immunoreceptor tyrosine-based inhibition motifs (ITIMs), the inhibitory function of HLA class I molecules, HLA-E, on macrophage-mediated cytolysis was examined. The suppressive effect against proinflammatory cytokine production by macrophages was also examined. Complementary DNA (cDNA) of HLA-E, and CD47 were prepared and transfected into swine endothelial cells (SEC). The expression of the modified genes was evaluated by flow cytometry and macrophage-mediated cytolysis was assessed using in vitro generated macrophages. Transgenic expression of HLA-E significantly suppressed the macrophage-mediated cytotoxicity. HLA-E transgenic expression demonstrated a significant suppression equivalent to CD47 transgenic expression. Furthermore, transgenic HLA-E suppressed the production of pro-inflammatory cytokines by inflammatory macrophages. These results indicate that generating transgenic HLA-E pigs might protect porcine grafts from, not only NK cytotoxicity, but also macrophage-mediated cytotoxicity. © 2013 Elsevier B.V. All rights reserved.

Upregulation of Monocyte/Macrophage HGFIN (Gpnmb/Osteoactivin) Expression in End-Stage Renal Disease

PubMed Central

Vaziri, Nosratola D.; Yuan, Jun; Adler, Sharon G.

2010-01-01

Background and objectives: Hematopoietic growth factor–inducible neurokinin 1 (HGFIN), also known as Gpnmb and osteoactivin, is a transmembrane glycoprotein that is expressed in numerous cells, including osteoclasts, macrophages, and dendritic cells. It serves as an osteoblast differentiation factor, participates in bone mineralization, and functions as a negative regulator of inflammation in macrophages. Although measurable at low levels in monocytes, monocyte-to-macrophage transformation causes substantial increase in HGFIN expression. HGFIN is involved in systemic inflammation, bone demineralization, and soft tissue vascular calcification. Design, setting, participants, & measurements: We explored HGFIN expression in monocytes and monocyte-derived macrophages in 21 stable hemodialysis patients and 22 control subjects. Results: Dialysis patients exhibited marked upregulation of colony-stimulating factor and IL-6 and significant downregulation of IL-10 in intact monocytes and transformed macrophages. HGFIN expression in intact monocytes was negligible in control subjects but conspicuously elevated (8.6-fold) in dialysis patients. As expected, in vitro monocyte-to-macrophage transformation resulted in marked upregulation of HGFIN in cells obtained from both groups but much more so in dialysis patients (17.5-fold higher). Upregulation of HGFIN and inflammatory cytokines in the uremic monocyte-derived macrophages occurred when grown in the presence of either normal or uremic serum, suggesting the enduring effect of the in vivo uremic milieu on monocyte/macrophage phenotype and function. Conclusions: Uremic macrophages exhibit increased HGFIN gene and protein expression and heightened expression of proinflammatory and a suppressed expression of anti-inflammatory cytokines. Further studies are needed to determine the role of heightened monocyte/macrophage HGFIN expression in the pathogenesis of ESRD-induced inflammation and vascular and soft tissue calcification. PMID:19833906

Oxygen Tension Modulates Differentiation and Primary Macrophage Functions in the Human Monocytic THP-1 Cell Line

PubMed Central

Grodzki, Ana Cristina G.; Giulivi, Cecilia; Lein, Pamela J.

2013-01-01

The human THP-1 cell line is widely used as an in vitro model system for studying macrophage differentiation and function. Conventional culture conditions for these cells consist of ambient oxygen pressure (∼20% v/v) and medium supplemented with the thiol 2-mercaptoethanol (2-ME) and serum. In consideration of the redox activities of O2 and 2-ME, and the extensive experimental evidence supporting a role for reactive oxygen species (ROS) in the differentiation and function of macrophages, we addressed the question of whether culturing THP-1 cells under a more physiologically relevant oxygen tension (5% O2) in the absence of 2-ME and serum would alter THP-1 cell physiology. Comparisons of cultures maintained in 18% O2 versus 5% O2 indicated that reducing oxygen tension had no effect on the proliferation of undifferentiated THP-1 cells. However, decreasing the oxygen tension to 5% O2 significantly increased the rate of phorbol ester-induced differentiation of THP-1 cells into macrophage-like cells as well as the metabolic activity of both undifferentiated and PMA-differentiated THP-1 cells. Removal of both 2-ME and serum from the medium decreased the proliferation of undifferentiated THP-1 cells but increased metabolic activity and the rate of differentiation under either oxygen tension. In differentiated THP-1 cells, lowering the oxygen tension to 5% O2 decreased phagocytic activity, the constitutive release of β-hexosaminidase and LPS-induced NF-κB activation but enhanced LPS-stimulated release of cytokines. Collectively, these data demonstrate that oxygen tension influences THP-1 cell differentiation and primary macrophage functions, and suggest that culturing these cells under tightly regulated oxygen tension in the absence of exogenous reducing agent and serum is likely to provide a physiologically relevant baseline from which to study the role of the local redox environment in regulating THP-1 cell physiology. PMID:23355903

Functional Relevance of Protein Glycosylation to the Pro-Inflammatory Effects of Extracellular Matrix Metalloproteinase Inducer (EMMPRIN) on Monocytes/Macrophages

PubMed Central

Ge, Heng; Yuan, Wei; Liu, Jidong; He, Qing; Ding, Song; Pu, Jun; He, Ben

2015-01-01

Background and Objective Extracellular matrix metalloproteinase inducer (EMMPRIN) is an important pro-inflammatory protein involved in the cellular functions of monocytes/macrophages. We have hypothesized that high-level heterogeneousness of protein glycosylation of EMMPRIN may have functional relevance to its biological effects and affect the inflammatory activity of monocytes/macrophages. Methods The glycosylation patterns of EMMPRIN expressed by monocytes/macrophages (THP-1 cells) in response to different extracellular stimuli were observed, and the structures of different glycosylation forms were identified. After the purification of highly- and less-glycosylated proteins respectively, the impacts of different glycosylation forms on the pro-inflammatory effects of EMMPRIN were examined in various aspects, such as cell adhesion to endothelial cells, cell migrations, cytokine expression, and activation of inflammatory signalling pathway. Results 1) It was mainly the highly-glycosylated form of EMMPRIN (HG-EMMPRIN) that increased after being exposed to inflammatory signals (PMA and H2O2). 2) Glycosylation of EMMPRIN in monocytes/macrophages led to N-linked-glycans being added to the protein, with the HG form containing complex-type glycans and the less-glycosylated form (LG) the simple type. 3) Only the HG-EMMPRIN but not the LG-EMMPRIN exhibited pro-inflammatory effects and stimulated inflammatory activities of the monocytes/macrophages (i.e., activation of ERK1/2 and NF-κB pathway, enhanced monocyte-endothelium adhesion, cell migration and matrix metalloproteinase -9 expression). Conclusions Post-transcriptional glycosylation represents an important mechanism that determines the biological effects of EMMPRIN in monocytes/macrophages. Glycosylation of EMMPRIN may serve as a potential target for regulating the inflammatory activities of monocytes/macrophages. PMID:25658763

Cot/tpl2 participates in the activation of macrophages by adiponectin.

PubMed

Sanz-Garcia, Carlos; Nagy, Laura E; Lasunción, Miguel A; Fernandez, Margarita; Alemany, Susana

2014-06-01

Whereas the main function of APN is to enhance insulin activity, it is also involved in modulating the macrophage phenotype. Here, we demonstrate that at physiological concentrations, APN activates Erk1/2 via the IKKβ-p105/NF-κΒ1-Cot/tpl2 intracellular signal transduction cassette in macrophages. In peritoneal macrophages stimulated with APN, Cot/tpl2 influences the ability to phagocytose beads. However, Cot/tpl2 did not modulate the known capacity of APN to decrease lipid content in peritoneal macrophages in response to treatment with oxLDL or acLDL. A microarray analysis of gene-expression profiles in BMDMs exposed to APN revealed that APN modulated the expression of ∼3300 genes; the most significantly affected biological functions were the inflammatory and the infectious disease responses. qRT-PCR analysis of WT and Cot/tpl2 KO macrophages stimulated with APN for 0, 3, and 18 h revealed that Cot/tpl2 participated in the up-regulation of APN target inflammatory mediators included in the cytokine-cytokine receptor interaction pathway (KEGG ID 4060). In accordance with these data, macrophages stimulated with APN increased secretion of cytokines and chemokines, including IL-1β, IL-1α, TNF-α, IL-10, IL-12, IL-6, and CCL2. Moreover, Cot/tpl2 also played an important role in the production of these inflammatory mediators upon stimulation of macrophages with APN. It has been reported that different types of signals that stimulate TLRs, IL-1R, TNFR, FcγR, and proteinase-activated receptor-1 activate Cot/tpl2. Here, we demonstrate that APN is a new signal that activates the IKKβ-p105/NF-κΒ1-Cot/tpl2-MKK1/2-Erk1/2 axis in macrophages. Furthermore, this signaling cassette modulates the biological functions triggered by APN in macrophages. © 2014 Society for Leukocyte Biology.

Cot/tpl2 participates in the activation of macrophages by adiponectin

PubMed Central

Sanz-Garcia, Carlos; Nagy, Laura E.; Lasunción, Miguel A.; Fernandez, Margarita; Alemany, Susana

2014-01-01

Whereas the main function of APN is to enhance insulin activity, it is also involved in modulating the macrophage phenotype. Here, we demonstrate that at physiological concentrations, APN activates Erk1/2 via the IKKβ-p105/NF-κΒ1-Cot/tpl2 intracellular signal transduction cassette in macrophages. In peritoneal macrophages stimulated with APN, Cot/tpl2 influences the ability to phagocytose beads. However, Cot/tpl2 did not modulate the known capacity of APN to decrease lipid content in peritoneal macrophages in response to treatment with oxLDL or acLDL. A microarray analysis of gene-expression profiles in BMDMs exposed to APN revealed that APN modulated the expression of ∼3300 genes; the most significantly affected biological functions were the inflammatory and the infectious disease responses. qRT-PCR analysis of WT and Cot/tpl2 KO macrophages stimulated with APN for 0, 3, and 18 h revealed that Cot/tpl2 participated in the up-regulation of APN target inflammatory mediators included in the cytokine–cytokine receptor interaction pathway (KEGG ID 4060). In accordance with these data, macrophages stimulated with APN increased secretion of cytokines and chemokines, including IL-1β, IL-1α, TNF-α, IL-10, IL-12, IL-6, and CCL2. Moreover, Cot/tpl2 also played an important role in the production of these inflammatory mediators upon stimulation of macrophages with APN. It has been reported that different types of signals that stimulate TLRs, IL-1R, TNFR, FcγR, and proteinase-activated receptor-1 activate Cot/tpl2. Here, we demonstrate that APN is a new signal that activates the IKKβ-p105/NF-κΒ1-Cot/tpl2-MKK1/2-Erk1/2 axis in macrophages. Furthermore, this signaling cassette modulates the biological functions triggered by APN in macrophages. PMID:24532642

METEORIN-LIKE is a cytokine associated with barrier tissues and alternatively activated macrophages

PubMed Central

Ushach, Irina; Burkhardt, Amanda M.; Martinez, Cynthia; Hevezi, Peter A.; Gerber, Peter Arne; Buhren, Bettina Alexandra; Schrumpf, Holger; Valle-Rios, Ricardo; Vazquez, Monica I.; Homey, Bernhard; Zlotnik, Albert

2014-01-01

Cytokines are involved in many functions of the immune system including initiating, amplifying and resolving immune responses. Through bioinformatics analyses of a comprehensive database of gene expression (BIGE: Body Index of Gene Expression) we observed that a small secreted protein encoded by a poorly characterized gene called meteorin-like (METRNL), is highly expressed in mucosal tissues, skin and activated macrophages. Further studies indicate that Metrnl is produced by Alternatively Activated Macrophages (AAM) and M-CSF cultured bone marrow macrophages (M2-like macrophages). In the skin, METRNL is expressed by resting fibroblasts and IFNγ-treated keratinocytes. A screen of human skin-associated diseases showed significant over-expression of METRNL in psoriasis, prurigo nodularis, actinic keratosis and atopic dermatitis. METRNL is also up-regulated in synovial membranes of human rheumatoid arthritis. Taken together, these results indicate that Metrnl represents a novel cytokine, which is likely involved in both innate and acquired immune responses. PMID:25486603

Macrophages are necessary for epimorphic regeneration in African spiny mice.

PubMed

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.

Macrophages are necessary for epimorphic regeneration in African spiny mice

PubMed Central

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

2017-01-01

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. DOI: http://dx.doi.org/10.7554/eLife.24623.001 PMID:28508748

Expression analysis of G Protein-Coupled Receptors in mouse macrophages

PubMed Central

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

2008-01-01

Background 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). Results 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. Conclusion 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. PMID:18442421

Cytoskeletal protein transformation in HIV-1-infected macrophage giant cells.

PubMed

Kadiu, Irena; Ricardo-Dukelow, Mary; Ciborowski, Pawel; Gendelman, Howard E

2007-05-15

The mechanisms linking HIV-1 replication, macrophage biology, and multinucleated giant cell formation are incompletely understood. With the advent of functional proteomics, the characterization, regulation, and transformation of HIV-1-infected macrophage-secreted proteins can be ascertained. To these ends, we performed proteomic analyses of culture fluids derived from HIV-1 infected monocyte-derived macrophages. Robust reorganization, phosphorylation, and exosomal secretion of the cytoskeletal proteins profilin 1 and actin were observed in conjunction with productive viral replication and giant cell formation. Actin and profilin 1 recruitment to the macrophage plasma membrane paralleled virus-induced cytopathicity, podosome formation, and cellular fusion. Poly-l-proline, an inhibitor of profilin 1-mediated actin polymerization, inhibited cytoskeletal transformations and suppressed, in part, progeny virion production. These data support the idea that actin and profilin 1 rearrangement along with exosomal secretion affect viral replication and cytopathicity. Such events favor the virus over the host cell and provide insights into macrophage defense mechanisms used to contain viral growth and how they may be affected during progressive HIV-1 infection.

LRH-1 mediates anti-inflammatory and antifungal phenotype of IL-13-activated macrophages through the PPARγ ligand synthesis

PubMed Central

Lefèvre, Lise; Authier, Hélène; Stein, Sokrates; Majorel, Clarisse; Couderc, Bettina; Dardenne, Christophe; Eddine, Mohamad Ala; Meunier, Etienne; Bernad, José; Valentin, Alexis; Pipy, Bernard; Schoonjans, Kristina; Coste, Agnès

2015-01-01

Liver receptor homologue-1 (LRH-1) is a nuclear receptor involved in the repression of inflammatory processes in the hepatointestinal tract. Here we report that LRH-1 is expressed in macrophages and induced by the Th2 cytokine IL-13 via a mechanism involving STAT6. We show that loss-of-function of LRH-1 in macrophages impedes IL-13-induced macrophage polarization due to impaired generation of 15-HETE PPARγ ligands. The incapacity to generate 15-HETE metabolites is at least partially caused by the compromised regulation of CYP1A1 and CYP1B1. Mice with LRH-1-deficient macrophages are, furthermore, highly susceptible to gastrointestinal and systemic Candida albicans infection. Altogether, these results identify LRH-1 as a critical component of the anti-inflammatory and fungicidal response of alternatively activated macrophages that acts upstream from the IL-13-induced 15-HETE/PPARγ axis. PMID:25873311

Sulforaphane regulates phenotypic and functional switching of both induced and spontaneously differentiating human monocytes.

PubMed

Pal, Sanjima; Konkimalla, V Badireenath

2016-06-01

At the site of inflammation, switching default on polarization of monocyte differentiation into classically activated macrophages (M1 type) is one of the pathogenic outcomes in several inflammatory autoimmune diseases, such as rheumatoid arthritis and osteoarthritis. In rheumatoid and osteoarthritis, a soluble collagen known as self-antigen is considered as a biomarker and acts as an important inflammatory mediator. In the present study, we investigated the effects of sulforaphane (SFN) on phenotypic changes and functional switching during in vitro induced and spontaneous differentiation of monocytes/macrophages, whose conditions were established with THP1 induced by PMA, and human peripheral blood monocytes, respectively. SFN at non-cytotoxic concentration (10μM) blocked soluble collagen induced inflammatory responses specific to M1 macrophages, COX-2, iNOS, surface CD14, CD197 expressions and production of IL12p70, suggesting that signals induced by SFN eventually shifted macrophage polarization to a direction specific to M2 macrophages (CD36high CD197extremely low). Results obtained with the induction of inflammatory conditions specific to M1 macrophages followed by SFN treatment showed that MAPKs were involved in the M1 to M2 phenotype switching. This immune-modulatory nature of SFN provides a clear indication for its ability to alleviate chronic inflammatory diseases by targeting monocytes/macrophages. Copyright © 2016 Elsevier B.V. All rights reserved.

Telomerase Activation in Atherosclerosis and Induction of Telomerase Reverse Transcriptase Expression by Inflammatory Stimuli in Macrophages

PubMed Central

Gizard, Florence; Heywood, Elizabeth B.; Findeisen, Hannes M.; Zhao, Yue; Jones, Karrie L.; Cudejko, Cèline; Post, Ginell R.; Staels, Bart; Bruemmer, Dennis

2010-01-01

Objective Telomerase serves as a critical regulator of tissue renewal. Although telomerase activity is inducible in response to various environmental cues, it remains unknown whether telomerase is activated during the inflammatory remodeling underlying atherosclerosis formation. To address this question, we investigated in the present study the regulation of telomerase in macrophages and during atherosclerosis development in LDL-receptor-deficient mice. Methods and Results We demonstrate that inflammatory stimuli activate telomerase in macrophages by inducing the expression of the catalytic subunit telomerase reverse transcriptase (TERT). Reporter and chromatin immunoprecipitation assays identified a previously unrecognized NF-κB response element in the TERT promoter, to which NF-κB is recruited during inflammation. Inhibition of NF-κB signaling completely abolished the induction of TERT expression, characterizing TERT as a bona fide NF-κB target gene. Furthermore, functional experiments revealed that TERT-deficiency results in a senescent cell phenotype. Finally, we demonstrate high levels of TERT expression in macrophages of human atherosclerotic lesions and establish that telomerase is activated during atherosclerosis development in LDL-receptor-deficient mice. Conclusion These results characterize TERT as a previously unrecognized NF-κB target gene in macrophages and demonstrate that telomerase is activated during atherosclerosis. This induction of TERT expression prevents macrophage senescence and may have important implications for the development of atherosclerosis. PMID:21106948

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

PubMed Central

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

Transcriptional Regulation of CXCL5 in HIV-1-Infected Macrophages and Its Functional Consequences on CNS Pathology.

PubMed

Guha, Debjani; Klamar, Cynthia R; Reinhart, Todd; Ayyavoo, Velpandi

2015-05-01

Human immunodeficiency virus-1 (HIV-1)-infected monocytes/macrophages and microglia release increased levels of proinflammatory cytokines and chemokines, including ELR+ (containing glutamic acid-leucine-arginine motif) chemokines. To investigate the role of HIV-1 infection on chemokine regulation, monocyte-derived macrophages (MDMs) from normal donors were infected with HIV-1 and the expression of chemokines and their downstream biological functions were evaluated. Among the tested chemokines, CXCL5 was upregulated significantly both at the mRNA and protein level in the HIV-1-infected MDMs compared with mock-infected cultures. Upregulation of CXCL5 in the HIV-1-infected MDMs is, in part, regulated by increased interleukin-1β (IL-1β) production and phosphorylation of ERK1/2. Functional analyses indicate that HIV-1-induced overexpression of CXCL5 has enhanced the ability to attract neutrophils, as observed by chemotaxis assay. However, exposure of NT2, SH-SY5Y cells, and primary neurons to HIV-1-infected MDM supernatants resulted in cell death that was not rescued by anti-CXCL5 antibody suggesting that CXCL5 does not have direct effect on neuronal death. Together, these results suggest that the increased level of CXCL5 in tissue compartments, including the central nervous system of HIV-1-infected individuals might alter the inflammatory response through the infiltration of neutrophils into tissue compartment, thus causing secondary effects on resident cells.

Prolonged Injury and Altered Lung Function after Ozone Inhalation in Mice with Chronic Lung Inflammation

PubMed Central

Groves, Angela M.; Gow, Andrew J.; Massa, Christopher B.; Laskin, Jeffrey D.

2012-01-01

Surfactant protein–D (Sftpd) is a pulmonary collectin important in down-regulating macrophage inflammatory responses. In these experiments, we analyzed the effects of chronic macrophage inflammation attributable to loss of Sftpd on the persistence of ozone-induced injury, macrophage activation, and altered functioning in the lung. Wild-type (Sftpd+/+) and Sftpd−/− mice (aged 8 wk) were exposed to air or ozone (0.8 parts per million, 3 h). Bronchoalveolar lavage (BAL) fluid and tissue were collected 72 hours later. In Sftpd−/− mice, but not Sftpd+/+ mice, increased BAL protein and nitrogen oxides were observed after ozone inhalation, indicating prolonged lung injury and oxidative stress. Increased numbers of macrophages were also present in BAL fluid and in histologic sections from Sftpd−/− mice. These cells were enlarged and foamy, suggesting that they were activated. This conclusion was supported by findings of increased BAL chemotactic activity, and increased expression of inducible nitric oxide synthase in lung macrophages. In both Sftpd+/+ and Sftpd−/− mice, inhalation of ozone was associated with functional alterations in the lung. Although these alterations were limited to central airway mechanics in Sftpd+/+ mice, both central airway and parenchymal mechanics were modified by ozone exposure in Sftpd−/− mice. The most notable changes were evident in resistance and elastance spectra and baseline lung function, and in lung responsiveness to changes in positive end-expiratory pressure. These data demonstrate that a loss of Sftpd is associated with prolonged lung injury, oxidative stress, and macrophage accumulation and activation in response to ozone, and with more extensive functional changes consistent with the loss of parenchymal integrity. PMID:22878412

Inhibition of Neddylation Represses Lipopolysaccharide-induced Proinflammatory Cytokine Production in Macrophage Cells

PubMed Central

Chang, Fang-Mei; Reyna, Sara M.; Granados, Jose C.; Wei, Sung-Jen; Innis-Whitehouse, Wendy; Maffi, Shivani K.; Rodriguez, Edward; Slaga, Thomas J.; Short, John D.

2012-01-01

Cullin-RING E3 ligases (CRLs) are a class of ubiquitin ligases that control the proteasomal degradation of numerous target proteins, including IκB, and the activity of these CRLs are positively regulated by conjugation of a Nedd8 polypeptide onto Cullin proteins in a process called neddylation. CRL-mediated degradation of IκB, which normally interacts with and retains NF-κB in the cytoplasm, permits nuclear translocation and transactivation of the NF-κB transcription factor. Neddylation occurs through a multistep enzymatic process involving Nedd8 activating enzymes, and recent studies have shown that the pharmacological agent, MLN4924, can potently inhibit Nedd8 activating enzymes, thereby preventing neddylation of Cullin proteins and preventing the degradation of CRL target proteins. In macrophages, regulation of NF-κB signaling functions as a primary pathway by which infectious agents such as lipopolysaccharides (LPSs) cause the up-regulation of proinflammatory cytokines. Here we have analyzed the effects of MLN4924, and compared the effects of MLN4924 with a known anti-inflammatory agent (dexamethasone), on certain proinflammatory cytokines (TNF-α and IL-6) and the NF-κB signaling pathway in LPS-stimulated macrophages. We also used siRNA to block neddylation to assess the role of this molecular process during LPS-induced cytokine responsiveness. Our results demonstrate that blocking neddylation, either pharmacologically or using siRNA, abrogates the increase in certain proinflammatory cytokines secreted from macrophages in response to LPS. In addition, we have shown that MLN4924 and dexamethasone inhibit LPS-induced cytokine up-regulation at the transcriptional level, albeit through different molecular mechanisms. Thus, neddylation represents a novel molecular process in macrophages that can be targeted to prevent and/or treat the LPS-induced up-regulation of proinflammatory cytokines and the disease processes associated with their up-regulation. PMID:22927439

CNS Macrophages Control Neurovascular Development via CD95L.

PubMed

Chen, Si; Tisch, Nathalie; Kegel, Marcel; Yerbes, Rosario; Hermann, Robert; Hudalla, Hannes; Zuliani, Cecilia; Gülcüler, Gülce Sila; Zwadlo, Klara; von Engelhardt, Jakob; Ruiz de Almodóvar, Carmen; Martin-Villalba, Ana

2017-05-16

The development of neurons and vessels shares striking anatomical and molecular features, and it is presumably orchestrated by an overlapping repertoire of extracellular signals. CNS macrophages have been implicated in various developmental functions, including the morphogenesis of neurons and vessels. However, whether CNS macrophages can coordinately influence neurovascular development and the identity of the signals involved therein is unclear. Here, we demonstrate that activity of the cell surface receptor CD95 regulates neuronal and vascular morphogenesis in the post-natal brain and retina. Furthermore, we identify CNS macrophages as the main source of CD95L, and macrophage-specific deletion thereof reduces both neurovascular complexity and synaptic activity in the brain. CD95L-induced neuronal and vascular growth is mediated through src-family kinase (SFK) and PI3K signaling. Together, our study highlights a coordinated neurovascular development instructed by CNS macrophage-derived CD95L, and it underlines the importance of macrophages for the establishment of the neurovascular network during CNS development. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Fas-Fas ligand interactions are essential for the binding to and killing of activated macrophages by gamma delta T cells.

PubMed

Dalton, Jane E; Howell, Gareth; Pearson, Jayne; Scott, Phillip; Carding, Simon R

2004-09-15

Gammadelta T cells have a direct role in resolving the host immune response to infection by eliminating populations of activated macrophages. Macrophage reactivity resides within the Vgamma1/Vdelta6.3 subset of gammadelta T cells, which have the ability to kill activated macrophages following infection with Listeria monocytogenes (Lm). However, it is not known how gammadelta T cell macrophage cytocidal activity is regulated, or what effector mechanisms gammadelta T cells use to kill activated macrophages. Using a macrophage-T cell coculture system in which peritoneal macrophages from naive or Lm-infected TCRdelta-/- mice were incubated with splenocytes from wild-type and Fas ligand (FasL)-deficient mice (gld), the ability of Vgamma1 T cells to bind macrophages was shown to be dependent upon Fas-FasL interactions. Combinations of anti-TCR and FasL Abs completely abolished binding to and killing of activated macrophages by Vgamma1 T cells. In addition, confocal microscopy showed that Fas and the TCR colocalized on Vgamma1 T cells at points of contact with macrophages. Collectively, these studies identify an accessory or coreceptor-like function for Fas-FasL that is essential for the interaction of Vgamma1 T cells with activated macrophages and their elimination during the resolution stage of pathogen-induced immune responses. Copyright 2004 The American Association of Immunologists, Inc.

Arnica montana Stimulates Extracellular Matrix Gene Expression in a Macrophage Cell Line Differentiated to Wound-Healing Phenotype.

PubMed

Marzotto, Marta; Bonafini, Clara; Olioso, Debora; Baruzzi, Anna; Bettinetti, Laura; Di Leva, Francesca; Galbiati, Elisabetta; Bellavite, Paolo

2016-01-01

Arnica montana (Arnica m.) is used for its purported anti-inflammatory and tissue healing actions after trauma, bruises, or tissue injuries, but its cellular and molecular mechanisms are largely unknown. This work tested Arnica m. effects on gene expression using an in vitro model of macrophages polarized towards a "wound-healing" phenotype. The monocyte-macrophage human THP-1 cell line was cultured and differentiated with phorbol-myristate acetate and Interleukin-4, then exposed for 24h to Arnica m. centesimal (c) dilutions 2c, 3c, 5c, 9c, 15c or Control. Total RNA was isolated and cDNA libraries were sequenced with a NextSeq500 sequencer. Genes with significantly positive (up-regulated) or negative (down-regulated) fold changes were defined as differentially expressed genes (DEGs). A total of 20 DEGs were identified in Arnica m. 2c treated cells. Of these, 7 genes were up-regulated and 13 were down-regulated. The most significantly up-regulated function concerned 4 genes with a conserved site of epidermal growth factor-like region (p<0.001) and three genes of proteinaceous extracellular matrix, including heparin sulphate proteoglycan 2 (HSPG2), fibrillin 2 (FBN2), and fibronectin (FN1) (p<0.01). Protein assay confirmed a statistically significant increase of fibronectin production (p<0.05). The down-regulated transcripts derived from mitochondrial genes coding for some components of electron transport chain. The same groups of genes were also regulated by increasing dilutions of Arnica m. (3c, 5c, 9c, 15c), although with a lower effect size. We further tested the healing potential of Arnica m. 2c in a scratch model of wound closure based on the motility of bone marrow-derived macrophages and found evidence of an accelerating effect on cell migration in this system. The results of this work, taken together, provide new insights into the action of Arnica m. in tissue healing and repair, and identify extracellular matrix regulation by macrophages as a therapeutic target.

Arnica montana Stimulates Extracellular Matrix Gene Expression in a Macrophage Cell Line Differentiated to Wound-Healing Phenotype

PubMed Central

Marzotto, Marta; Bonafini, Clara; Olioso, Debora; Baruzzi, Anna; Bettinetti, Laura; Di Leva, Francesca; Galbiati, Elisabetta; Bellavite, Paolo

2016-01-01

Arnica montana (Arnica m.) is used for its purported anti-inflammatory and tissue healing actions after trauma, bruises, or tissue injuries, but its cellular and molecular mechanisms are largely unknown. This work tested Arnica m. effects on gene expression using an in vitro model of macrophages polarized towards a “wound-healing” phenotype. The monocyte-macrophage human THP-1 cell line was cultured and differentiated with phorbol-myristate acetate and Interleukin-4, then exposed for 24h to Arnica m. centesimal (c) dilutions 2c, 3c, 5c, 9c, 15c or Control. Total RNA was isolated and cDNA libraries were sequenced with a NextSeq500 sequencer. Genes with significantly positive (up-regulated) or negative (down-regulated) fold changes were defined as differentially expressed genes (DEGs). A total of 20 DEGs were identified in Arnica m. 2c treated cells. Of these, 7 genes were up-regulated and 13 were down-regulated. The most significantly up-regulated function concerned 4 genes with a conserved site of epidermal growth factor-like region (p<0.001) and three genes of proteinaceous extracellular matrix, including heparin sulphate proteoglycan 2 (HSPG2), fibrillin 2 (FBN2), and fibronectin (FN1) (p<0.01). Protein assay confirmed a statistically significant increase of fibronectin production (p<0.05). The down-regulated transcripts derived from mitochondrial genes coding for some components of electron transport chain. The same groups of genes were also regulated by increasing dilutions of Arnica m. (3c, 5c, 9c, 15c), although with a lower effect size. We further tested the healing potential of Arnica m. 2c in a scratch model of wound closure based on the motility of bone marrow-derived macrophages and found evidence of an accelerating effect on cell migration in this system. The results of this work, taken together, provide new insights into the action of Arnica m. in tissue healing and repair, and identify extracellular matrix regulation by macrophages as a therapeutic target. PMID:27832158

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

NASA Technical Reports Server (NTRS)

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.

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

PubMed Central

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

1998-01-01

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

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

PubMed

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

2005-08-01

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

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

USDA-ARS?s Scientific Manuscript database

Two major functions of the intestinal epithelium are to act as a physical barrier and to regulate the movement of nutrients, ions and fluid. Nematode infection induces alterations in smooth and epithelial cell function, including increased fluid in the intestinal lumen, which are attributed to a ST...

Mycobacterium tuberculosis Infection Manipulates the Glycosylation Machinery and the N-Glycoproteome of Human Macrophages and Their Microparticles.

PubMed

Hare, Nathan J; Lee, Ling Y; Loke, Ian; Britton, Warwick J; Saunders, Bernadette M; Thaysen-Andersen, Morten

2017-01-06

Tuberculosis (TB) remains a prevalent and lethal infectious disease. The glycobiology associated with Mycobacterium tuberculosis infection of frontline alveolar macrophages is still unresolved. Herein, we investigated the regulation of protein N-glycosylation in human macrophages and their secreted microparticles (MPs) used for intercellular communication upon M. tb infection. LC-MS/MS-based proteomics and glycomics were performed to monitor the regulation of glycosylation enzymes and receptors and the N-glycome in in vitro-differentiated macrophages and in isolated MPs upon M. tb infection. Infection promoted a dramatic regulation of the macrophage proteome. Most notably, significant infection-dependent down-regulation (4-26 fold) of 11 lysosomal exoglycosidases, e.g., β-galactosidase, β-hexosaminidases and α-/β-mannosidases, was observed. Relative weak infection-driven transcriptional regulation of these exoglycosidases and a stronger augmentation of the extracellular hexosaminidase activity demonstrated that the lysosome-centric changes may originate predominantly from infection-induced secretion of the lysosomal content. The macrophages showed heterogeneous N-glycan profiles and displayed significant up-regulation of complex-type glycosylation and concomitant down-regulation of paucimannosylation upon infection. Complementary intact N-glycopeptide analysis supported a subcellular-specific manipulation of the glycosylation machinery and altered glycosylation patterns of lysosomal N-glycoproteins within infected macrophages. Interestingly, the corresponding macrophage-derived MPs displayed unique N-glycome and proteome signatures supporting a preferential packaging from plasma membranes. The MPs were devoid of infection-dependent N-glycosylation signatures, but interestingly displayed increased levels of the glyco-initiating oligosaccharyltransferase complex and associated α-glucosidases that correlated with increased formation, N-glycan precursor levels and N-glycan density of infected MPs. In conclusion, this system-wide study provides new insight into the host- and pathogen-driven N-glycoproteome manipulation of macrophages in TB.

Proteoglycan 4 regulates macrophage function without altering atherosclerotic lesion formation in a murine bone marrow-specific deletion model.

PubMed

Nahon, Joya E; Hoekstra, Menno; Havik, Stefan R; Van Santbrink, Peter J; Dallinga-Thie, Geesje M; Kuivenhoven, Jan-Albert; Geerling, Janine J; Van Eck, Miranda

2018-05-05

Proteoglycan 4 (Prg4) has a high structural similarity with the established atherosclerosis-modulating proteoglycan versican, but its role in atherogenesis is still unknown. Therefore, the impact of Prg4 deficiency on macrophage function in vitro and atherosclerosis susceptibility in vivo was investigated. The presence and localization of Prg4 was studied in atherosclerotic lesions. Furthermore, the effect of Prg4 deficiency on macrophage foam cell formation, cholesterol efflux and lipopolysaccharide (LPS) response was determined. Finally, susceptibility for atherosclerotic lesion formation was investigated in bone marrow-specific Prg4 knockout (KO) mice. Prg4 mRNA expression was induced 91-fold (p<0.001) in murine initial atherosclerotic lesions and Prg4 protein co-localized with human lesional macrophages. Murine Prg4 KO macrophages showed increased foam cell formation (+2.1-fold, p<0.01). In parallel, the expression of the cholesterol efflux genes ATP-binding cassette transporter A1 and scavenger receptor type B1 was lower (-35%, p<0.05;-40%, p<0.05) in Prg4 KO macrophages. This translated into an impaired cholesterol efflux to high-density lipoprotein (-13%, p<0.001) and apolipoprotein A1 (-8%, p<0.05). Furthermore, Prg4 KO macrophages showed an impaired LPS-induced rise in TNFα secretion as compared to wild-type controls (-31%, p<0.001), indicating a reduced inflammatory response. Combined, these pro- and anti-atherogenic effects did not translate into a significant difference in atherosclerotic lesion formation upon bone marrow-specific deletion of Prg4 in low-density lipoprotein receptor KO mice. Prg4 is present in macrophages in both murine and human atherosclerotic lesions and critically influences macrophage function, but deletion of Prg4 in bone marrow-derived cells does not affect atherosclerotic lesion development. Copyright © 2018 Elsevier B.V. All rights reserved.

A MicroRNA93-IRF9-IRG1-Itaconic Acid Pathway Modulates M2-like-Macrophage Polarization to Revascularize Ischemic Muscle

PubMed Central

Ganta, Vijay Chaitanya; Choi, Min Hyub; Kutateladze, Anna; Fox, Todd E.; Farber, Charles R.; Annex, Brian H.

2017-01-01

Background Currently no therapies exist for treating, and improving outcomes in patients with severe peripheral arterial disease (PAD). MicroRNA93 (miR93) has been shown to favorably modulate angiogenesis and reduce tissue loss in genetic PAD models. However, the cell specific function, downstream mechanisms or signaling involved in miR93 mediated ischemic muscle neovascularization is not clear. Macrophages were best known to modulate arteriogenic response in PAD and the extent of arteriogenic response induced by macrophages is dependent on greater M2 to M1-activation/polarization state. In the current study, we identified a novel mechanism by which miR93 regulates macrophage-polarization to promote angiogenesis and arteriogenesis to revascularize ischemic muscle in experimental-PAD. Methods In vitro (macrophages, endothelial cells, skeletal muscle cells under normal and hypoxia serum starvation (HSS) conditions) and in vivo experiments in preclinical-PAD models (unilateral femoral artery ligation and resection)) were conducted to examine the role of miR93-interferon regulatory factor-9 (IRF9)-immune responsive gene-1 (IRG1)-itaconic acid pathway in macrophage-polarization, angiogenesis, arteriogenesis and perfusion recovery. Results In vivo, compared to wild type (WT) controls, miR106b-93-25 cluster deficient mice (miR106b-93-25−/−) showed decreased angiogenesis and arteriogenesis correlating with increased M1-like-macrophages following experimental-PAD. Intra-muscular delivery of miR93 in miR106b-93-25−/− PAD mice increased angiogenesis, arteriogenesis, the extent of perfusion which correlated with more M2-like-macrophages in the proximal and distal hind-limb muscles. In vitro, miR93 promotes and sustains M2-like-polarization even under M1-like-polarizing conditions (HSS). Delivery of bone marrow derived macrophages from miR106b-93-25−/− to WT ischemic-muscle decreased angiogenesis, arteriogenesis and perfusion, while transfer of wild-type macrophages to miR106b-93-25−/− had the opposite effect. Systematic analysis of top-differentially upregulated genes from RNA-sequencing between miR106b-93-25−/− and WT ischemic-muscle showed that miR93 regulates IRG1 function to modulate itaconic acid production and macrophage-polarization. 3′UTR luciferase-assays performed to determine whether IRG1 is a direct target of miR93 revealed that IRG1 is not a miR93 target but IRF9 that can regulate IRG1-expression is a miR93 target. In vitro, increased expression of IRF9, IRG1 and itaconic acid treatment significantly decreased endothelial angiogenic potential. Conclusion We conclude that miR93 inhibits IRF9 to decrease IRG1-itaconic acid production to induce M2-like-polarization in ischemic muscle to enhance angiogenesis, arteriogenesis and perfusion recovery in experimental-PAD. PMID:28356443

A droplet-merging platform for comparative functional analysis of m1 and m2 macrophages in response to e. coli-induced stimuli.

PubMed

Hondroulis, Evangelia; Movila, Alexandru; Sabhachandani, Pooja; Sarkar, Saheli; Cohen, Noa; Kawai, Toshihisa; Konry, Tania

2017-03-01

Microfluidic droplets are used to isolate cell pairs and prevent crosstalk with neighboring cells, while permitting free motility and interaction within the confined space. Dynamic analysis of cellular heterogeneity in droplets has provided insights in various biological processes. Droplet manipulation methods such as fusion and fission make it possible to precisely regulate the localized environment of a cell in a droplet and deliver reagents as required. Droplet fusion strategies achieved by passive mechanisms preserve cell viability and are easier to fabricate and operate. Here, we present a simple and effective method for the co-encapsulation of polarized M1 and M2 macrophages with Escherichia coli (E. coli) by passive merging in an integrated droplet generation, merging, and docking platform. This approach facilitated live cell profiling of effector immune functions in situ and quantitative functional analysis of macrophage heterogeneity. Biotechnol. Bioeng. 2017;114: 705-709. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

A Molecular Analysis of the Induction of Class II Major Histocompatibility Antigen Expression on Murine Macrophages by Interferon-Gamma and Its Down-Regulation by Interferon-Alpha/Beta and Dexamethasone

DTIC Science & Technology

1989-11-09

Approved for public release , distribution unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT 15. SUBJECT TERMS 16. SECURITY CLASSIFICATION OF: 17...kilobasepairs KCl, potassium chloride LFA, lymphocyte function associated LPS, lipopolysaccharide LT, lymphotoxin LV, lentivirus J.LCi, microcurie ...process has also been studied extensively. IL 1 is released from macrophages following contact with helper T cells and one of its targets is the

Protein kinase cα regulates the expression of complement receptor Ig in human monocyte-derived macrophages.

PubMed

Ma, Yuefang; Usuwanthim, Kanchana; Munawara, Usma; Quach, Alex; Gorgani, Nick N; Abbott, Catherine A; Hii, Charles S; Ferrante, Antonio

2015-03-15

The complement receptor Ig (CRIg) is selectively expressed by macrophages. This receptor not only promotes the rapid phagocytosis of bacteria by macrophages but also has anti-inflammatory and immunosuppressive functions. Previous findings have suggested that protein kinase C (PKC) may be involved in the regulation of CRIg expression in human macrophages. We have now examined the role of PKCα in CRIg expression in human monocyte-derived macrophages (MDM). Macrophages nucleofected with plasmid containing short hairpin RNA against PKCα showed markedly reduced expression of PKCα, but normal PKCζ expression, by Western blotting analysis, and vice versa. PKCα-deficient MDM showed increased expression of CRIg mRNA and protein (both the long and short form), an increase in phagocytosis of complement-opsonized Candida albicans, and decreased production of TNF-α and IL-6. TNF-α caused a marked decrease in CRIg expression, and addition of anti-TNF mAb to the TNF-α-producing MDMs increased CRIg expression. PKCα-deficient macrophages also showed significantly less bacterial LPS-induced downregulation of CRIg. In contrast, cells deficient in PKCα showed decreased expression of CR type 3 (CR3) and decreased production of TNF-α and IL-6 in response to LPS. MDM developed under conditions that increased expression of CRIg over CR3 showed significantly reduced production of TNF-α in response to opsonized C. albicans. The findings indicate that PKCα promotes the downregulation of CRIg and upregulation of CR3 expression and TNF-α and IL-6 production, a mechanism that may promote inflammation. Copyright © 2015 by The American Association of Immunologists, Inc.

Netrin-1 regulates the inflammatory response of neutrophils and macrophages, and suppresses ischemic acute kidney injury by inhibiting COX-2 mediated PGE2 production

PubMed Central

Ranganathan, Punithavathi Vilapakkam; Jayakumar, Calpurnia; Mohamed, Riyaz; Dong, Zheng; Ramesh, Ganesan

2012-01-01

Netrin-1 regulates inflammation but the mechanism by which this occurs is unknown. Here we explore the role of netrin-1 in regulating the production of the prostanoid metabolite PGE2 from neutrophils in in vitro and in vivo disease models. Ischemia reperfusion in wild-type and RAG-1 knockout mice induced severe kidney injury that was associated with a large increase in neutrophil infiltration and COX-2 expression in the infiltrating leukocytes. Administration of netrin-1 suppressed COX-2 expression, PGE2 and thromboxane production, and neutrophil infiltration into the kidney. This was associated with reduced apoptosis, inflammatory cytokine and chemokine expression, and improved kidney function. Treatment with the PGE2 receptor EP4 agonist enhanced neutrophil infiltration and renal injury which was not inhibited by netrin-1. Consistent with in vivo data, both LPS and IFNγ-induced inflammatory cytokine production in macrophages and IL-17-induced IFNγ production in neutrophils were suppressed by netrin-1 in vitro by suppression of COX-2 expression. Moreover, netrin-1 regulates COX-2 expression at the transcriptional level through the regulation of NFκB activation. Thus, netrin-1 regulates the inflammatory response of neutrophils and macrophages through suppression of COX-2 mediated PGE2 production. This could be a potential drug for treating many inflammatory immune disorders. PMID:23447066

Innate cellular sources of interleukin-17A regulate macrophage accumulation in cigarette- smoke-induced lung inflammation in mice.

PubMed

Bozinovski, Steven; Seow, Huei Jiunn; Chan, Sheau Pyng Jamie; Anthony, Desiree; McQualter, Jonathan; Hansen, Michelle; Jenkins, Brendan J; Anderson, Gary P; Vlahos, Ross

2015-11-01

Cigarette smoke (CS) is the major cause of chronic obstructive pulmonary disease (COPD). Interleukin-17A (IL-17A) is a pivotal cytokine that regulates lung immunity and inflammation. The aim of the present study was to investigate how IL-17A regulates CS-induced lung inflammation in vivo. IL-17A knockout (KO) mice and neutralization of IL-17A in wild-type (WT) mice reduced macrophage and neutrophil recruitment and chemokine (C-C motif) ligand 2 (CCL2), CCL3 and matrix metalloproteinase (MMP)-12 mRNA expression in response to acute CS exposure. IL-17A expression was increased in non-obese diabetic (NOD) severe combined immunodeficiency SCID) mice with non-functional B- and T-cells over a 4-week CS exposure period, where macrophages accumulated to the same extent as in WT mice. Gene expression analysis by QPCR (quantitative real-time PCR) of isolated immune cell subsets detected increased levels of IL-17A transcript in macrophages, neutrophils and NK/NKT cells in the lungs of CS-exposed mice. In order to further explore the relative contribution of innate immune cellular sources, intracellular IL-17A staining was performed. In the present study, we demonstrate that CS exposure primes natural killer (NK), natural killer T (NKT) and γδ T-cells to produce more IL-17A protein and CS alone increased the frequency of IL17+ γδ T-cells in the lung, whereas IL-17A protein was not detected in macrophages and neutrophils. Our data suggest that activation of innate cellular sources of IL-17A is an essential mediator of macrophage accumulation in CS-exposed lungs. Targeting non-conventional T-cell sources of IL-17A may offer an alternative strategy to reduce pathogenic macrophages in COPD. © 2015 Authors; published by Portland Press Limited.

Innate cellular sources of interleukin-17A regulate macrophage accumulation in cigarette- smoke-induced lung inflammation in mice

PubMed Central

Bozinovski, Steven; Seow, Huei Jiunn; Chan, Sheau Pyng Jamie; Anthony, Desiree; McQualter, Jonathan; Hansen, Michelle; Jenkins, Brendan J.; Anderson, Gary P.

2015-01-01

Cigarette smoke (CS) is the major cause of chronic obstructive pulmonary disease (COPD). Interleukin-17A (IL-17A) is a pivotal cytokine that regulates lung immunity and inflammation. The aim of the present study was to investigate how IL-17A regulates CS-induced lung inflammation in vivo. IL-17A knockout (KO) mice and neutralization of IL-17A in wild-type (WT) mice reduced macrophage and neutrophil recruitment and chemokine (C-C motif) ligand 2 (CCL2), CCL3 and matrix metalloproteinase (MMP)-12 mRNA expression in response to acute CS exposure. IL-17A expression was increased in non-obese diabetic (NOD) severe combined immunodeficiency SCID) mice with non-functional B- and T-cells over a 4-week CS exposure period, where macrophages accumulated to the same extent as in WT mice. Gene expression analysis by QPCR (quantitative real-time PCR) of isolated immune cell subsets detected increased levels of IL-17A transcript in macrophages, neutrophils and NK/NKT cells in the lungs of CS-exposed mice. In order to further explore the relative contribution of innate immune cellular sources, intracellular IL-17A staining was performed. In the present study, we demonstrate that CS exposure primes natural killer (NK), natural killer T (NKT) and γδ T-cells to produce more IL-17A protein and CS alone increased the frequency of IL17+ γδ T-cells in the lung, whereas IL-17A protein was not detected in macrophages and neutrophils. Our data suggest that activation of innate cellular sources of IL-17A is an essential mediator of macrophage accumulation in CS-exposed lungs. Targeting non-conventional T-cell sources of IL-17A may offer an alternative strategy to reduce pathogenic macrophages in COPD. PMID:26201093

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

PubMed

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

2017-10-17

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

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

PubMed

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.

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

PubMed

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.

The Inositol Phosphatase SHIP-1 Inhibits NOD2-Induced NF-κB Activation by Disturbing the Interaction of XIAP with RIP2

PubMed Central

Condé, Claude; Rambout, Xavier; Lebrun, Marielle; Lecat, Aurore; Di Valentin, Emmanuel; Dequiedt, Franck; Piette, Jacques

2012-01-01

SHIP-1 is an inositol phosphatase predominantly expressed in hematopoietic cells. Over the ten past years, SHIP-1 has been described as an important regulator of immune functions. Here, we characterize a new inhibitory function for SHIP-1 in NOD2 signaling. NOD2 is a crucial cytoplasmic bacterial sensor that activates proinflammatory and antimicrobial responses upon bacterial invasion. We observed that SHIP-1 decreases NOD2-induced NF-κB activation in macrophages. This negative regulation relies on its interaction with XIAP. Indeed, we observed that XIAP is an essential mediator of the NOD2 signaling pathway that enables proper NF-κB activation in macrophages. Upon NOD2 activation, SHIP-1 C-terminal proline rich domain (PRD) interacts with XIAP, thereby disturbing the interaction between XIAP and RIP2 in order to decrease NF-κB signaling. PMID:22815893

Gla-rich protein function as an anti-inflammatory agent in monocytes/macrophages: Implications for calcification-related chronic inflammatory diseases

PubMed Central

Viegas, Carla S. B.; Costa, Rúben M.; Santos, Lúcia; Videira, Paula A.; Silva, Zélia; Araújo, Nuna; Macedo, Anjos L.; Matos, António P.; Vermeer, Cees; Simes, Dina C.

2017-01-01

Calcification-related chronic inflammatory diseases are multifactorial pathological processes, involving a complex interplay between inflammation and calcification events in a positive feed-back loop driving disease progression. Gla-rich protein (GRP) is a vitamin K dependent protein (VKDP) shown to function as a calcification inhibitor in cardiovascular and articular tissues, and proposed as an anti-inflammatory agent in chondrocytes and synoviocytes, acting as a new crosstalk factor between these two interconnected events in osteoarthritis. However, a possible function of GRP in the immune system has never been studied. Here we focused our investigation in the involvement of GRP in the cell inflammatory response mechanisms, using a combination of freshly isolated human leucocytes and undifferentiated/differentiated THP-1 cell line. Our results demonstrate that VKDPs such as GRP and matrix gla protein (MGP) are synthesized and γ-carboxylated in the majority of human immune system cells either involved in innate or adaptive immune responses. Stimulation of THP-1 monocytes/macrophages with LPS or hydroxyapatite (HA) up-regulated GRP expression, and treatments with GRP or GRP-coated basic calcium phosphate crystals resulted in the down-regulation of mediators of inflammation and inflammatory cytokines, independently of the protein γ-carboxylation status. Moreover, overexpression of GRP in THP-1 cells rescued the inflammation induced by LPS and HA, by down-regulation of the proinflammatory cytokines TNFα, IL-1β and NFkB. Interestingly, GRP was detected at protein and mRNA levels in extracellular vesicles released by macrophages, which may act as vehicles for extracellular trafficking and release. Our data indicate GRP as an endogenous mediator of inflammatory responses acting as an anti-inflammatory agent in monocytes/macrophages. We propose that in a context of chronic inflammation and calcification-related pathologies, GRP might act as a novel molecular mediator linking inflammation and calcification events, with potential therapeutic application. PMID:28542410

Cytokine overproduction and crosslinker hypersensitivity are unlinked in Fanconi anemia macrophages.

PubMed

Garbati, Michael R; Hays, Laura E; Rathbun, R Keaney; Jillette, Nathaniel; Chin, Kathy; Al-Dhalimy, Muhsen; Agarwal, Anupriya; Newell, Amy E Hanlon; Olson, Susan B; Bagby, Grover C

2016-03-01

The Fanconi anemia proteins participate in a canonical pathway that repairs cross-linking agent-induced DNA damage. Cells with inactivated Fanconi anemia genes are universally hypersensitive to such agents. Fanconi anemia-deficient hematopoietic stem cells are also hypersensitive to inflammatory cytokines, and, as importantly, Fanconi anemia macrophages overproduce such cytokines in response to TLR4 and TLR7/8 agonists. We questioned whether TLR-induced DNA damage is the primary cause of aberrantly regulated cytokine production in Fanconi anemia macrophages by quantifying TLR agonist-induced TNF-α production, DNA strand breaks, crosslinker-induced chromosomal breakage, and Fanconi anemia core complex function in Fanconi anemia complementation group C-deficient human and murine macrophages. Although both M1 and M2 polarized Fanconi anemia cells were predictably hypersensitive to mitomycin C, only M1 macrophages overproduced TNF-α in response to TLR-activating signals. DNA damaging agents alone did not induce TNF-α production in the absence of TLR agonists in wild-type or Fanconi anemia macrophages, and mitomycin C did not enhance TLR responses in either normal or Fanconi anemia cells. TLR4 and TLR7/8 activation induced cytokine overproduction in Fanconi anemia macrophages. Also, although TLR4 activation was associated with induced double strand breaks, TLR7/8 activation was not. That DNA strand breaks and chromosome breaks are neither necessary nor sufficient to account for the overproduction of inflammatory cytokines by Fanconi anemia cells suggests that noncanonical anti-inflammatory functions of Fanconi anemia complementation group C contribute to the aberrant macrophage phenotype and suggests that suppression of macrophage/TLR hyperreactivity might prevent cytokine-induced stem cell attrition in Fanconi anemia. © Society for Leukocyte Biology.

Development of mannose functionalized dendrimeric nanoparticles for targeted delivery to macrophages: use of this platform to modulate atherosclerosis.

PubMed

He, Hongliang; Yuan, Quan; Bie, Jinghua; Wallace, Ryan L; Yannie, Paul J; Wang, Jing; Lancina, Michael G; Zolotarskaya, Olga Yu; Korzun, William; Yang, Hu; Ghosh, Shobha

2018-03-01

Dysfunctional macrophages underlie the development of several diseases including atherosclerosis where accumulation of cholesteryl esters and persistent inflammation are 2 of the critical macrophage processes that regulate the progression as well as stability of atherosclerotic plaques. Ligand-dependent activation of liver-x-receptor (LXR) not only enhances mobilization of stored cholesteryl ester but also exerts anti-inflammatory effects mediated via trans-repression of proinflammatory transcription factor nuclear factor kappa B. However, increased hepatic lipogenesis by systemic administration of LXR ligands (LXR-L) has precluded their therapeutic use. The objective of the present study was to devise a strategy to selectively deliver LXR-L to atherosclerotic plaque-associated macrophages while limiting hepatic uptake. Mannose-functionalized dendrimeric nanoparticles (mDNP) were synthesized to facilitate active uptake via the mannose receptor expressed exclusively by macrophages using polyamidoamine dendrimer. Terminal amine groups were used to conjugate mannose and LXR-L T091317 via polyethylene glycol spacers. mDNP-LXR-L was effectively taken up by macrophages (and not by hepatocytes), increased expression of LXR target genes (ABCA1/ABCG1), and enhanced cholesterol efflux. When administered intravenously to LDLR-/- mice with established plaques, significant accumulation of fluorescently labeled mDNP-LXR-L was seen in atherosclerotic plaque-associated macrophages. Four weekly injections of mDNP-LXR-L led to significant reduction in atherosclerotic plaque progression, plaque necrosis, and plaque inflammation as assessed by expression of nuclear factor kappa B target gene matrix metalloproteinase 9; no increase in hepatic lipogenic genes or plasma lipids was observed. These studies validate the development of a macrophage-specific delivery platform for the delivery of anti-atherosclerotic agents directly to the plaque-associated macrophages to attenuate plaque burden. Copyright © 2017 Elsevier Inc. All rights reserved.

Regulation of Mitochondria Function by TRAF3 in B Lymphocytes and B Cell Malignancies

DTIC Science & Technology

2015-10-01

1, 2014. 2. Chair, Block Symposia of Innate Immune Responses in Monocytes/Macrophages, Dendritic Cells , and Myeloid Cells , the Annual Meeting of...Xie P. TRAF3-mediated regulation of innate immunity and inflammation. Research Forum, Department of Cell Biology and Neuroscience, Rutgers...TRAF3: a regulator of innate immunity and inflammation. Department of Cell & Molecular Physiology, University of Loyola, Chicago, IL. Aug. 27, 2014 9

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

PubMed Central

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 expression of cytokines in macrophages. This finding suggests that CAEV may modulate the accessory functions of infected macrophages and the antiviral immune response in vivo. PMID:9311828

Macrophage Fusion Is Controlled by the Cytoplasmic Protein Tyrosine Phosphatase PTP-PEST/PTPN12

PubMed Central

Rhee, Inmoo; Davidson, Dominique; Souza, Cleiton Martins; Vacher, Jean

2013-01-01

Macrophages can undergo cell-cell fusion, leading to the formation of multinucleated giant cells and osteoclasts. This process is believed to promote the proteolytic activity of macrophages toward pathogens, foreign bodies, and extracellular matrices. Here, we examined the role of PTP-PEST (PTPN12), a cytoplasmic protein tyrosine phosphatase, in macrophage fusion. Using a macrophage-targeted PTP-PEST-deficient mouse, we determined that PTP-PEST was not needed for macrophage differentiation or cytokine production. However, it was necessary for interleukin-4-induced macrophage fusion into multinucleated giant cells in vitro. It was also needed for macrophage fusion following implantation of a foreign body in vivo. Moreover, in the RAW264.7 macrophage cell line, PTP-PEST was required for receptor activator of nuclear factor kappa-B ligand (RANKL)-triggered macrophage fusion into osteoclasts. PTP-PEST had no impact on expression of fusion mediators such as β-integrins, E-cadherin, and CD47, which enable macrophages to become fusion competent. However, it was needed for polarization of macrophages, migration induced by the chemokine CC chemokine ligand 2 (CCL2), and integrin-induced spreading, three key events in the fusion process. PTP-PEST deficiency resulted in specific hyperphosphorylation of the protein tyrosine kinase Pyk2 and the adaptor paxillin. Moreover, a fusion defect was induced upon treatment of normal macrophages with a Pyk2 inhibitor. Together, these data argue that macrophage fusion is critically dependent on PTP-PEST. This function is seemingly due to the ability of PTP-PEST to control phosphorylation of Pyk2 and paxillin, thereby regulating cell polarization, migration, and spreading. PMID:23589331

Macrophage fusion is controlled by the cytoplasmic protein tyrosine phosphatase PTP-PEST/PTPN12.

PubMed

Rhee, Inmoo; Davidson, Dominique; Souza, Cleiton Martins; Vacher, Jean; Veillette, André

2013-06-01

Macrophages can undergo cell-cell fusion, leading to the formation of multinucleated giant cells and osteoclasts. This process is believed to promote the proteolytic activity of macrophages toward pathogens, foreign bodies, and extracellular matrices. Here, we examined the role of PTP-PEST (PTPN12), a cytoplasmic protein tyrosine phosphatase, in macrophage fusion. Using a macrophage-targeted PTP-PEST-deficient mouse, we determined that PTP-PEST was not needed for macrophage differentiation or cytokine production. However, it was necessary for interleukin-4-induced macrophage fusion into multinucleated giant cells in vitro. It was also needed for macrophage fusion following implantation of a foreign body in vivo. Moreover, in the RAW264.7 macrophage cell line, PTP-PEST was required for receptor activator of nuclear factor kappa-B ligand (RANKL)-triggered macrophage fusion into osteoclasts. PTP-PEST had no impact on expression of fusion mediators such as β-integrins, E-cadherin, and CD47, which enable macrophages to become fusion competent. However, it was needed for polarization of macrophages, migration induced by the chemokine CC chemokine ligand 2 (CCL2), and integrin-induced spreading, three key events in the fusion process. PTP-PEST deficiency resulted in specific hyperphosphorylation of the protein tyrosine kinase Pyk2 and the adaptor paxillin. Moreover, a fusion defect was induced upon treatment of normal macrophages with a Pyk2 inhibitor. Together, these data argue that macrophage fusion is critically dependent on PTP-PEST. This function is seemingly due to the ability of PTP-PEST to control phosphorylation of Pyk2 and paxillin, thereby regulating cell polarization, migration, and spreading.

Role of KCa3.1 Channels in Macrophage Polarization and Its Relevance in Atherosclerotic Plaque Instability.

PubMed

Xu, Rende; Li, Chenguang; Wu, Yizhe; Shen, Li; Ma, Jianying; Qian, Juying; Ge, Junbo

2017-02-01

Emerging evidence indicates that proinflammatory macrophage polarization imbalance plays a key role in atherosclerotic plaque progression and instability. The calcium-activated potassium channel KCa3.1 is critically involved in macrophage activation and function. However, the role of KCa3.1 in macrophage polarization is unknown. This study investigates the potential role of KCa3.1 in transcriptional regulation in macrophage polarization and its relationship to plaque instability. Human monocytes were differentiated into macrophages using macrophage colony-stimulating factor. Macrophages were then polarized into proinflammatory M1 cells by interferon-γ and lipopolysaccharide and into alternative M2 macrophages by interleukin-4. A model for plaque instability was induced by combined partial ligation of the left renal artery and left common carotid artery in apolipoprotein E knockout mice. Significant upregulation of KCa3.1 expression was observed during the differentiation of human monocytes into macrophages. Blocking KCa3.1 significantly reduced the expression of proinflammatory genes during macrophages polarization. Further mechanistic studies indicated that blocking KCa3.1 inhibited macrophage differentiation toward the M1 phenotype by downregulating signal transducer and activator of transcription-1 phosphorylation. In animal models, KCa3.1 blockade therapy strikingly reduced the incidence of plaque rupture and luminal thrombus in carotid arteries, decreased the expression of markers associated with M1 macrophage polarization, and enhanced the expression of M2 markers within atherosclerotic lesions. These results suggest that blocking KCa3.1 suppresses plaque instability in advanced stages of atherosclerosis by inhibiting macrophage polarization toward an M1 phenotype. © 2016 American Heart Association, Inc.

Macrophage migration inhibitory factor counter-regulates dexamethasone-induced annexin 1 expression and influences the release of eicosanoids in murine macrophages.

PubMed

Sun, Yu; Wang, Yu; Li, Jia-Hui; Zhu, Shi-Hui; Tang, Hong-Tai; Xia, Zhao-Fan

2013-10-01

Macrophage migration inhibitory factor (MIF), a pro-inflammatory cytokine and glucocorticoid (GC) counter-regulator, has emerged as an important modulator of inflammatory responses. However, the molecular mechanisms of MIF counter-regulation of GC still remain incomplete. In the present study, we investigated whether MIF mediated the counter-regulation of the anti-inflammatory effect of GC by affecting annexin 1 in RAW 264.7 macrophages. We found that stimulation of RAW 264.7 macrophages with lipopolysaccharide (LPS) resulted in down-regulation of annexin 1, while GC dexamethasone (Dex) or Dex plus LPS led to significant up-regulation of annexin 1 expression. RNA interference-mediated knockdown of intracellular MIF increased annexin 1 expression with or without incubation of Dex, whereas Dex-induced annexin 1 expression was counter-regulated by the exogenous application of recombinant MIF. Moreover, recombinant MIF counter-regulated, in a dose-dependent manner, inhibition of cytosolic phospholipase A2α (cPLA2α) activation and prostaglandin E2 (PGE2 ) and leukotriene B4 (LTB4 ) release by Dex in RAW 264.7 macrophages stimulated with LPS. Endogenous depletion of MIF enhanced the effects of Dex, reflected by further decease of cPLA2α expression and lower PGE2 and LTB4 release in RAW 264.7 macrophages. Based on these data, we suggest that MIF counter-regulates Dex-induced annexin 1 expression, further influencing the activation of cPLA2α and the release of eicosanoids. These findings will add new insights into the mechanisms of MIF counter-regulation of GC. © 2013 John Wiley & Sons Ltd.

Phosphatidic acid inhibits ceramide 1-phosphate-stimulated macrophage migration.

PubMed

Ouro, Alberto; Arana, Lide; Rivera, Io-Guané; Ordoñez, Marta; Gomez-Larrauri, Ana; Presa, Natalia; Simón, Jorge; Trueba, Miguel; Gangoiti, Patricia; Bittman, Robert; Gomez-Muñoz, Antonio

2014-12-15

Ceramide 1-phosphate (C1P) was recently demonstrated to potently induce cell migration. This action could only be observed when C1P was applied exogenously to cells in culture, and was inhibited by pertussis toxin. However, the mechanisms involved in this process are poorly understood. In this work, we found that phosphatidic acid (PA), which is structurally related to C1P, displaced radiolabeled C1P from its membrane-binding site and inhibited C1P-stimulated macrophage migration. This effect was independent of the saturated fatty acid chain length or the presence of a double bond in each of the fatty acyl chains of PA. Treatment of RAW264.7 macrophages with exogenous phospholipase D (PLD), an enzyme that produces PA from membrane phospholipids, also inhibited C1P-stimulated cell migration. Likewise, PA or exogenous PLD inhibited C1P-stimulated extracellularly regulated kinases (ERK) 1 and 2 phosphorylation, leading to inhibition of cell migration. However, PA did not inhibit C1P-stimulated Akt phosphorylation. It is concluded that PA is a physiological regulator of C1P-stimulated macrophage migration. These actions of PA may have important implications in the control of pathophysiological functions that are regulated by C1P, including inflammation and various cellular processes associated with cell migration such as organogenesis or tumor metastasis. Copyright © 2014 Elsevier Inc. All rights reserved.

Activation by nitric oxide of an oxidative-stress response that defends Escherichia coli against activated macrophages.

PubMed Central

Nunoshiba, T; deRojas-Walker, T; Wishnok, J S; Tannenbaum, S R; Demple, B

1993-01-01

Nitric oxide is a free radical (NO) formed biologically through the oxidation of L-arginine by nitric oxide synthases. NO is produced transiently in mammalian cells for intercellular signaling and in copious quantities to cause cytostasis and cytotoxicity. In the latter situation, NO is a deliberate cytotoxic product of activated macrophages, along with other reactive oxygen species such as hydrogen peroxide (H2O2) and superoxide (O2-). Escherichia coli has a complex set of responses to H2O2 and O2- that involves approximately 80 inducible proteins; we wondered whether these bacteria might induce analogous defenses against nitric oxide. We show here that a multigene system controlled by the redox-sensitive transcriptional regulator SoxR is activated by NO in vivo. This induction confers bacterial resistance to activated murine macrophages with kinetics that parallel the production of NO by these cells. Elimination of specific SoxR-regulated genes diminishes the resistance of these bacteria to the cytotoxic macrophages. The required functions include manganese-containing superoxide dismutase, endonuclease IV (a DNA-repair enzyme for oxidative damage), and micF, an antisense regulator of the outer membrane porin OmpF. These results demonstrate that SoxR is a sensor for cellular exposure to NO, and that the soxRS response system may contribute to bacterial virulence. PMID:8234347

Lineage-specific functions of Bcl-6 in immunity and inflammation are mediated through distinct biochemical mechanisms

PubMed Central

Huang, Chuanxin; Hatzi, Katerina; Melnick, Ari

2013-01-01

The transcription factor Bcl-6 orchestrates the germinal center reaction through its actions in B and T cells, and regulates inflammatory signaling in macrophages. We report that genetic replacement by mutant Bcl-6, which cannot bind corepressors to its BTB domain, disrupted germinal center formation and immunoglobulin affinity maturation, due to a defect in B cell proliferation and survival. In contrast, BTB loss of function had no effect on T follicular helper cell differentiation and function, nor other T helper subsets. Bcl6 null mice displayed a lethal inflammatory phenotype, whereas BTB mutant mice experienced normal healthy lives with no inflammation. Bcl-6 repression of inflammatory responses in macrophages was accordingly independent of the BTB domain repressor function. Bcl-6 thus mediates its actions through lineage-specific biochemical functions. PMID:23455674

Epigenetic programming during monocyte to macrophage differentiation and trained innate immunity

PubMed Central

Saeed, Sadia; Quintin, Jessica; Kerstens, Hindrik H.D.; Rao, Nagesha A; Aghajanirefah, Ali; Matarese, Filomena; Cheng, Shih-Chin; Ratter, Jacqueline; Berentsen, Kim; van der Ent, Martijn A.; Sharifi, Nilofar; Janssen-Megens, Eva M.; Huurne, Menno Ter; Mandoli, Amit; van Schaik, Tom; Ng, Aylwin; Burden, Frances; Downes, Kate; Frontini, Mattia; Kumar, Vinod; Giamarellos-Bourboulis, Evangelos J; Ouwehand, Willem H; van der Meer, Jos W.M.; Joosten, Leo A.B.; Wijmenga, Cisca; Martens, Joost H.A.; Xavier, Ramnik J.; Logie, Colin; Netea, Mihai G.; Stunnenberg, Hendrik G.

2014-01-01

Structured Abstract Introduction Monocytes circulate in the bloodstream for up to 3–5 days. Concomitantly, immunological imprinting of either tolerance (immunosuppression) or trained immunity (innate immune memory) determines the functional fate of monocytes and monocyte-derived macrophages, as observed after infection or vaccination. Methods Purified circulating monocytes from healthy volunteers were differentiated under the homeostatic M-CSF concentrations present in human serum. During the first 24 hours, trained immunity was induced by β-glucan (BG) priming, while post-sepsis immunoparalysis was mimicked by exposure to LPS, generating endotoxin-induced tolerance. Epigenomic profiling of the histone marks H3K4me1, H3K4me3 and H3K27ac, DNase I accessibility and RNA sequencing were performed at both the start of the experiment (ex vivo monocytes) and at the end of the six days of in vitro culture (macrophages). Results Compared to monocytes (Mo), naïve macrophages (Mf) display a remodeled metabolic enzyme repertoire and attenuated innate inflammatory pathways; most likely necessary to generate functional tissue macrophages. Epigenetic profiling uncovered ~8000 dynamic regions associated with ~11000 DNase I hypersensitive sites. Changes in histone acetylation identified most dynamic events. Furthermore, these regions of differential histone marks displayed some degree of DNase I accessibility that was already present in monocytes. H3K4me1 mark increased in parallel with de novo H3K27ac deposition at distal regulatory regions; H3K4me1 mark remained even after the loss of H3K27ac, marking decommissioned regulatory elements. β-glucan priming specifically induced ~3000 distal regulatory elements, whereas LPS-tolerization uniquely induced H3K27ac at ~500 distal regulatory regions. At the transcriptional level, we identified co-regulated gene modules during monocyte to macrophage differentiation, as well as discordant modules between trained and tolerized cells. These indicate that training likely involves an increased expression of modules expressed in naïve macrophages, including genes that code for metabolic enzymes. On the other hand, endotoxin tolerance involves gene modules that are more active in monocytes than in naïve macrophages. About 12% of known human transcription factors display variation in expression during macrophage differentiation, training and tolerance. We also observed transcription factor motifs in DNase I hypersensitive sites at condition-specific dynamic epigenomic regions, implying that specific transcription factors are required for trained and tolerized macrophage epigenetic and transcriptional programs. Finally, our analyses and functional validation indicate that the inhibition of cAMP generation blocked trained immunity in vitro and during an in vivo model of lethal C. albicans infection, abolishing the protective effects of trained immunity. Discussion We documented the importance of epigenetic regulation of the immunological pathways underlying monocyte-to-macrophage differentiation and trained immunity. These dynamic epigenetic elements may inform on potential pharmacological targets that modulate innate immunity. Altogether, we uncovered the epigenetic and transcriptional programs of monocyte differentiation to macrophages that distinguish tolerant and trained macrophage phenotypes, providing a resource to further understand and manipulate immune-mediated responses. PMID:25258085

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

PubMed

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

2012-05-01

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

Macrophage Phenotype Controls Long-Term AKI Outcomes—Kidney Regeneration versus Atrophy

PubMed Central

Gröbmayr, Regina; Ryu, Mi; Lorenz, Georg; Hartter, Ingo; Mulay, Shrikant R.; Susanti, Heni Eka; Kobayashi, Koichi S.; Flavell, Richard A.; Anders, Hans-Joachim

2014-01-01

The mechanisms that determine full recovery versus subsequent progressive CKD after AKI are largely unknown. Because macrophages regulate inflammation as well as epithelial recovery, we investigated whether macrophage activation influences AKI outcomes. IL-1 receptor–associated kinase-M (IRAK-M) is a macrophage-specific inhibitor of Toll-like receptor (TLR) and IL-1 receptor signaling that prevents polarization toward a proinflammatory phenotype. In postischemic kidneys of wild-type mice, IRAK-M expression increased for 3 weeks after AKI and declined thereafter. However, genetic depletion of IRAK-M did not affect immunopathology and renal dysfunction during early postischemic AKI. Regarding long-term outcomes, wild-type kidneys regenerated completely within 5 weeks after AKI. In contrast, IRAK-M−/− kidneys progressively lost up to two-thirds of their original mass due to tubule loss, leaving atubular glomeruli and interstitial scarring. Moreover, M1 macrophages accumulated in the renal interstitial compartment, coincident with increased expression of proinflammatory cytokines and chemokines. Injection of bacterial CpG DNA induced the same effects in wild-type mice, and TNF-α blockade with etanercept partially prevented renal atrophy in IRAK-M−/− mice. These results suggest that IRAK-M induction during the healing phase of AKI supports the resolution of M1 macrophage– and TNF-α–dependent renal inflammation, allowing structural regeneration and functional recovery of the injured kidney. Conversely, IRAK-M loss-of-function mutations or transient exposure to bacterial DNA may drive persistent inflammatory mononuclear phagocyte infiltrates, which impair kidney regeneration and promote CKD. Overall, these results support a novel role for IRAK-M in the regulation of wound healing and tissue regeneration. PMID:24309188

F4/80: the macrophage-specific adhesion-GPCR and its role in immunoregulation.

PubMed

Lin, Hsi-Hsien; Stacey, Martin; Stein-Streilein, Joan; Gordon, Siamon

2010-01-01

As a macrophage-restricted reagent, the generation and application of the F4/80 mAb has greatly benefited the phenotypic characterization of mouse tissue macrophages for three decades. Following the molecular identification of the F4/80 antigen as an EGF-TM7 member of the adhesion-GPCR family, great interest was ignited to understand its cell type-specific expression pattern as well as its functional role in macrophage biology. Recent studies have shown that the F4/80 gene is regulated by a novel set of transcription factors that recognized a unique promoter sequence. Gene targeting experiments have produced two F4/80 knock out animal models and showed that F4/80 is not required for normal macrophage development. Nevertheless, the F4/80 receptor was found to be necessary for the induction of efferent CD8+ regulatory T cells responsible for peripheral immune tolerance. The identification of cellular ligands for F4/80 and delineation of its signaling pathway remain elusive but are critical to understand the in vivo role of this macrophage-specific adhesion-GPCR.

Methamphetamine and HIV-1 gp120 Effects on Lipopolysaccharide Stimulated Matrix Metalloproteinase-9 Production by Human Monocyte-Derived Macrophages

PubMed Central

Reynolds, Jessica L.; Mahajan, Supriya D.; Aalinkeel, Ravikumar; Nair, Bindukumar; Sykes, Donald E.; Schwartz, Stanley A.

2011-01-01

Monocytes/macrophages are a primary source of human immunodeficiency virus (HIV-1) in the central nervous system (CNS). Macrophages infected with HIV-1 produce a plethora of factors, including matrix metalloproteinase-9 (MMP-9) that may contribute to the development of HIV-1-associated neurocognitive disorders (HAND). MMP-9 plays a pivotal role in the turnover of the extracellular matrix (ECM) and functions to remodel cellular architecture. We have investigated the role of methamphetamine and HIV-1 gp120 in the regulation of lipopolysaccaride (LPS) induced-MMP-9 production in monocyte-derived macrophages (MDM). Here, we show that LPS-induced MMP-9 gene expression and protein secretion are potentiated by incubation with methamphetamine alone and gp120 alone. Further, concomitant incubation with gp120 and methamphetamine potentiated LPS-induced MMP-9 expression and biological activity in MDM. Collectively methamphetamine and gp120 effects on MMPs may modulate remodeling of the extracellular environment enhancing migration of monocytes/macrophages to the CNS. PMID:21425912

DUSP3 genetic deletion confers M2-like−macrophage-dependent tolerance to septic shock

PubMed Central

Singh, Pratibha; Dejager, Lien; Amand, Mathieu; Theatre, Emilie; Vandereyken, Maud; Zurashvilli, Tinatin; Singh, Maneesh; Mack, Matthias; Timmermans, Steven; Musumeci, Lucia; Dejardin, Emmanuel; Mustelin, Tomas; Van Ginderachter, Jo A.; Moutschen, Michel; Oury, Cécile; Libert, Claude; Rahmouni, Souad

2015-01-01

DUSP3 is a small dual-specificity protein phosphatase with an unknown physiological function. We report that DUSP3 is strongly expressed in human and mouse monocytes and macrophages and that its deficiency in mice promotes tolerance to lipopolysaccharide (LPS)-induced endotoxin shock and to polymicrobial septic shock following cecal ligation and puncture. By using adoptive transfer experiments, we demonstrate that resistance to endotoxin is macrophage-dependent and transferable and that this protection is associated with a striking increase of M2-like macrophages in DUSP3−/− mice in both the LPS and cecal ligation and puncture models. We show that the altered response of DUSP3−/− mice to sepsis is reflected in decreased TNF production and impaired ERK1/2 activation. Our results demonstrate that DUSP3 plays a key and non-redundant role as a regulator of innate immune responses by mechanisms involving the control of ERK1/2 activation, TNF secretion and macrophage polarization. PMID:25876765

Steroids Regulate CXCL4 in the Human Endometrium During Menstruation to Enable Efficient Endometrial Repair

PubMed Central

Maybin, Jacqueline A.; Thiruchelvam, Uma; Madhra, Mayank; Saunders, Philippa T.K.

2017-01-01

Context: Repair of the endometrial surface at menstruation must be efficient to minimize blood loss and optimize reproductive function. The mechanism and regulation of endometrial repair remain undefined. Objective: To determine the presence/regulation of CXCL4 in the human endometrium as a putative repair factor at menses. Patients/Setting: Endometrial tissue was collected throughout the menstrual cycle from healthy women attending the gynecology department. Menstrual blood loss was objectively measured in a subset, and heavy menstrual bleeding (HMB) was defined as >80 mL per cycle. Monocytes were isolated from peripheral blood. Design: CXCL4 messenger RNA (mRNA) and protein were identified by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. The function/regulation of endometrial CXCL4 was explored by in vitro cell culture. Results: CXCL4 mRNA concentrations were significantly increased during menstruation. Intense staining for CXCL4 was detected in late secretory and menstrual tissue, localized to stromal, epithelial and endothelial cells. Colocalization identified positive staining in CD68+ macrophages. Treatment of human endometrial stromal and endothelial cells (hESCs and HEECs, respectively) with steroids revealed differential regulation of CXCL4. Progesterone withdrawal resulted in significant increases in CXCL4 mRNA and protein in hESCs, whereas cortisol significantly increased CXCL4 in HEECs. In women with HMB, CXCL4 was reduced in endothelial cells during the menstrual phase compared with women with normal menstrual bleeding. Cortisol-exposed macrophages displayed increased chemotaxis toward CXCL4 compared with macrophages incubated with estrogen or progesterone. Conclusions: These data implicate CXCL4 in endometrial repair after menses. Reduced cortisol at the time of menses may contribute to delayed endometrial repair and HMB, in part by mechanisms involving aberrant expression of CXCL4. PMID:28323919

Steroids Regulate CXCL4 in the Human Endometrium During Menstruation to Enable Efficient Endometrial Repair.

PubMed

Maybin, Jacqueline A; Thiruchelvam, Uma; Madhra, Mayank; Saunders, Philippa T K; Critchley, Hilary O D

2017-06-01

Repair of the endometrial surface at menstruation must be efficient to minimize blood loss and optimize reproductive function. The mechanism and regulation of endometrial repair remain undefined. To determine the presence/regulation of CXCL4 in the human endometrium as a putative repair factor at menses. Endometrial tissue was collected throughout the menstrual cycle from healthy women attending the gynecology department. Menstrual blood loss was objectively measured in a subset, and heavy menstrual bleeding (HMB) was defined as >80 mL per cycle. Monocytes were isolated from peripheral blood. CXCL4 messenger RNA (mRNA) and protein were identified by quantitative reverse transcription polymerase chain reaction and immunohistochemistry. The function/regulation of endometrial CXCL4 was explored by in vitro cell culture. CXCL4 mRNA concentrations were significantly increased during menstruation. Intense staining for CXCL4 was detected in late secretory and menstrual tissue, localized to stromal, epithelial and endothelial cells. Colocalization identified positive staining in CD68+ macrophages. Treatment of human endometrial stromal and endothelial cells (hESCs and HEECs, respectively) with steroids revealed differential regulation of CXCL4. Progesterone withdrawal resulted in significant increases in CXCL4 mRNA and protein in hESCs, whereas cortisol significantly increased CXCL4 in HEECs. In women with HMB, CXCL4 was reduced in endothelial cells during the menstrual phase compared with women with normal menstrual bleeding. Cortisol-exposed macrophages displayed increased chemotaxis toward CXCL4 compared with macrophages incubated with estrogen or progesterone. These data implicate CXCL4 in endometrial repair after menses. Reduced cortisol at the time of menses may contribute to delayed endometrial repair and HMB, in part by mechanisms involving aberrant expression of CXCL4. Copyright © 2017 by the Endocrine Society

Neutrophils induce macrophage anti-inflammatory reprogramming by suppressing NF-κB activation.

PubMed

Marwick, John A; Mills, Ross; Kay, Oliver; Michail, Kyriakos; Stephen, Jillian; Rossi, Adriano G; Dransfield, Ian; Hirani, Nikhil

2018-06-04

Apoptotic cells modulate the function of macrophages to control and resolve inflammation. Here, we show that neutrophils induce a rapid and sustained suppression of NF-κB signalling in the macrophage through a unique regulatory relationship which is independent of apoptosis. The reduction of macrophage NF-κB activation occurs through a blockade in transforming growth factor β-activated kinase 1 (TAK1) and IKKβ activation. As a consequence, NF-κB (p65) phosphorylation is reduced, its translocation to the nucleus is inhibited and NF-κB-mediated inflammatory cytokine transcription is suppressed. Gene Set Enrichment Analysis reveals that this suppression of NF-κB activation is not restricted to post-translational modifications of the canonical NF-κB pathway, but is also imprinted at the transcriptional level. Thus neutrophils exert a sustained anti-inflammatory phenotypic reprogramming of the macrophage, which is reflected by the sustained reduction in the release of pro- but not anti- inflammatory cytokines from the macrophage. Together, our findings identify a novel apoptosis-independent mechanism by which neutrophils regulate the mediator profile and reprogramming of monocytes/macrophages, representing an important nodal point for inflammatory control.

Deletion of calponin 2 in macrophages attenuates the severity of inflammatory arthritis in mice.

PubMed

Huang, Qi-Quan; Hossain, M Moazzem; Sun, Wen; Xing, Lianping; Pope, Richard M; Jin, J-P

2016-10-01

Calponin is an actin cytoskeleton-associated protein that regulates motility-based cellular functions. Three isoforms of calponin are present in vertebrates, among which calponin 2 encoded by the Cnn2 gene is expressed in multiple types of cells, including blood cells from the myeloid lineage. Our previous studies demonstrated that macrophages from Cnn2 knockout (KO) mice exhibit increased migration and phagocytosis. Intrigued by an observation that monocytes and macrophages from patients with rheumatoid arthritis had increased calponin 2, we investigated anti-glucose-6-phosphate isomerase serum-induced arthritis in Cnn2-KO mice for the effect of calponin 2 deletion on the pathogenesis and pathology of inflammatory arthritis. The results showed that the development of arthritis was attenuated in systemic Cnn2-KO mice with significantly reduced inflammation and bone erosion than that in age- and stain background-matched C57BL/6 wild-type mice. In vitro differentiation of calponin 2-null mouse bone marrow cells produced fewer osteoclasts with decreased bone resorption. The attenuation of inflammatory arthritis was confirmed in conditional myeloid cell-specific Cnn2-KO mice. The increased phagocytotic activity of calponin 2-null macrophages may facilitate the clearance of autoimmune complexes and the resolution of inflammation, whereas the decreased substrate adhesion may reduce osteoclastogenesis and bone resorption. The data suggest that calponin 2 regulation of cytoskeleton function plays a novel role in the pathogenesis of inflammatory arthritis, implicating a potentially therapeutic target. Copyright © 2016 the American Physiological Society.

Interleukin 10 (IL-10)-mediated Immunosuppression

PubMed Central

Mittal, Sharad K.; Cho, Kyung-Jin; Ishido, Satoshi; Roche, Paul A.

2015-01-01

Efficient immune responses require regulated antigen presentation to CD4 T cells. IL-10 inhibits the ability of dendritic cells (DCs) and macrophages to stimulate antigen-specific CD4 T cells; however, the mechanisms by which IL-10 suppresses antigen presentation remain poorly understood. We now report that IL-10 stimulates expression of the E3 ubiquitin ligase March-I in activated macrophages, thereby down-regulating MHC-II, CD86, and antigen presentation to CD4 T cells. By contrast, IL-10 does not stimulate March-I expression in DCs, does not suppress MHC-II or CD86 expression on either resting or activated DCs, and does not affect antigen presentation by activated DCs. IL-10 does, however, inhibit the process of DC activation itself, thereby reducing the efficiency of antigen presentation in a March-I-independent manner. Thus, IL-10 suppression of antigen presenting cell function in macrophages is March-I-dependent, whereas in DCs, suppression is March- I-independent. PMID:26408197

Proinflammatory TLR signaling is regulated by a TRAF2-dependent proteolysis mechanism in macrophages

PubMed Central

Jin, Jin; Xiao, Yichuan; Hu, Hongbo; Zou, Qiang; Li, Yanchuan; Gao, Yanpan; Ge, Wei; Cheng, Xuhong; Sun, Shao-Cong

2014-01-01

Signal transduction from toll-like receptors (TLRs) is important for innate immunity against infections, but deregulated TLR signaling contributes to inflammatory disorders. Here we show that myeloid cell-specific ablation of TRAF2 greatly promotes TLR-stimulated proinflammatory cytokine expression in macrophages and exacerbates colitis in an animal model of inflammatory bowel disease. TRAF2 deficiency does not enhance upstream signaling events, but it causes accumulation of two transcription factors, c-Rel and IRF5, known to mediate proinflammatory cytokine induction. Interestingly, TRAF2 controls the fate of c-Rel and IRF5 via a proteasome-dependent mechanism that also requires TRAF3 and the E3 ubiquitin ligase cIAP. We further show that TRAF2 also regulates inflammatory cytokine production in tumor-associated macrophages and facilitates tumor growth. These findings demonstrate an unexpected anti-inflammatory function of TRAF2 and suggest a proteasome-dependent mechanism that limits the proinflammatory TLR signaling. PMID:25565375

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

PubMed

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.

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

Huang, Yuan; Luo, Fangjun; Li, Hui

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 Hoechstmore » 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.« less

Trypanosoma cruzi Exploits Wnt Signaling Pathway to Promote Its Intracellular Replication in Macrophages.

PubMed

Volpini, Ximena; Ambrosio, Laura F; Fozzatti, Laura; Insfran, Constanza; Stempin, Cinthia C; Cervi, Laura; Motran, Claudia Cristina

2018-01-01

During the acute phase of Trypanosoma cruzi infection, macrophages can act as host cells for the parasites as well as effector cells in the early anti-parasitic immune response. Thus, the targeting of specific signaling pathways could modulate macrophages response to restrict parasite replication and instruct an appropriate adaptive response. Recently, it has become evident that Wnt signaling has immunomodulatory functions during inflammation and infection. Here, we tested the hypothesis that during T. cruzi infection, the activation of Wnt signaling pathway in macrophages plays a role in modulating the inflammatory/tolerogenic response and therefore regulating the control of parasite replication. In this report, we show that early after T. cruzi infection of bone marrow-derived macrophages (BMM), β-catenin was activated and Wnt3a, Wnt5a, and some Frizzled receptors as well as Wnt/β-catenin pathway's target genes were upregulated, with Wnt proteins signaling sustaining the activation of Wnt/β-catenin pathway and then activating the Wnt/Ca +2 pathway. Wnt signaling pathway activation was critical to sustain the parasite's replication in BMM; since the treatments with specific inhibitors of β-catenin transcriptional activation or Wnt proteins secretion limited the parasite replication. Mechanistically, inhibition of Wnt signaling pathway armed BMM to fight against T. cruzi by inducing the production of pro-inflammatory cytokines and indoleamine 2,3-dioxygenase activity and by downregulating arginase activity. Likewise, in vivo pharmacological inhibition of the Wnts' interaction with its receptors controlled the parasite replication and improved the survival of lethally infected mice. It is well established that T. cruzi infection activates a plethora of signaling pathways that ultimately regulate immune mediators to determine the modulation of a defined set of effector functions in macrophages. In this study, we have revealed a new signaling pathway that is activated by the interaction between protozoan parasites and host innate immunity, establishing a new conceptual framework for the development of new therapies.

PPE57 induces activation of macrophages and drives Th1-type immune responses through TLR2.

PubMed

Xu, Ying; Yang, Enzhuo; Huang, Qi; Ni, Wenwen; Kong, Cong; Liu, Guoyuan; Li, Guanghua; Su, Haibo; Wang, Honghai

2015-06-01

Proline-glutamic acid (PE) and proline-proline-glutamic acid (PPE) are related proteins exclusive to Mycobacteria that play diverse roles in modulating critical innate immune pathways. In this study, we observed that the PPE57 protein is associated with the cell wall and is exposed on the cell surface. PPE57 enhances Mycobacterium spp. entering into macrophages and plays a role in macrophage phagocytosis. To explore the underlying mechanism, we demonstrated that PPE57 is able to recognise Toll-like receptor 2 (TLR2) and further induce macrophage activation by augmenting the expression of several cell surface molecules (CD40, CD80, CD86 and MHC class II) and pro-inflammatory cytokines (TNF-α, IL-6 and IL-12p40) within macrophages. These molecules are involved in the mitogen-activated protein kinase (MAPK) and nuclear factor κB (NF-κB) signalling pathways. We demonstrated that PPE57 effectively polarises T cells to secrete interferon (IFN)-γ and IL-2 and to up-regulate CXCR3 expression in vivo and in vitro, suggesting that this protein may contribute to Th1 polarisation during the immune response. Moreover, recombinant Bacillus Calmette-Guérin (BCG) over-expressing PPE57 could provide better protective efficacy against Mycobacterium tuberculosis challenge compared with BCG. Taken together, our data provides several pieces of evidence that PPE57 may regulate innate and adaptive immunity by interacting with TLR2. These findings indicate that PPE57 protein is a potential antigen for the rational design of an efficient vaccine against M. tuberculosis. PPE57 is located on the cell surface and enhances mycobacterium entry into macrophage. PPE57 interacts directly with TLR2 on macrophages. PPE57 plays a key role in the activation of macrophages in a TLR2-dependent manner. PPE57 induces a Th1 immune response via TLR2-mediated macrophage functions. Recombinant BCG over-expressing PPE57 could improve protective efficacy against M. tuberculosis.

Interferon-gamma regulates nucleoside transport systems in macrophages through signal transduction and activator of transduction factor 1 (STAT1)-dependent and -independent signalling pathways.

PubMed Central

Soler, Concepció; Felipe, Antonio; García-Manteiga, José; Serra, Maria; Guillén-Gómez, Elena; Casado, F Javier; MacLeod, Carol; Modolell, Manuel; Pastor-Anglada, Marçal; Celada, Antonio

2003-01-01

The expressions of CNT and ENT (concentrative and equilibrative nucleoside transporters) in macrophages are differentially regulated by IFN-gamma (interferon-gamma). This cytokine controls gene expression through STAT1-dependent and/or -independent pathways (where STAT1 stands for signal transduction and activator of transcription 1). In the present study, the role of STAT1 in the response of nucleoside transporters to IFN-gamma was studied using macrophages from STAT1 knockout mice. IFN-gamma triggered an inhibition of ENT1-related nucleoside transport activity through STAT1-dependent mechanisms. Such inhibition of macrophage growth and ENT1 activity by IFN-gamma is required for DNA synthesis. Interestingly, IFN-gamma led to an induction of the CNT1- and CNT2-related nucleoside transport activities independent of STAT1, thus ensuring the supply of extracellular nucleosides for the STAT1-independent RNA synthesis. IFN-gamma up-regulated CNT2 mRNA and CNT1 protein levels and down-regulated ENT1 mRNA in both wild-type and STAT1 knockout macrophages. This is consistent with a STAT1-independent, long-term-mediated, probably transcription-dependent, regulation of nucleoside transporter genes. Moreover, STAT1-dependent post-transcriptional mechanisms are implicated in the regulation of ENT1 activity. Although nitric oxide is involved in the regulation of ENT1 activity in B-cells at a post-transcriptional level, our results show that STAT1-dependent induction of nitric oxide by IFN-gamma is not implicated in the regulation of ENT1 activity in macrophages. Our results indicate that both STAT1-dependent and -independent pathways are involved in the regulation of nucleoside transporters by IFN-gamma in macrophages. PMID:12868960

Src is required for migration, phagocytosis, and interferon beta production in Toll-like receptor-engaged macrophages.

PubMed

Maa, Ming-Chei; Leu, Tzeng-Horng

2016-06-01

As an evolutionarily conserved mechanism, innate immunity controls self-nonself discrimination to protect a host from invasive pathogens. Macrophages are major participants of the innate immune system. Through the activation of diverse Toll-like receptors (TLRs), macrophages are triggered to initiate a variety of functions including locomotion, phagocytosis, and secretion of cytokines that requires the participation of tyrosine kinases. Fgr, Hck, and Lyn are myeloid-specific Src family kinases. Despite their constitutively high expression in macrophages, their absence does not impair LPS responsiveness. In contrast, Src, a barely detectable tyrosine kinase in resting macrophages, becomes greatly inducible in response to TLR engagement, implicating its role in macrophage activation. Indeed, silencing Src suppresses the activated TLR-mediated migration, phagocytosis, and interferon-beta (IFN-β) secretion in macrophages. And these physiological defects can be restored by the introduction of siRNA-resistant Src. Notably, the elevated expression and activity of Src is inducible nitric oxide synthase (iNOS)-dependent. Due to (1) iNOS being a NF-κB target, which can be induced by various TLR ligands, (2) Src can mediate NF-κB activation, therefore, there ought to exist a loop of signal amplification that regulates macrophage physiology in response to the engagement of TLRs.

Rictor/mammalian target of rapamycin complex 2 promotes macrophage activation and kidney fibrosis.

PubMed

Ren, Jiafa; Li, Jianzhong; Feng, Ye; Shu, Bingyan; Gui, Yuan; Wei, Wei; He, Weichun; Yang, Junwei; Dai, Chunsun

2017-08-01

Mammalian target of rapamycin (mTOR) signalling controls many essential cellular functions. However, the role of Rictor/mTOR complex 2 (mTORC2) in regulating macrophage activation and kidney fibrosis remains largely unknown. We report here that Rictor/mTORC2 was activated in macrophages from the fibrotic kidneys of mice. Ablation of Rictor in macrophages reduced kidney fibrosis, inflammatory cell accumulation, macrophage proliferation and polarization after unilateral ureter obstruction or ischaemia/reperfusion injury. In bone marrow-derived macrophages (BMMs), deletion of Rictor or blockade of protein kinase Cα inhibited cell migration. Additionally, deletion of Rictor or blockade of Akt abolished interleukin-4-stimulated or transforming growth factor (TGF)-β1-stimulated macrophage M2 polarization. Furthermore, deletion of Rictor downregulated TGF-β1-stimulated upregulation of multiple profibrotic cytokines, including platelet-derived growth factor, vascular endothelial growth factor and connective tissue growth factor, in BMMs. Conditioned medium from TGF-β1-pretreated Rictor -/- macrophages stimulated fibroblast activation less efficiently than that from TGF-β1-pretreated Rictor +/+ macrophages. These results demonstrate that Rictor/mTORC2 signalling can promote macrophage activation and kidney fibrosis. Targeting this signalling pathway in macrophages may shine light on ways to protect against kidney fibrosis in patients with chronic kidney diseases. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

FLT1 signaling in metastasis-associated macrophages activates an inflammatory signature that promotes breast cancer metastasis

PubMed Central

Zhang, Hui; Li, Jiufeng; He, Tianfang; Yeo, Eun-Jin; Soong, Daniel Y.H.; Carragher, Neil O.; Munro, Alison; Chang, Alvin; Bresnick, Anne R.; Lang, Richard A.

2015-01-01

Although the link between inflammation and cancer initiation is well established, its role in metastatic diseases, the primary cause of cancer deaths, has been poorly explored. Our previous studies identified a population of metastasis-associated macrophages (MAMs) recruited to the lung that promote tumor cell seeding and growth. Here we show that FMS-like tyrosine kinase 1 (Flt1, also known as VEGFR1) labels a subset of macrophages in human breast cancers that are significantly enriched in metastatic sites. In mouse models of breast cancer pulmonary metastasis, MAMs uniquely express FLT1. Using several genetic models, we show that macrophage FLT1 signaling is critical for metastasis. FLT1 inhibition does not affect MAM recruitment to metastatic lesions but regulates a set of inflammatory response genes, including colony-stimulating factor 1 (CSF1), a central regulator of macrophage biology. Using a gain-of-function approach, we show that CSF1-mediated autocrine signaling in MAMs is downstream of FLT1 and can restore the tumor-promoting activity of FLT1-inhibited MAMs. Thus, CSF1 is epistatic to FLT1, establishing a link between FLT1 and inflammatory responses within breast tumor metastases. Importantly, FLT1 inhibition reduces tumor metastatic efficiency even after initial seeding, suggesting that these pathways represent therapeutic targets in metastatic disease. PMID:26261265

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

PubMed

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

2003-01-01

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

iRhom2 regulates cell surface expression of CSF1R and non-steady state myelopoiesis in mice

PubMed Central

Qing, Xiaoping; Lavin, Yonit; Redecha, Patricia; Issuree, Priya D.; Maretzky, Thorsten; Merad, Miriam; McIlwain, David; Mak, Tak W.; Overall, Christopher M.

2016-01-01

The colony stimulating factor 1 receptor (CSF1R) functions as the major receptor for macrophage colony stimulating factor (CSF1) with crucial roles in regulating myelopoeisis. CSF1R can be proteolytically released from the cell surface by A disintegrin and metalloprotease 17 (ADAM17). Here we identified CSF1R as a major substrate of ADAM17 in an unbiased degradomics screen. We explored the impact of CSF1R shedding by ADAM17 and its upstream regulator, inactive rhomboid protein 2 (iRhom2, gene name Rhbdf2), on homeostatic development of mouse myeloid cells. In iRhom2−/− mice, we found constitutive accumulation of membrane-bound CSF1R on myeloid cells at steady state, although cell numbers of these populations were not altered. However, in the context of mixed bone marrow (BM) chimera, under competitive pressure, iRhom2−/− BM progenitor-derived monocytes, tissue macrophages and lung DCs showed a repopulation advantage over those derived from wild type (WT) BM progenitors, suggesting enhanced CSF1R signaling in the absence of iRhom2. In vitro experiments indicate that iRhom2−/− Lin−SCA-1+c-Kit+ (LSKs) cells, but not granulocyte-macrophage progenitors (GMPs), had faster growth rates than WT cells in response to CSF1. Our results shed light on an important role of iRhom2/ADAM17 pathway in regulation of CSF1R shedding and repopulation of monocytes, macrophages and DCs. PMID:27601030

CARD9 knockout ameliorates myocardial dysfunction associated with high fat diet-induced obesity.

PubMed

Cao, Li; Qin, Xing; Peterson, Matthew R; Haller, Samantha E; Wilson, Kayla A; Hu, Nan; Lin, Xin; Nair, Sreejayan; Ren, Jun; He, Guanglong

2016-03-01

Obesity is associated with chronic inflammation which plays a critical role in the development of cardiovascular dysfunction. Because the adaptor protein caspase recruitment domain-containing protein 9 (CARD9) in macrophages regulates innate immune responses via activation of pro-inflammatory cytokines, we hypothesize that CARD9 mediates the pro-inflammatory signaling associated with obesity en route to myocardial dysfunction. C57BL/6 wild-type (WT) and CARD9(-/-) mice were fed normal diet (ND, 12% fat) or a high fat diet (HFD, 45% fat) for 5months. At the end of 5-month HFD feeding, cardiac function was evaluated using echocardiography. Cardiomyocytes were isolated and contractile properties were measured. Immunofluorescence was performed to detect macrophage infiltration in the heart. Heart tissue homogenates, plasma, and supernatants from isolated macrophages were collected to measure the concentrations of pro-inflammatory cytokines using ELISA kits. Western immunoblotting analyses were performed on heart tissue homogenates and isolated macrophages to explore the underlying signaling mechanism(s). CARD9 knockout alleviated HFD-induced insulin resistance and glucose intolerance, prevented myocardial dysfunction with preserved cardiac fractional shortening and cardiomyocyte contractile properties. CARD9 knockout also significantly decreased the number of infiltrated macrophages in the heart with reduced myocardium-, plasma-, and macrophage-derived cytokines including IL-6, IL-1β and TNFα. Finally, CARD9 knockout abrogated the increase of p38 MAPK phosphorylation, the decrease of LC3BII/LC3BI ratio and the up-regulation of p62 expression in the heart induced by HFD feeding and restored cardiac autophagy signaling. In conclusion, CARD9 knockout ameliorates myocardial dysfunction associated with HFD-induced obesity, potentially through reduction of macrophage infiltration, suppression of p38 MAPK phosphorylation, and preservation of autophagy in the heart. Copyright © 2016 Elsevier Ltd. All rights reserved.

Changes in macrophage transcriptome associate with systemic sclerosis and mediate GSDMA contribution to disease risk

PubMed Central

Koturan, Surya; Ko, Jeong-Hun; Fonseca, Carmen; Harmston, Nathan; Game, Laurence; Martin, Javier; Ong, Voon; Abraham, David J; Denton, Christopher P; Behmoaras, Jacques; Petretto, Enrico

2018-01-01

Objectives Several common and rare risk variants have been reported for systemic sclerosis (SSc), but the effector cell(s) mediating the function of these genetic variants remains to be elucidated. While innate immune cells have been proposed as the critical targets to interfere with the disease process underlying SSc, no studies have comprehensively established their effector role. Here we investigated the contribution of monocyte-derived macrophages (MDMs) in mediating genetic susceptibility to SSc. Methods We carried out RNA sequencing and genome-wide genotyping in MDMs from 57 patients with SSc and 15 controls. Our differential expression and expression quantitative trait locus (eQTL) analysis in SSc was further integrated with epigenetic, expression and eQTL data from skin, monocytes, neutrophils and lymphocytes. Results We identified 602 genes upregulated and downregulated in SSc macrophages that were significantly enriched for genes previously implicated in SSc susceptibility (P=5×10−4), and 270 cis-regulated genes in MDMs. Among these, GSDMA was reported to carry an SSc risk variant (rs3894194) regulating expression of neighbouring genes in blood. We show that GSDMA is upregulated in SSc MDMs (P=8.4×10−4) but not in the skin, and is a significant eQTL in SSc macrophages and lipopolysaccharide/interferon gamma (IFNγ)-stimulated monocytes. Furthermore, we identify an SSc macrophage transcriptome signature characterised by upregulation of glycolysis, hypoxia and mTOR signalling and a downregulation of IFNγ response pathways. Conclusions Our data further establish the link between macrophages and SSc, and suggest that the contribution of the rs3894194 risk variant to SSc susceptibility can be mediated by GSDMA expression in macrophages. PMID:29348297

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

PubMed

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

2018-01-01

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

Interleukin-4 Is Essential for Microglia/Macrophage M2 Polarization and Long-Term Recovery After Cerebral Ischemia.

PubMed

Liu, Xiangrong; Liu, Jia; Zhao, Shangfeng; Zhang, Haiyue; Cai, Wei; Cai, Mengfei; Ji, Xunming; Leak, Rehana K; Gao, Yanqin; Chen, Jun; Hu, Xiaoming

2016-02-01

Interleukin-4 (IL-4) is a unique cytokine that may contribute to brain repair by regulating microglia/macrophage functions. Thus, we examined the effect of IL-4 on long-term recovery and microglia/macrophage polarization in 2 well-established stroke models. Transient middle cerebral artery occlusion or permanent distal middle cerebral artery occlusion was induced in wild-type and IL-4 knockout C57/BL6 mice. In a separate cohort of wild-type animals, IL-4 (60 ng/d for 7 days) or vehicle was infused into the cerebroventricle after transient middle cerebral artery occlusion. Behavioral outcomes were assessed by the Rotarod, corner, foot fault, and Morris water maze tests. Neuronal tissue loss was verified by 2 independent neuron markers. Markers of classically activated (M1) and alternatively activated (M2) microglia were assessed by real-time polymerase chain reaction, immunofluorescence, and flow cytometry. Loss of IL-4 exacerbated sensorimotor deficits and impaired cognitive functions ≤21 days post injury. In contrast to the delayed deterioration of neurological functions, IL-4 deficiency increased neuronal tissue loss only in the acute phase (5 days) after stroke and had no impact on neuronal tissue loss 14 or 21 days post injury. Loss of IL-4 promoted expression of M1 microglia/macrophage markers and impaired expression of M2 markers at 5 and 14 days post injury. Administration of IL-4 into the ischemic brain also enhanced long-term functional recovery. The cytokine IL-4 improves long-term neurological outcomes after stroke, perhaps through M2 phenotype induction in microglia/macrophages. These results are the first to suggest that immunomodulation with IL-4 is a promising approach to promote long-term functional recovery after stroke. © 2016 American Heart Association, Inc.

Kupffer cells express a unique combination of phenotypic and functional characteristics compared with splenic and peritoneal macrophages.

PubMed

Movita, Dowty; Kreefft, Kim; Biesta, Paula; van Oudenaren, Adri; Leenen, Pieter J M; Janssen, Harry L A; Boonstra, Andre

2012-10-01

The immunostimulatory role of Kupffer cells in various inflammatory liver diseases is still not fully understood. In this study, phenotypic and functional aspects of Kupffer cells from healthy C57BL/6 mice were analyzed and compared with those of splenic and peritoneal macrophages to generate a blueprint of the cells under steady-state conditions. In the mouse liver, only one population of Kupffer cells was identified as F4/80(high)CD11b(low) cells. We observed that freshy isolated Kupffer cells are endocytic and show a relatively high basal ROS content. Interestingly, despite expression of TLR mRNA on Kupffer cells, ligation of TLR4, TLR7/8, and TLR9 resulted in a weak induction of IL-10, low or undetectable levels of IL-12p40 and TNF, and up-regulation of CD40 on the surface. Kupffer cells and splenic macrophages show functional similarities, in comparison with peritoneal macrophages, as reflected by comparable levels of TLR4, TLR7/8, and TLR9 mRNA and low or undetectable levels of TNF and IL-12p40 produced upon TLR ligation. The unique, functional characteristics of Kupffer cells, demonstrated in this study, suggest that Kupffer cells under steady-state conditions are specialized as phagocytes to clear and degrade particulates and only play a limited immunoregulatory role via the release of soluble mediators.

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

PubMed Central

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

2017-01-01

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

Vascular endothelial growth factor modified macrophages transdifferentiate into endothelial-like cells and decrease foam cell formation.

PubMed

Yan, Dan; He, Yujuan; Dai, Jun; Yang, Lili; Wang, Xiaoyan; Ruan, Qiurong

2017-06-30

Macrophages are largely involved in the whole process of atherosclerosis from an initiation lesion to an advanced lesion. Endothelial disruption is the initial step and macrophage-derived foam cells are the hallmark of atherosclerosis. Promotion of vascular integrity and inhibition of foam cell formation are two important strategies for preventing atherosclerosis. How can we inhibit even the reverse negative role of macrophages in atherosclerosis? The present study was performed to investigate if overexpressing endogenous human vascular endothelial growth factor (VEGF) could facilitate transdifferentiation of macrophages into endothelial-like cells (ELCs) and inhibit foam cell formation. We demonstrated that VEGF-modified macrophages which stably overexpressed human VEGF (hVEGF 165 ) displayed a high capability to alter their phenotype and function into ELCs in vitro Exogenous VEGF could not replace endogenous VEGF to induce the transdifferentiation of macrophages into ELCs in vitro We further showed that VEGF-modified macrophages significantly decreased cytoplasmic lipid accumulation after treatment with oxidized LDL (ox-LDL). Moreover, down-regulation of CD36 expression in these cells was probably one of the mechanisms of reduction in foam cell formation. Our results provided the in vitro proof of VEGF-modified macrophages as atheroprotective therapeutic cells by both promotion of vascular repair and inhibition of foam cell formation. © 2017 The Author(s).

Mice Lacking Alternatively Activated (M2) Macrophages Show Impairments in Restorative Sleep after Sleep Loss and in Cold Environment.

PubMed

Massie, Ashley; Boland, Erin; Kapás, Levente; Szentirmai, Éva

2018-06-05

The relationship between sleep, metabolism and immune functions has been described, but the cellular components of the interaction are incompletely identified. We previously reported that systemic macrophage depletion results in sleep impairment after sleep loss and in cold environment. These findings point to the role of macrophage-derived signals in maintaining normal sleep. Macrophages exist either in resting form, classically activated, pro-inflammatory (M1) or alternatively activated, anti-inflammatory (M2) phenotypes. In the present study we determined the contribution of M2 macrophages to sleep signaling by using IL-4 receptor α-chain-deficient [IL-4Rα knockout (KO)] mice, which are unable to produce M2 macrophages. Sleep deprivation induced robust increases in non-rapid-eye-movement sleep (NREMS) and slow-wave activity in wild-type (WT) animals. NREMS rebound after sleep deprivation was ~50% less in IL-4Rα KO mice. Cold exposure induced reductions in rapid-eye-movement sleep (REMS) and NREMS in both WT and KO mice. These differences were augmented in IL-4Rα KO mice, which lost ~100% more NREMS and ~25% more REMS compared to WTs. Our finding that M2 macrophage-deficient mice have the same sleep phenotype as mice with global macrophage depletion reconfirms the significance of macrophages in sleep regulation and suggests that the main contributors are the alternatively activated M2 cells.

Substrate Stiffness Regulates Proinflammatory Mediator Production through TLR4 Activity in Macrophages

PubMed Central

Previtera, Michelle L.; Sengupta, Amitabha

2015-01-01

Clinical data show that disease adversely affects tissue elasticity or stiffness. While macrophage activity plays a critical role in driving disease pathology, there are limited data available on the effects of tissue stiffness on macrophage activity. In this study, the effects of substrate stiffness on inflammatory mediator production by macrophages were investigated. Bone marrow–derived macrophages were grown on polyacrylamide gels that mimicked the stiffness of a variety of soft biological tissues. Overall, macrophages grown on soft substrates produced less proinflammatory mediators than macrophages grown on stiff substrates when the endotoxin LPS was added to media. In addition, the pathways involved in stiffness–regulated proinflammation were investigated. The TLR4 signaling pathway was examined by evaluating TLR4, p–NF–κB p65, MyD88, and p–IκBα expression as well as p–NF–κB p65 translocation. Expression and translocation of the various signaling molecules were higher in macrophages grown on stiff substrates than on soft substrates. Furthermore, TLR4 knockout experiments showed that TLR4 activity enhanced proinflammation on stiff substrates. In conclusion, these results suggest that proinflammatory mediator production initiated by TLR4 is mechanically regulated in macrophages. PMID:26710072

Nicotine can skew the characterization of the macrophage type-1 (M{Phi}1) phenotype differentiated with granulocyte-macrophage colony-stimulating factor to the M{Phi}2 phenotype

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

Yanagita, Manabu; Kobayashi, Ryohei; Murakami, Shinya, E-mail: ipshinya@dent.osaka-u.ac.jp

Macrophages (M{Phi}s) exhibit functional heterogeneity and plasticity in the local microenvironment. Recently, it was reported that M{Phi}s can be divided into proinflammatory M{Phi}s (M{Phi}1) and anti-inflammatory M{Phi}s (M{Phi}2) based on their polarized functional properties. Here, we report that nicotine, the major ingredient of cigarette smoke, can modulate the characteristics of M{Phi}1. Granulocyte-macrophage colony-stimulating factor-driven M{Phi}1 with nicotine (Ni-M{Phi}1) showed the phenotypic characteristics of M{Phi}2. Like M{Phi}2, Ni-M{Phi}1 exhibited antigen-uptake activities. Ni-M{Phi}1 suppressed IL-12, but maintained IL-10 and produced high amounts of MCP-1 upon lipopolysaccharide stimulation compared with M{Phi}1. Moreover, we observed strong proliferative responses of T cells to lipopolysaccharide-stimulated M{Phi}1,more » whereas Ni-M{Phi}1 reduced T cell proliferation and inhibited IFN-{gamma} production by T cells. These results suggest that nicotine can change the functional characteristics of M{Phi} and skew the M{Phi}1 phenotype to M{Phi}2. We propose that nicotine is a potent regulator that modulates immune responses in microenvironments.« less

Recruiting specialized macrophages across the borders to restore brain functions.

PubMed

Corraliza, Inés

2014-01-01

Although is well accepted that the central nervous system has an immune privilege protected by the blood-brain barrier (BBB) and maintained by the glia, it is also known that in homeostatic conditions, peripheral immune cells are able to penetrate to the deepest regions of brain without altering the structural integrity of the BBB. Nearly all neurological diseases, including degenerative, autoimmune or infectious ones, compromising brain functions, develop with a common pattern of inflammation in which macrophages and microglia activation have been regarded often as the "bad guys." However, recognizing the huge heterogeneity of macrophage populations and also the different expression properties of microglia, there is increasing evidence of alternative conditions in which these cells, if primed and addressed in the correct direction, could be essential for reparative and regenerative functions. The main proposal of this review is to integrate studies about macrophage's biology at the brain borders where the ultimate challenge is to penetrate through the BBB and contribute to change or even stop the course of disease. Thanks to the efforts made in the last century, this special wall is currently recognized as a highly regulated cooperative structure, in which their components form neurovascular units. This new scenario prompted us to review the precise cross-talk between the mind and body modes of immune response.

A WAVE2-Abi1 complex mediates CSF-1-induced F-actin-rich membrane protrusions and migration in macrophages.

PubMed

Kheir, Wassim Abou; Gevrey, Jean-Claude; Yamaguchi, Hideki; Isaac, Beth; Cox, Dianne

2005-11-15

Colony-stimulating factor 1 (CSF-1) is an important physiological chemoattractant for macrophages. The mechanisms by which CSF-1 elicits the formation of filamentous actin (F-actin)-rich membrane protrusions and induces macrophage migration are not fully understood. In particular, very little is known regarding the contribution of the different members of the Wiskott-Aldrich Syndrome protein (WASP) family of actin regulators in response to CSF-1. Although a role for WASP itself in macrophage chemotaxis has been previously identified, no data was available regarding the function of WASP family verprolin-homologous (WAVE) proteins in this cell type. We found that WAVE2 was the predominant isoform to be expressed in primary macrophages and in cells derived from the murine monocyte/macrophage RAW264.7 cell line (RAW/LR5). CSF-1 treatment of macrophages resulted in WAVE2 accumulation in F-actin-rich protrusions induced by CSF-1. Inhibition of WAVE2 function by expressing a dominant-negative mutant or introducing anti-WAVE2 antibodies in RAW/LR5 cells, as well as reduction of endogenous WAVE2 expression by RNA-mediated interference (RNAi), resulted in a significant reduction of CSF-1-elicited F-actin protrusions. WAVE2 was found in a protein complex together with Abelson kinase interactor 1 (Abi1) in resting or stimulated cells. Both WAVE2 and Abi1 were recruited to and necessary for the formation of F-actin protrusions in response to CSF-1. Reducing the levels of WAVE2, directly or by targeting Abi1, resulted in an impaired cell migration to CSF-1. Altogether these data identify a WAVE2-Abi1 complex crucial for the normal actin cytoskeleton reorganization and migration of macrophages in response to CSF-1.

Atypical chemokine receptor ACKR2 controls branching morphogenesis in the developing mammary gland

PubMed Central

Hewit, Kay D.; Pallas, Kenneth J.; Cairney, Claire J.; Lee, Kit M.; Hansell, Christopher A.; Stein, Torsten

2017-01-01

Macrophages are important regulators of branching morphogenesis during development and postnatally in the mammary gland. Regulation of macrophage dynamics during these processes can therefore have a profound impact on development. We demonstrate here that the developing mammary gland expresses high levels of inflammatory CC-chemokines, which are essential in vivo regulators of macrophage migration. We further demonstrate that the atypical chemokine receptor ACKR2, which scavenges inflammatory CC-chemokines, is differentially expressed during mammary gland development. We have previously shown that ACKR2 regulates macrophage dynamics during lymphatic vessel development. Here, we extend these observations to reveal a novel role for ACKR2 in regulating the postnatal development of the mammary gland. Specifically, we show that Ackr2−/− mice display precocious mammary gland development. This is associated with increased macrophage recruitment to the developing gland and increased density of the ductal epithelial network. These data demonstrate that ACKR2 is an important regulator of branching morphogenesis in diverse biological contexts and provide the first evidence of a role for chemokines and their receptors in postnatal development processes. PMID:27888192

Macrophage uptake and accumulation of folates are polarization-dependent in vitro and in vivo and are regulated by activin A.

PubMed

Samaniego, Rafael; Palacios, Blanca Soler; Domiguez-Soto, Ángeles; Vidal, Carlos; Salas, Azucena; Matsuyama, Takami; Sánchez-Torres, Carmen; de la Torre, Inmaculada; Miranda-Carús, Maria Eugenia; Sánchez-Mateos, Paloma; Puig-Kröger, Amaya

2014-05-01

Vitamin B9, commonly known as folate, is an essential cofactor for one-carbon metabolism that enters cells through three major specialized transporter molecules (RFC, FR, and PCFT), which differ in expression pattern, affinity for substrate, and ligand-binding pH dependency. We now report that the expression of the folate transporters differs between macrophage subtypes and explains the higher accumulation of 5-MTHF-the major folate form found in serum-in M2 macrophages in vitro and in vivo. M1 macrophages display a higher expression of RFC, whereas FRβ and PCFT are preferentially expressed by anti-inflammatory and homeostatic M2 macrophages. These differences are also seen in macrophages from normal tissues involved in folate transit (placenta, liver, colon) and inflamed tissues (ulcerative colitis, RA), as M2-like macrophages from normal tissues express FRβ and PCFT, whereas TNF-α-expressing M1 macrophages from inflamed tissues are RFC+. Besides, we provide evidences that activin A is a critical factor controlling the set of folate transporters in macrophages, as it down-regulates FRβ, up-regulates RFC expression, and modulates 5-MTHF uptake. All of these experiments support the notion that folate handling is dependent on the stage of macrophage polarization. © 2014 Society for Leukocyte Biology.

Akt3 kinase suppresses pinocytosis of low-density lipoprotein by macrophages via a novel WNK/SGK1/Cdc42 protein pathway

PubMed Central

Ding, Liang; Zhang, Lifang; Kim, Michael; Byzova, Tatiana; Podrez, Eugene

2017-01-01

Fluid-phase pinocytosis of LDL by macrophages is regarded as a novel promising target to reduce macrophage cholesterol accumulation in atherosclerotic lesions. The mechanisms of regulation of fluid-phase pinocytosis in macrophages and, specifically, the role of Akt kinases are poorly understood. We have found previously that increased lipoprotein uptake via the receptor-independent process in Akt3 kinase-deficient macrophages contributes to increased atherosclerosis in Akt3−/− mice. The mechanism by which Akt3 deficiency promotes lipoprotein uptake in macrophages is unknown. We now report that Akt3 constitutively suppresses macropinocytosis in macrophages through a novel WNK1/SGK1/Cdc42 pathway. Mechanistic studies have demonstrated that the lack of Akt3 expression in murine and human macrophages results in increased expression of with-no-lysine kinase 1 (WNK1), which, in turn, leads to increased activity of serum and glucocorticoid-inducible kinase 1 (SGK1). SGK1 promotes expression of the Rho family GTPase Cdc42, a positive regulator of actin assembly, cell polarization, and pinocytosis. Individual suppression of WNK1 expression, SGK1, or Cdc42 activity in Akt3-deficient macrophages rescued the phenotype. These results demonstrate that Akt3 is a specific negative regulator of macropinocytosis in macrophages. PMID:28389565

Kcnn4 Is a Regulator of Macrophage Multinucleation in Bone Homeostasis and Inflammatory Disease

PubMed Central

Kang, Heeseog; Kerloc’h, Audrey; Rotival, Maxime; Xu, Xiaoqing; Zhang, Qing; D’Souza, Zelpha; Kim, Michael; Scholz, Jodi Carlson; Ko, Jeong-Hun; Srivastava, Prashant K.; Genzen, Jonathan R.; Cui, Weiguo; Aitman, Timothy J.; Game, Laurence; Melvin, James E.; Hanidu, Adedayo; Dimock, Janice; Zheng, Jie; Souza, Donald; Behera, Aruna K.; Nabozny, Gerald; Cook, H. Terence; Bassett, J.H. Duncan; Williams, Graham R.; Li, Jun; Vignery, Agnès; Petretto, Enrico; Behmoaras, Jacques

2014-01-01

Summary Macrophages can fuse to form osteoclasts in bone or multinucleate giant cells (MGCs) as part of the immune response. We use a systems genetics approach in rat macrophages to unravel their genetic determinants of multinucleation and investigate their role in both bone homeostasis and inflammatory disease. We identify a trans-regulated gene network associated with macrophage multinucleation and Kcnn4 as being the most significantly trans-regulated gene in the network and induced at the onset of fusion. Kcnn4 is required for osteoclast and MGC formation in rodents and humans. Genetic deletion of Kcnn4 reduces macrophage multinucleation through modulation of Ca2+ signaling, increases bone mass, and improves clinical outcome in arthritis. Pharmacological blockade of Kcnn4 reduces experimental glomerulonephritis. Our data implicate Kcnn4 in macrophage multinucleation, identifying it as a potential therapeutic target for inhibition of bone resorption and chronic inflammation. PMID:25131209

The E3 ubiquitin ligase, HECTD1, is involved in ABCA1-mediated cholesterol export from macrophages.

PubMed

Aleidi, Shereen M; Yang, Alryel; Sharpe, Laura J; Rao, Geetha; Cochran, Blake J; Rye, Kerry-Anne; Kockx, Maaike; Brown, Andrew J; Gelissen, Ingrid C

2018-04-01

The ABC lipid transporters, ABCA1 and ABCG1, are essential for maintaining lipid homeostasis in cells such as macrophages by exporting excess cholesterol to extracellular acceptors. These transporters are highly regulated at the post-translational level, including protein ubiquitination. Our aim was to investigate the role of the E3 ubiquitin ligase HECTD1, recently identified as associated with ABCG1, on ABCG1 and ABCA1 protein levels and cholesterol export function. Here, we show that HECTD1 protein is widely expressed in a range of human and murine primary cells and cell lines, including macrophages, neuronal cells and insulin secreting β-cells. siRNA knockdown of HECTD1 unexpectedly decreased overexpressed ABCG1 protein levels and cell growth, but increased native ABCA1 protein in CHO-K1 cells. Knockdown of HECTD1 in unloaded THP-1 macrophages did not affect ABCG1 but significantly increased ABCA1 protein levels, in wild-type as well as THP-1 cells that do not express ABCG1. Cholesterol export from macrophages to apoA-I over time was increased after knockdown of HECTD1, however these effects were not sustained in cholesterol-loaded cells. In conclusion, we have identified a new candidate, the E3 ubiquitin ligase HECTD1, that may be involved in the regulation of ABCA1-mediated cholesterol export from unloaded macrophages to apoA-I. The exact mechanism by which this ligase affects this pathway remains to be elucidated. Copyright © 2018 Elsevier B.V. All rights reserved.

Cot/Tpl2 regulates IL-23 p19 expression in LPS-stimulated macrophages through ERK activation.

PubMed

Kakimoto, K; Musikacharoen, T; Chiba, N; Bandow, K; Ohnishi, T; Matsuguchi, T

2010-03-01

We have previously reported that a serine/threonine protein kinase, Cot/Tpl2, is a negative regulator of Th1-type immunity through inhibiting IL-12 expression in antigen presenting cells (APCs) stimulated by Toll-like receptor (TLR) ligands. We here show that Cot/Tpl2(-/-) macrophages produce significantly less IL-23, an important regulator of Th17-type response, than the wild-type counterparts in response to lipopolysaccharide (LPS), which is a ligand for TLR4. The decreased IL-23 production in Cot/Tpl2(-/-) macrophages is, at least partly, regulated at the transcriptional level, as the LPS-mediated IL-23 p19 mRNA induction was significantly less in Cot/Tpl2(-/-) macrophages. Chemical inhibition of extracellular signal-regulated kinase (ERK) activity similarly inhibited IL-23 expression in LPS-stimulated wild-type macrophages. As Cot/Tpl2 is an essential upstream component of the ERK activation pathway of LPS, it is suggested that Cot/Tpl2 positively regulates IL-23 expression through ERK activation. These results indicate that Cot/Tpl2 may be involved in balancing Th1/Th17 differentiation by regulating the expression ratio of IL-12 and IL-23 in APCs.

Role of the testis interstitial compartment in spermatogonial stem cell function

PubMed Central

Potter, Sarah J.; DeFalco, Tony

2017-01-01

Male fertility is maintained through intricate cellular and molecular interactions that ensure spermatogonial stem cells (SSCs) proceed in a step-wise differentiation process through spermatogenesis and spermiogenesis to produce sperm. SSCs lie within the seminiferous tubule compartment, which provides a nurturing environment for the development of sperm. Cells outside of the tubules, such as interstitial and peritubular cells, also help direct SSC activity. This review focuses on interstitial (interstitial macrophages, Leydig cells, and vasculature) and peritubular (peritubular macrophages, peritubular myoid cells) cells and their role in regulating SSC self-renewal and differentiation in mammals. Leydig cells, the major steroidogenic cells in the testis, influence SSCs through secreted factors, such as insulin growth factor 1 (IGF1) and colony stimulating factor 1 (CSF1). Macrophages interact with SSCs through various potential mechanisms, such as CSF1 and retinoic acid (RA), to induce proliferation or differentiation of SSCs, respectively. Vasculature influences SSC dynamics through CSF1, vascular endothelial growth factor (VEGF), and regulating oxygen levels. Lastly, peritubular myoid cells produce one of the most well-known factors that is required for SSC self-renewal, glial cell line derived neurotrophic factor (GDNF), as well as CSF1. Overall, SSC interactions with interstitial and peritubular cells are critical for SSC function and are an important underlying factor promoting male fertility. PMID:28115580

Intracellular degradation of functionalized carbon nanotube/iron oxide hybrids is modulated by iron via Nrf2 pathway

PubMed Central

Elgrabli, Dan; Dachraoui, Walid; Marmier, Hélène de; Ménard-Moyon, Cécilia; Bégin, Dominique; Bégin-Colin, Sylvie; Bianco, Alberto; Alloyeau, Damien; Gazeau, Florence

2017-01-01

The in vivo fate and biodegradability of carbon nanotubes is still a matter of debate despite tremendous applications. In this paper we describe a molecular pathway by which macrophages degrade functionalized multi-walled carbon nanotubes (CNTs) designed for biomedical applications and containing, or not, iron oxide nanoparticles in their inner cavity. Electron microscopy and Raman spectroscopy show that intracellularly-induced structural damages appear more rapidly for iron-free CNTs in comparison to iron-loaded ones, suggesting a role of iron in the degradation mechanism. By comparing the molecular responses of macrophages derived from THP1 monocytes to both types of CNTs, we highlight a molecular mechanism regulated by Nrf2/Bach1 signaling pathways to induce CNT degradation via NOX2 complex activation and O2•−, H2O2 and OH• production. CNT exposure activates an oxidative stress-dependent production of iron via Nrf2 nuclear translocation, Ferritin H and Heme oxygenase 1 translation. Conversely, Bach1 was translocated to the nucleus of cells exposed to iron-loaded CNTs to recycle embedded iron. Our results provide new information on the role of oxidative stress, iron metabolism and Nrf2-mediated host defence for regulating CNT fate in macrophages. PMID:28120861

Macrophages Inhibit Neovascularization in a Murine Model of Age-Related Macular Degeneration

PubMed Central

Apte, Rajendra S; Richter, Jennifer; Herndon, John; Ferguson, Thomas A

2006-01-01

Background Age-related macular degeneration (AMD) is the leading cause of blindness in people over 50 y of age in at least three continents. Choroidal neovascularization (CNV) is the process by which abnormal blood vessels develop underneath the retina. CNV develops in 10% of patients with AMD but accounts for up to 90% of the blindness from AMD. Although the precise etiology of CNV in AMD remains unknown, the macrophage component of the inflammatory response, which has been shown to promote tumor growth and support atherosclerotic plaque formation, is thought to stimulate aberrant angiogenesis in blinding eye diseases. The current theory is that macrophage infiltration promotes the development of neovascularization in CNV. Methods and Findings We examined the role of macrophages in a mouse model of CNV. IL-10 −/− mice, which have increased inflammation in response to diverse stimuli, have significantly reduced CNV with increased macrophage infiltrates compared to wild type. Prevention of macrophage entry into the eye promoted neovascularization while direct injection of macrophages significantly inhibited CNV. Inhibition by macrophages was mediated by the TNF family death molecule Fas ligand (CD95-ligand). Conclusions Immune vascular interactions can be highly complex. Normal macrophage function is critical in controlling pathologic neovascularization in the eye. IL-10 regulates macrophage activity in the eye and is an attractive therapeutic target in order to suppress or inhibit CNV in AMD that can otherwise lead to blindness. PMID:16903779

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

PubMed

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

2016-08-01

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

ROS is Required for Alternatively Activated Macrophage Differentiation | Center for Cancer Research

Cancer.gov

Macrophages are key regulators in host inflammatory responses. Granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) are responsible for inducing macrophage differentiation from monocytes. GM-CSF or M-CSF-differentiated macrophages can be further differentiated, or polarized, to more specialized cells. Classically activated,

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

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

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

Highlights: Black-Right-Pointing-Pointer Rosiglitazone attenuated postincisional pain. Black-Right-Pointing-Pointer Rosiglitazone alters macrophage polarization to F4/80{sup +}CD206{sup +} M2 macrophages at the incisional sites. Black-Right-Pointing-Pointer Transplantation of rosiglitazone-treated macrophages produced analgesic effects. -- Abstract: Acute inflammation triggered by macrophage infiltration to injured tissue promotes wound repair and may induce pain hypersensitivity. Peroxisome proliferator-activated receptor {gamma} (PPAR){gamma} signaling is known to regulate heterogeneity of macrophages, which are often referred to as classically activated (M1) and alternatively activated (M2) macrophages. M1 macrophages have considerable antimicrobial activity and produce a wide variety of proinflammatory cytokines. In contrast, M2 macrophages are involved in anti-inflammatory and homeostatic functionsmore » linked to wound healing and tissue repair. Although it has been suggested that PPAR{gamma} agonists attenuate pain hypersensitivity, the molecular mechanism of macrophage-mediated effects of PPAR{gamma} signaling on pain development has not been explored. In this study, we investigated the link between the phenotype switching of macrophage polarization induced by PPAR{gamma} signaling and the development of acute pain hypersensitivity. Local administration of rosiglitazone significantly ameliorated hypersensitivity to heat and mechanical stimuli, and paw swelling. Consistent with the down-regulation of nuclear factor {kappa}B (NF{kappa}B) phosphorylation by rosiglitazone at the incisional sites, the number of F4/80{sup +}iNOS{sup +} M1 macrophages was decreased whereas numbers of F4/80{sup +}CD206{sup +} M2 macrophages were increased in rosiglitazone-treated incisional sites 24 h after the procedure. In addition, gene induction of anti-inflammatory M2-macrophage-associated markers such as arginase1, FIZZ1 and interleukin (IL)-10 were significantly increased, whereas M1-macrophage-related molecules such as integrin {alpha}X, IL-1{beta}, MIP2{alpha} and leptin were decreased at rosiglitazone-treated incisional sites. Moreover, transplantation of rosiglitazone-treated peritoneal macrophages into the incisional sites significantly attenuated hyperalgesia. We speculate that local administration of rosiglitazone significantly alleviated the development of postincisional pain, possibly through regulating macrophage polarity at the inflamed site. PPAR{gamma} signaling in macrophages may be a potential therapeutic target for the treatment of acute pain development.« less

IL-4 and IL-13 employ discrete signaling pathways for target gene expression in alternatively activated monocytes/macrophages.

PubMed

Bhattacharjee, Ashish; Shukla, Meenakshi; Yakubenko, Valentin P; Mulya, Anny; Kundu, Suman; Cathcart, Martha K

2013-01-01

Monocytes/macrophages are innate immune cells that play a crucial role in the resolution of inflammation. In the presence of the Th2 cytokines interleukin-4 (IL-4) and interleukin-13 (IL-13), they display an anti-inflammatory profile and this activation pathway is known as alternative activation. In this study we compare and differentiate pathways mediated by IL-4 and IL-13 activation of human monocytes/macrophages. Here we report differential regulation of IL-4 and IL-13 signaling in monocytes/macrophages starting from IL-4/IL-13 cytokine receptors to Jak/Stat-mediated signaling pathways that ultimately control expression of several inflammatory genes. Our data demonstrate that although the receptor-associated tyrosine kinases Jak2 and Tyk2 are activated after the recruitment of IL-13 to its receptor (containing IL-4Rα and IL-13Rα1), IL-4 stimulates Jak1 activation. We further show that Jak2 is upstream of Stat3 activation and Tyk2 controls Stat1 and Stat6 activation in response to IL-13 stimulation. In contrast, Jak1 regulates Stat3 and Stat6 activation in IL-4-induced monocytes. Our results further reveal that although IL-13 utilizes both IL-4Rα/Jak2/Stat3 and IL-13Rα1/Tyk2/Stat1/Stat6 signaling pathways, IL-4 can use only the IL-4Rα/Jak1/Stat3/Stat6 cascade to regulate the expression of some critical inflammatory genes, including 15-lipoxygenase, monoamine oxidase A (MAO-A), and the scavenger receptor CD36. Moreover, we demonstrate here that IL-13 and IL-4 can uniquely affect the expression of particular genes such as dual-specificity phosphatase 1 and tissue inhibitor of metalloprotease-3 and do so through different Jaks. As evidence of differential regulation of gene function by IL-4 and IL-13, we further report that MAO-A-mediated reactive oxygen species generation is influenced by different Jaks. Collectively, these results have major implications for understanding the mechanism and function of alternatively activated monocytes/macrophages by IL-4 and IL-13 and add novel insights into the pathogenesis and potential treatment of various inflammatory diseases. Copyright © 2012 Elsevier Inc. All rights reserved.

Testosterone regulates 3T3-L1 pre-adipocyte differentiation and epididymal fat accumulation in mice through modulating macrophage polarization.

PubMed

Ren, Xiaojiao; Fu, Xiaojian; Zhang, Xinhua; Chen, Shiqiang; Huang, Shuguang; Yao, Lun; Liu, Guoquan

2017-09-15

Low testosterone levels are strongly related to obesity in males. The balance between the classically M1 and alternatively M2 polarized macrophages also plays a critical role in obesity. It is not clear whether testosterone regulates macrophage polarization and then affects adipocyte differentiation. In this report, we demonstrate that testosterone strengthens interleukin (IL) -4-induced M2 polarization and inhibits lipopolysaccharide (LPS)-induced M1 polarization, but has no direct effect on adipocyte differentiation. Cellular signaling studies indicate that testosterone regulates macrophage polarization through the inhibitory regulative G-protein (Gαi) mainly, rather than via androgen receptors, and phosphorylation of Akt. Moreover, testosterone inhibits pre-adipocyte differentiation induced by M1 macrophage medium. Lowering of serum testosterone in mice by injecting a luteinizing hormone receptor (LHR) peptide increases epididymal white adipose tissue. Testosterone supplementation reverses this effect. Therefore, our findings indicate that testosterone inhibits pre-adipocyte differentiation by switching macrophages to M2 polarization through the Gαi and Akt signaling pathways. Copyright © 2017 Elsevier Inc. All rights reserved.

CSF-1 Receptor Signaling in Myeloid Cells

PubMed Central

Stanley, E. Richard; Chitu, Violeta

2014-01-01

The CSF-1 receptor (CSF-1R) is activated by the homodimeric growth factors colony-stimulating factor-1 (CSF-1) and interleukin-34 (IL-34). It plays important roles in development and in innate immunity by regulating the development of most tissue macrophages and osteoclasts, of Langerhans cells of the skin, of Paneth cells of the small intestine, and of brain microglia. It also regulates the differentiation of neural progenitor cells and controls functions of oocytes and trophoblastic cells in the female reproductive tract. Owing to this broad tissue expression pattern, it plays a central role in neoplastic, inflammatory, and neurological diseases. In this review we summarize the evolution, structure, and regulation of expression of the CSF-1R gene. We review, the structures of CSF-1, IL-34, and the CSF-1R and the mechanism of ligand binding to and activation of the receptor. We further describe the pathways regulating macrophage survival, proliferation, differentiation, and chemotaxis downstream from the CSF-1R. PMID:24890514

Sympathetic innervation regulates macrophage activity in rats with polycystic ovary.

PubMed

Figueroa, Florencia; Mendoza, Gisela; Cardozo, Darío; Mohamed, Fabián; Oliveros, Liliana; Forneris, Myriam

2018-07-01

Polycystic ovarian syndrome (PCOS) is a low-grade inflammatory disease characterized by hyperandrogenism and ovarian hyperinnervation. The aim of this work is to investigate whether in vivo bilateral superior ovarian nerve (SON) section in adult rats with estradiol valerate-induced PCOS (PCO rats) affects macrophage spleen cells (MФ) and modifies the steroidogenic ability of their secretions. Culture media of MФ from PCO rats and PCO rats with SON section (PCO-SON rats) were used to stimulate in vitro intact ovaries. Compared with macrophages PCO, macrophages from PCO-SON rats released less tumor necrosis factor-α and nitric oxide, expressed lower Bax and Nfkb mRNA and showed reduced TUNEL staining. Also, in PCO rats, the SON section decreased kisspeptin and nerve growth factor mRNA expressions, without changes in Trka receptor mRNA levels. Macrophage secretions from PCO-SON rats decreased androstenedione and stimulated progesterone release in PCO ovaries, compared to macrophage secretions from PCO rats. No changes were observed in ovarian estradiol response. These findings emphasize the importance of the SON in spleen MΦ, since its manipulation leads to secondary modifications of immunological and neural mediators, which might influence ovarian steroidogenesis. In PCO ovaries, the reduction of androstenedione and the improvement of progesterone release induced by PCO-SON MΦ secretion, might be beneficial considering the hormonal anomalies characteristic of PCOS. We present functional evidence that modulation of the immune-endocrine function by peripheral sympathetic nervous system might have implications for understanding the pathophysiology of PCOS. © 2018 Society for Endocrinology.

Nuclear DAMP complex-mediated RAGE-dependent macrophage cell death

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

Chen, Ruochan; Department of Infectious Diseases and State Key Lab of Viral Hepatitis, Xiangya Hospital, Central South University, Changsha, Hunan 410008; Fu, Sha

High mobility group box 1 (HMGB1), histone, and DNA are essential nuclear components involved in the regulation of chromosome structure and function. In addition to their nuclear function, these molecules act as damage-associated molecular patterns (DAMPs) alone or together when released extracellularly. The synergistic effect of these nuclear DNA-HMGB1-histone complexes as DAMP complexes (nDCs) on immune cells remains largely unexplored. Here, we demonstrate that nDCs limit survival of macrophages (e.g., RAW264.7 and peritoneal macrophages) but not cancer cells (e.g., HCT116, HepG2 and Hepa1-6). nDCs promote production of inflammatory tumor necrosis factor α (TNFα) release, triggering reactive oxygen species-dependent apoptosis andmore » necrosis. Moreover, the receptor for advanced glycation end products (RAGE), but not toll-like receptor (TLR)-4 and TLR-2, was required for Akt-dependent TNFα release and subsequent cell death following treatment with nDCs. Genetic depletion of RAGE by RNAi, antioxidant N-Acetyl-L-cysteine, and TNFα neutralizing antibody significantly attenuated nDC-induced cell death. These findings provide evidence supporting novel signaling mechanisms linking nDCs and inflammation in macrophage cell death. - Highlights: • Nuclear DAMP complexes (nDCs) selectively induce cell death in macrophages, but not cancer cells. • TNFα-mediated oxidative stress is required for nDC-induced death. • RAGE-mediated Akt activation is required for nDC-induced TNFα release. • Blocking RAGE and TNFα inhibits nDC-induced macrophage cell death.« less

Macrophage arginase-1 controls bacterial growth and pathology in hypoxic tuberculosis granulomas

PubMed Central

Duque-Correa, María A.; Kühl, Anja A.; Rodriguez, Paulo C.; Zedler, Ulrike; Schommer-Leitner, Sandra; Rao, Martin; Weiner, January; Hurwitz, Robert; Qualls, Joseph E.; Kosmiadi, George A.; Murray, Peter J.; Kaufmann, Stefan H. E.; Reece, Stephen T.

2014-01-01

Lung granulomas develop upon Mycobacterium tuberculosis (Mtb) infection as a hallmark of human tuberculosis (TB). They are structured aggregates consisting mainly of Mtb-infected and -uninfected macrophages and Mtb-specific T cells. The production of NO by granuloma macrophages expressing nitric oxide synthase-2 (NOS2) via l-arginine and oxygen is a key protective mechanism against mycobacteria. Despite this protection, TB granulomas are often hypoxic, and bacterial killing via NOS2 in these conditions is likely suboptimal. Arginase-1 (Arg1) also metabolizes l-arginine but does not require oxygen as a substrate and has been shown to regulate NOS2 via substrate competition. However, in other infectious diseases in which granulomas occur, such as leishmaniasis and schistosomiasis, Arg1 plays additional roles such as T-cell regulation and tissue repair that are independent of NOS2 suppression. To address whether Arg1 could perform similar functions in hypoxic regions of TB granulomas, we used a TB murine granuloma model in which NOS2 is absent. Abrogation of Arg1 expression in macrophages in this setting resulted in exacerbated lung granuloma pathology and bacterial burden. Arg1 expression in hypoxic granuloma regions correlated with decreased T-cell proliferation, suggesting that Arg1 regulation of T-cell immunity is involved in disease control. Our data argue that Arg1 plays a central role in the control of TB when NOS2 is rendered ineffective by hypoxia. PMID:25201986

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

PubMed Central

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

2013-01-01

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

Avian macrophage: effector functions in health and disease.

PubMed

Qureshi, M A; Heggen, C L; Hussain, I

2000-01-01

Monocytes-macrophages, cells belonging to the mononuclear phagocytic system, are considered as the first line of immunological defense. Being mobile scavenger cells, macrophages participate in innate immunity by serving as phagocytic cells. These cells arise in the bone marrow and subsequently enter the blood circulation as blood monocytes. Upon migration to various tissues, monocytes mature and differentiate into tissue macrophages. Macrophages then initiate the 'acquired' immune response in their capacity as antigen processing and presenting cells. While responding to their tissue microenvironment or exogenous antigenic challenge, macrophages may secrete several immunoregulatory cytokines or metabolites. Being the first line of immunological defense, macrophages therefore represent an important step during interaction with infectious agents. The outcome of the macrophage-pathogen interaction depends upon several factors including the stage of macrophage activation, the nature of the infectious agent, the level of genetic control on macrophage function as well as environmental and nutritional factors that may modulate macrophage activation and functions. Research in avian macrophages has lagged behind that in mammals. This has been largely due to the lack of harvestable resident macrophages from the chicken peritoneal cavity. However, the development of elicitation protocols to harvest inflammatory abdominal macrophages and the availability of transformed chicken macrophage cell lines, has enabled researchers to address several questions related to chicken macrophage biology and function in health and disease. In this manuscript the basic profiles of several macrophage effector functions are described. In addition, the interaction of macrophages with various pathogens as well as the effect of genetic and environmental factors on macrophage functional modulation is described.

Involvement of specific macrophage-lineage cells surrounding arterioles in barrier and scavenger function in brain cortex.

PubMed Central

Mato, M; Ookawara, S; Sakamoto, A; Aikawa, E; Ogawa, T; Mitsuhashi, U; Masuzawa, T; Suzuki, H; Honda, M; Yazaki, Y; Watanabe, E; Luoma, J; Yla-Herttuala, S; Fraser, I; Gordon, S; Kodama, T

1996-01-01

The transport of solutes between blood and brain is regulated by a specific barrier. Capillary endothelial cells of brain are known to mediate barrier function and facilitate transport. Here we report that specific cells surrounding arterioles, known as Mato's fluorescent granular perithelial (FGP) cells or perivascular microglial cells, contribute to the barrier function. Immunohistochemical and in situ hybridization studies indicate that, in normal brain cortex, type I and type II macrophage scavenger receptors are expressed only in FGP/perivascular microglial cells, and surface markers of macrophage lineage are also detected on them. These cells mediate the uptake of macromolecules, including modified low density lipoprotein, horseradish peroxidase, and ferritin injected either into the blood or into the cerebral ventricles. Accumulation of scavenged materials with aging or after the administration of a high-fat diet results in the formation of honeycomb-like foam cells and the narrowing of the lumen of arterioles in the brain cortex. These results indicate involvement of FGP/perivascular microglial cells in the barrier and scavenger functions in the central nervous system. Images Fig. 1 Fig. 2 Fig. 4 Fig. 5 Fig. 6 PMID:8622926

Ski can negatively regulates macrophage differentiation through its interaction with PU.1

PubMed Central

Ueki, N; Zhang, L; Haymann, MJ

2010-01-01

In the hematopoietic cell system, the oncoprotein Ski dramatically affects growth and differentiation programs, in some cases leading to malignant leukemia. However, little is known about the interaction partners or signaling pathways involved in the Ski-mediated block of differentiation in hematopoietic cells. Here we show that Ski interacts with PU.1, a lineage-specific transcription factor essential for terminal myeloid differentiation, and thereby represses PU.1-dependent transcriptional activation. Consistent with this, Ski inhibits the biological function of PU.1 to promote myeloid cells to differentiate into macrophage colony-stimulating factor receptor (M-CSFR)-positive macrophages. Using a Ski mutant deficient in PU.1 binding, we demonstrate that Ski–PU.1 interaction is critical for Ski's ability to repress PU.1-dependent transcription and block macrophage differentiation. Furthermore, we provide evidence that Ski-mediated repression of PU.1 is due to Ski's ability to recruit histone deacetylase 3 to PU.1 bound to DNA. Since inactivation of PU.1 is closely related to the development of myeloid leukemia and Ski strongly inhibits PU.1 function, we propose that aberrant Ski expression in certain types of myeloid cell lineages might contribute to leukemogenesis. PMID:17621263

Plasminogen promotes macrophage phagocytosis in mice

PubMed Central

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

2014-01-01

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

Temporal Changes in Myeloid Cells in the Cervix during Pregnancy and Parturition

PubMed Central

Timmons, Brenda C; Fairhurst, Anna–Marie; Mahendroo, Mala S

2009-01-01

Preterm birth occurs at a rate of 12.7% in the United States and is the primary cause of fetal morbidity in the first year of life as well as the cause of later health problems. Elucidation of mechanisms controlling cervical remodeling is critical for development of therapies to reduce the incidence of prematurity. The cervical extracellular matrix must be disorganized during labor to allow birth followed by a rapid repair postpartum. Leukocytes infiltrate the cervix prior to and after birth and are proposed to regulate matrix remodeling during cervical ripening via release of proteolytic enzymes. In the current study, flow cytometry and cell sorting were utilized to determine the role of immune cells in cervical matrix remodeling before, during, and after parturition. Markers of myeloid cell differentiation and activation were assessed to define phenotype and function. Tissue monocytes and eosinophils increased in the cervix prior to birth in a progesterone regulated fashion while macrophage numbers were unchanged. Neutrophils increased in the postpartum period. Increased mRNA expression of Csfr1 and markers of alternatively activated M2 macrophages during labor or shortly postpartum suggest a function of M2 macrophages in postpartum tissue repair. Changes in cervical myeloid cell numbers are not reflected in the peripheral blood. These data along with our previous studies suggest that myeloid derived cells do not orchestrate processes required for initiation of cervical ripening prior to birth. Additionally, macrophages with diverse phenotypes (M1 and M2) are present in the cervix and likely involved in the postpartum repair of tissue. PMID:19234164

Nitric Oxide and KLF4 Protein Epigenetically Modify Class II Transactivator to Repress Major Histocompatibility Complex II Expression during Mycobacterium bovis Bacillus Calmette-Guérin Infection*

PubMed Central

Ghorpade, Devram Sampat; Holla, Sahana; Sinha, Akhauri Yash; Alagesan, Senthil Kumar; Balaji, Kithiganahalli Narayanaswamy

2013-01-01

Pathogenic mycobacteria employ several immune evasion strategies such as inhibition of class II transactivator (CIITA) and MHC-II expression, to survive and persist in host macrophages. However, precise roles for specific signaling components executing down-regulation of CIITA/MHC-II have not been adequately addressed. Here, we demonstrate that Mycobacterium bovis bacillus Calmette-Guérin (BCG)-mediated TLR2 signaling-induced iNOS/NO expression is obligatory for the suppression of IFN-γ-induced CIITA/MHC-II functions. Significantly, NOTCH/PKC/MAPK-triggered signaling cross-talk was found critical for iNOS/NO production. NO responsive recruitment of a bifunctional transcription factor, KLF4, to the promoter of CIITA during M. bovis BCG infection of macrophages was essential to orchestrate the epigenetic modifications mediated by histone methyltransferase EZH2 or miR-150 and thus calibrate CIITA/MHC-II expression. NO-dependent KLF4 regulated the processing and presentation of ovalbumin by infected macrophages to reactive T cells. Altogether, our study delineates a novel role for iNOS/NO/KLF4 in dictating the mycobacterial capacity to inhibit CIITA/MHC-II-mediated antigen presentation by infected macrophages and thereby elude immune surveillance. PMID:23733190

Contrasting phagosome pH regulation and maturation in human M1 and M2 macrophages

PubMed Central

Canton, Johnathan; Khezri, Rojyar; Glogauer, Michael; Grinstein, Sergio

2014-01-01

Macrophages respond to changes in environmental stimuli by assuming distinct functional phenotypes, a phenomenon referred to as macrophage polarization. We generated classically (M1) and alternatively (M2) polarized macrophages—two extremes of the polarization spectrum—to compare the properties of their phagosomes. Specifically, we analyzed the regulation of the luminal pH after particle engulfment. The phagosomes of M1 macrophages had a similar buffering power and proton (equivalent) leakage permeability but significantly reduced proton-pumping activity compared with M2 phagosomes. As a result, only the latter underwent a rapid and profound acidification. By contrast, M1 phagosomes displayed alkaline pH oscillations, which were caused by proton consumption upon dismutation of superoxide, followed by activation of a voltage- and Zn2+-sensitive permeation pathway, likely HV1 channels. The paucity of V-ATPases in M1 phagosomes was associated with, and likely caused by, delayed fusion with late endosomes and lysosomes. The delayed kinetics of maturation was, in turn, promoted by the failure of M1 phagosomes to acidify. Thus, in M1 cells, elimination of pathogens through deployment of the microbicidal NADPH oxidase is given priority at the expense of delayed acidification. By contrast, M2 phagosomes proceed to acidify immediately in order to clear apoptotic bodies rapidly and effectively. PMID:25165138

Impact of Leishmania metalloprotease GP63 on macrophage signaling

PubMed Central

Isnard, Amandine; Shio, Marina T.; Olivier, Martin

2012-01-01

The intramacrophage protozoan parasites of Leishmania genus have developed sophisticated ways to subvert the innate immune response permitting their infection and propagation within the macrophages of the mammalian host. Several Leishmania virulence factors have been identified and found to be of importance for the development of leishmaniasis. However, recent findings are now further reinforcing the critical role played by the zinc-metalloprotease GP63 as a virulence factor that greatly influence host cell signaling mechanisms and related functions. GP63 has been found to be involved not only in the cleavage and degradation of various kinases and transcription factors, but also to be the major molecule modulating host negative regulatory mechanisms involving for instance protein tyrosine phosphatases (PTPs). Those latter being well recognized for their pivotal role in the regulation of a great number of signaling pathways. In this review article, we are providing a complete overview about the role of Leishmania GP63 in the mechanisms underlying the subversion of macrophage signaling and functions. PMID:22919663

Impact of Leishmania metalloprotease GP63 on macrophage signaling.

PubMed

Isnard, Amandine; Shio, Marina T; Olivier, Martin

2012-01-01

The intramacrophage protozoan parasites of Leishmania genus have developed sophisticated ways to subvert the innate immune response permitting their infection and propagation within the macrophages of the mammalian host. Several Leishmania virulence factors have been identified and found to be of importance for the development of leishmaniasis. However, recent findings are now further reinforcing the critical role played by the zinc-metalloprotease GP63 as a virulence factor that greatly influence host cell signaling mechanisms and related functions. GP63 has been found to be involved not only in the cleavage and degradation of various kinases and transcription factors, but also to be the major molecule modulating host negative regulatory mechanisms involving for instance protein tyrosine phosphatases (PTPs). Those latter being well recognized for their pivotal role in the regulation of a great number of signaling pathways. In this review article, we are providing a complete overview about the role of Leishmania GP63 in the mechanisms underlying the subversion of macrophage signaling and functions.

Regulation of reverse cholesterol transport - a comprehensive appraisal of available animal studies

PubMed Central

2012-01-01

Plasma levels of high density lipoprotein (HDL) cholesterol are strongly inversely correlated to the risk of atherosclerotic cardiovascular disease. A major recognized functional property of HDL particles is to elicit cholesterol efflux and consequently mediate reverse cholesterol transport (RCT). The recent introduction of a surrogate method aiming at determining specifically RCT from the macrophage compartment has facilitated research on the different components and pathways relevant for RCT. The current review provides a comprehensive overview of studies carried out on macrophage-specific RCT including a quick reference guide of available data. Knowledge and insights gained on the regulation of the RCT pathway are summarized. A discussion of methodological issues as well as of the respective relevance of specific pathways for RCT is also included. PMID:22458435

Maternal low protein diet leads to placental angiogenic compensation via dysregulated M1/M2 macrophages and TNFa expression in Sprague-Dawley rats

USDA-ARS?s Scientific Manuscript database

A maternal low-protein (LP) diet results in low birth weight, increased offspring rapid adipose tissue catch-up growth, adult obesity, and insulin resistance in Sprague-Dawley rats. The placenta functions to fulfill the fetus’ nutrient demands. Placental function is dependent on regulation of immune...

Suppression of human macrophage function in vitro by delta 9-tetrahydrocannabinol.

PubMed

Specter, S; Lancz, G; Goodfellow, D

1991-11-01

The ability of macrophages to function in the presence of delta 9-tetrahydrocannabinol (THC), the major psychoactive component in marijuana, was evaluated. THC added to macrophage cultures prepared from human peripheral blood inhibited macrophage spreading and phagocytosis of yeast. The effects of THC were concentration dependent, with inhibitory effects observed from 10 to 1 micrograms/ml or lower. These results suggest that macrophages are more sensitive to THC than are lymphocytes because macrophage functions were inhibited by THC at concentrations that did not affect lymphocyte function. Thus, inhibition of lymphocyte function(s) by THC could be attributed to a direct effect of the drug on macrophages which indirectly results in lowered lymphoid cell activity.

Quantitative Profiling of Protein S-Glutathionylation Reveals Redox-Dependent Regulation of Macrophage Function During Nanoparticle-Induced Oxidative Stress

DOE PAGES

Duan, Jicheng; Kodali, Vamsi K.; Gaffrey, Matthew J.; ...

2015-12-23

Engineered nanoparticles (ENPs) are emerging functional materials increasingly utilized for commercial and medical applications. Due to the potential hazard effects of ENPs to human health, it is significant to assess and understand the underlying mechanisms of nanotoxicity. Here, we investigate protein S-glutathionylation (SSG) as an underlying regulatory mechanism for ENP-induced oxidative stress in macrophages by applying a recently developed quantitative redox proteomics approach for site-specific measurements of SSG. Three high-volume production ENPs (SiO 2, Fe 3O 4 and CoO) were selected as representative ENPs with low, moderate, and high reactive oxygen species (ROS) activity, respectively. Among these nanoparticles, we observemore » that CoO led to the most significant dose-dependent oxidative stress and increase of protein SSG modifications in macrophages. Our site-specific SSG changes highlighted a broad set of redox sensitive proteins and their specific Cys residues potentially implicated in stress response. Functional analysis revealed that the most significantly enriched functional categories for SSG-modified proteins were stress response, cellular structure change, and cell death or survival. Moreover, ENPs-induce oxidative stress levels (CoO > Fe 3O 4 > SiO 2) were found to correlate well with the levels of impairment of macrophage phagocytic activity and the overall degrees of increases in SSG. RNA silencing knockdown experiment of glutaredoxin 1 (Grx1) also led to a decreased phagocytic activity in macrophages, which suggested a regulatory role of SSG in phagocytosis. Together, the results provided valuable insights of protein SSG as a potential regulatory mechanism in response to nanomaterial-induced oxidative stress and immunity dysfunction.« less

Cyclic GMP-dependent protein kinase II is necessary for macrophage M1 polarization and phagocytosis via toll-like receptor 2.

PubMed

Liao, Wei-Ting; You, Huey-Ling; Li, Changgui; Chang, Jan-Gowth; Chang, Shun-Jen; Chen, Chung-Jen

2015-05-01

Cyclic GMP-dependent protein kinase II (cGKII; PRKG2) phosphorylates a variety of biological targets and has been identified as a gout-susceptible gene. However, the regulatory role of cGKII in triggering gout disease has yet to be clarified. Thus, we plan to explore the specific function of cGKII in macrophages related to gout disease. By using cGKII gene knockdown method, we detected macrophage M1/M2 polarization, phagocytosis, and their responses to stimulation by monosodium urate (MSU). cGKII was highly expressed in M1 phenotype, but not in M2, and cGKII knockdown significantly inhibited macrophage M1 polarization by decreasing M1 chemokine markers (CXCL10 and CCL2) and downregulating phagocytosis function. We further identified that cGKII-associated phagocytosis was mediated by upregulating toll-like receptor 2 (TLR2) expression, but not by TLR4. Mimicking gout condition by MSU treatments, we found that MSU alone induced cGKII and TLR2 expression with increased M1 polarization markers and phagocytosis activity. It means that cGKII knockdown significantly inhibited this MSU-induced cGKII-TLR2-phagocytosis axis. Our study showed that cGKII plays a key role in M1 polarization, especially in TLR2-mediated phagocytosis under MSU exposure. The findings provide evidence for the possible role of cGKII as an inflammation exciter in gout disease. Gout-susceptible gene cGKII is necessary for macrophage M1 polarization. cGKII regulates M1 phagocytosis function via TLR2. Monosodium urate treatments increase cGKII expression and related function. This study reveals the role of cGKII in enhancing gouty inflammatory responses.

Quantitative Profiling of Protein S-Glutathionylation Reveals Redox-Dependent Regulation of Macrophage Function During Nanoparticle-Induced Oxidative Stress

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

Duan, Jicheng; Kodali, Vamsi K.; Gaffrey, Matthew J.

Engineered nanoparticles (ENPs) are emerging functional materials increasingly utilized for commercial and medical applications. Due to the potential hazard effects of ENPs to human health, it is significant to assess and understand the underlying mechanisms of nanotoxicity. Here, we investigate protein S-glutathionylation (SSG) as an underlying regulatory mechanism for ENP-induced oxidative stress in macrophages by applying a recently developed quantitative redox proteomics approach for site-specific measurements of SSG. Three high-volume production ENPs (SiO 2, Fe 3O 4 and CoO) were selected as representative ENPs with low, moderate, and high reactive oxygen species (ROS) activity, respectively. Among these nanoparticles, we observemore » that CoO led to the most significant dose-dependent oxidative stress and increase of protein SSG modifications in macrophages. Our site-specific SSG changes highlighted a broad set of redox sensitive proteins and their specific Cys residues potentially implicated in stress response. Functional analysis revealed that the most significantly enriched functional categories for SSG-modified proteins were stress response, cellular structure change, and cell death or survival. Moreover, ENPs-induce oxidative stress levels (CoO > Fe 3O 4 > SiO 2) were found to correlate well with the levels of impairment of macrophage phagocytic activity and the overall degrees of increases in SSG. RNA silencing knockdown experiment of glutaredoxin 1 (Grx1) also led to a decreased phagocytic activity in macrophages, which suggested a regulatory role of SSG in phagocytosis. Together, the results provided valuable insights of protein SSG as a potential regulatory mechanism in response to nanomaterial-induced oxidative stress and immunity dysfunction.« less

RNA sequencing demonstrates large-scale temporal dysregulation of gene expression in stimulated macrophages derived from MHC-defined chicken haplotypes.

PubMed

Irizarry, Kristopher J L; Downs, Eileen; Bryden, Randall; Clark, Jory; Griggs, Lisa; Kopulos, Renee; Boettger, Cynthia M; Carr, Thomas J; Keeler, Calvin L; Collisson, Ellen; Drechsler, Yvonne

2017-01-01

Discovering genetic biomarkers associated with disease resistance and enhanced immunity is critical to developing advanced strategies for controlling viral and bacterial infections in different species. Macrophages, important cells of innate immunity, are directly involved in cellular interactions with pathogens, the release of cytokines activating other immune cells and antigen presentation to cells of the adaptive immune response. IFNγ is a potent activator of macrophages and increased production has been associated with disease resistance in several species. This study characterizes the molecular basis for dramatically different nitric oxide production and immune function between the B2 and the B19 haplotype chicken macrophages.A large-scale RNA sequencing approach was employed to sequence the RNA of purified macrophages from each haplotype group (B2 vs. B19) during differentiation and after stimulation. Our results demonstrate that a large number of genes exhibit divergent expression between B2 and B19 haplotype cells both prior and after stimulation. These differences in gene expression appear to be regulated by complex epigenetic mechanisms that need further investigation.

Exposure to di(n-butyl)phthalate and benzo(a)pyrene alters IL-1{beta} secretion and subset expression of testicular macrophages, resulting in decreased testosterone production in rats

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

Zheng Shanjun; Key Laboratory of High Altitude Medicine, Ministry of Education, Chongqing 400038; Key Laboratory of High Altitude Physiology and High Altitude Disease, PLA, Chongqing 400038

Di(n-butyl)phthalate (DBP) and benzo(a)pyrene (BaP) are environmental endocrine disruptors that are potentially hazardous to humans. These chemicals affect testicular macrophage immuno-endocrine function and testosterone production. However, the underlying mechanisms for these effects are not fully understood. It is well known that interleukin-1 beta (IL-1{beta}), which is secreted by testicular macrophages, plays a trigger role in regulating Leydig cell steroidogenesis. The purpose of this study was to reveal the effects of co-exposure to DBP and BaP on testicular macrophage subset expression, IL-1{beta} secretion and testosterone production. Adult male Sprague-Dawley rats were randomly divided into seven groups; two groups received DBP plusmore » BaP (DBP + BaP: 50 + 1 or 250 + 5 mg/kg/day) four groups received DBP or BaP alone (DBP: 50 or 250 mg/kg/day; BaP: 1 or 5 mg/kg/day), and one group received vehicle alone (control). After co-exposure for 90 days, the relative expression of macrophage subsets and their functions changed. ED2{sup +} testicular macrophages (reactive with a differentiation-related antigen present on the resident macrophages) were activated and IL-1{beta} secretion was enhanced. DBP and BaP acted additively, as demonstrated by greater IL-1{beta} secretion relative to each compound alone. These observations suggest that exposure to DBP plus BaP exerted greater suppression on testosterone production compared with each compound alone. The altered balance in the subsets of testicular macrophages and the enhanced ability of resident testicular macrophages to secrete IL-1{beta}, resulted in enhanced production of IL-1{beta} as a potent steroidogenesis repressor. This may represent an important mechanism by which DBP and BaP repress steroidogenesis.« less

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

PubMed

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

2018-06-01

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

Melanocortin 1 Receptor Signaling Regulates Cholesterol Transport in Macrophages.

PubMed

Rinne, Petteri; Rami, Martina; Nuutinen, Salla; Santovito, Donato; van der Vorst, Emiel P C; Guillamat-Prats, Raquel; Lyytikäinen, Leo-Pekka; Raitoharju, Emma; Oksala, Niku; Ring, Larisa; Cai, Minying; Hruby, Victor J; Lehtimäki, Terho; Weber, Christian; Steffens, Sabine

2017-07-04

The melanocortin 1 receptor (MC1-R) is expressed by monocytes and macrophages, where it exerts anti-inflammatory actions on stimulation with its natural ligand α-melanocyte-stimulating hormone. The present study was designed to investigate the specific role of MC1-R in the context of atherosclerosis and possible regulatory pathways of MC1-R beyond anti-inflammation. Human and mouse atherosclerotic samples and primary mouse macrophages were used to study the regulatory functions of MC1-R. The impact of pharmacological MC1-R activation on atherosclerosis was assessed in apolipoprotein E-deficient mice. Characterization of human and mouse atherosclerotic plaques revealed that MC1-R expression localizes in lesional macrophages and is significantly associated with the ATP-binding cassette transporters ABCA1 and ABCG1, which are responsible for initiating reverse cholesterol transport. Using bone marrow-derived macrophages, we observed that α-melanocyte-stimulating hormone and selective MC1-R agonists similarly promoted cholesterol efflux, which is a counterregulatory mechanism against foam cell formation. Mechanistically, MC1-R activation upregulated the levels of ABCA1 and ABCG1. These effects were accompanied by a reduction in cell surface CD36 expression and in cholesterol uptake, further protecting macrophages from excessive lipid accumulation. Conversely, macrophages deficient in functional MC1-R displayed a phenotype with impaired efflux and enhanced uptake of cholesterol. Pharmacological targeting of MC1-R in atherosclerotic apolipoprotein E-deficient mice reduced plasma cholesterol levels and aortic CD36 expression and increased plaque ABCG1 expression and signs of plaque stability. Our findings identify a novel role for MC1-R in macrophage cholesterol transport. Activation of MC1-R confers protection against macrophage foam cell formation through a dual mechanism: It prevents cholesterol uptake while concomitantly promoting ABCA1- and ABCG1-mediated reverse cholesterol transport. © 2017 American Heart Association, Inc.

Akirin1 (Mighty), a novel promyogenic factor regulates muscle regeneration and cell chemotaxis

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

Salerno, Monica Senna; Dyer, Kelly; Bracegirdle, Jeremy

2009-07-15

Akirin1 (Mighty) is a downstream target gene of myostatin and has been shown to be a promyogenic factor. Although expressed in many tissues, akirin1 is negatively regulated by myostatin specifically in skeletal muscle tissue. In this manuscript we have characterized the possible function of akirin1 in postnatal muscle growth. Molecular and immunohistological analyses indicated that while low levels of akirin1 are associated with quiescent satellite cells (SC), higher levels of akirin1 are detected in activated proliferating SC indicating that akirin1 could be associated with satellite cell activation. In addition to SC, macrophages also express akirin1, and increased expression of akirin1more » resulted in more efficient chemotaxis of both macrophages and myoblasts. Akirin1 appears to regulate chemotaxis of both macrophages and myoblasts by reorganising actin cytoskeleton, leading to more efficient lamellipodia formation via a PI3 kinase dependent pathway. Expression analysis during muscle regeneration also indicated that akirin1 expression is detected very early (day 2) in regenerating muscle, and expression gradually peaks to coincide the nascent myotube formation stage of muscle regeneration. Based on these results we propose that akirin1 could be acting as a transducer of early signals of muscle regeneration. Thus, we speculate that myostatin regulates key steps of muscle regeneration including chemotaxis of inflammatory cells, SC activation and migration through akirin1.« less

The pathogenesis of diclofenac induced immunoallergic hepatitis in a canine model of liver injury

PubMed Central

Selvaraj, Saravanakumar; Oh, Jung-Hwa; Spanel, Reinhard; Länger, Florian; Han, Hyoung-Yun; Lee, Eun-Hee; Yoon, Seokjoo; Borlak, Jürgen

2017-01-01

Hypersensitivity to non-steroidal anti-inflammatory drugs is a common adverse drug reaction and may result in serious inflammatory reactions of the liver. To investigate mechanism of immunoallergic hepatitis beagle dogs were given 1 or 3 mg/kg/day (HD) oral diclofenac for 28 days. HD diclofenac treatment caused liver function test abnormalities, reduced haematocrit and haemoglobin but induced reticulocyte, WBC, platelet, neutrophil and eosinophil counts. Histopathology evidenced hepatic steatosis and glycogen depletion, apoptosis, acute lobular hepatitis, granulomas and mastocytosis. Whole genome scans revealed 663 significantly regulated genes of which 82, 47 and 25 code for stress, immune response and inflammation. Immunopathology confirmed strong induction of IgM, the complement factors C3&B, SAA, SERPING1 and others of the classical and alternate pathway. Alike, marked expression of CD205 and CD74 in Kupffer cells and lymphocytes facilitate antigen presentation and B-cell differentiation. The highly induced HIF1A and KLF6 protein expression in mast cells and macrophages sustain inflammation. Furthermore, immunogenomics discovered 24, 17, 6 and 11 significantly regulated marker genes to hallmark M1/M2 polarized macrophages, lymphocytic and granulocytic infiltrates; note, the latter was confirmed by CAE staining. Other highly regulated genes included alpha-2-macroglobulin, CRP, hepcidin, IL1R1, S100A8 and CCL20. Diclofenac treatment caused unprecedented induction of myeloperoxidase in macrophages and oxidative stress as shown by SOD1/SOD2 immunohistochemistry. Lastly, bioinformatics defined molecular circuits of inflammation and consisted of 161 regulated genes. Altogether, the mechanism of diclofenac induced liver hypersensitivity reactions involved oxidative stress, macrophage polarization, mastocytosis, complement activation and an erroneous programming of the innate and adaptive immune system. PMID:29296203

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

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

Shi, Liang; Chowdhury, Saiful M.; Smallwood, Heather S.

2009-08-01

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

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

PubMed

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

2017-07-04

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

Estrogen Represses Hepatocellular Carcinoma (HCC) Growth via Inhibiting Alternative Activation of Tumor-associated Macrophages (TAMs)*

PubMed Central

Yang, Weiwei; Lu, Yan; Xu, Yichen; Xu, Lizhi; Zheng, Wei; Wu, Yuanyuan; Li, Long; Shen, Pingping

2012-01-01

Hepatocarcinoma cancer (HCC), one of the most malignant cancers, occurs significantly more often in men than in women; however, little is known about its underlying molecular mechanisms. Here we identified that 17β-estradiol (E2) could suppress tumor growth via regulating the polarization of macrophages. We showed that E2 re-administration reduced tumor growth in orthotopic and ectopic mice HCC models. E2 functioned as a suppressor for macrophage alternative activation and tumor progression by keeping estrogen receptor β (ERβ) away from interacting with ATP5J (also known as ATPase-coupling factor 6), a part of ATPase, thus inhibiting the JAK1-STAT6 signaling pathway. These studies introduce a novel mechanism for suppressing male-predominant HCC. PMID:22908233

Harnessing the power of macrophages/monocytes for enhanced bone tissue engineering.

PubMed

Dong, Lei; Wang, Chunming

2013-06-01

Bone tissue engineering has attracted considerable attention as a promising treatment modality for severe bone degeneration. The pressing need for more sophisticated and fully functional bone substitutes has spurred a refocus on the development of bone constructs in a way more comparable to the physiological process. Current research is increasingly revealing the central roles of macrophages/monocytes in regulating bone development and repair, so we propose that these immunocytes can play a similar pivotal role in directing engineered bone regeneration. Accordingly, we discuss two possible strategies to exemplify how the distinctive power of macrophages/monocytes--particularly their cytokine-secretion ability and chemotactic response to foreign materials--can be harnessed to enhance the performance of bone tissue engineering applications. Copyright © 2013 Elsevier Ltd. All rights reserved.

Hck is a key regulator of gene expression in alternatively activated human monocytes.

PubMed

Bhattacharjee, Ashish; Pal, Srabani; Feldman, Gerald M; Cathcart, Martha K

2011-10-21

IL-13 is a Th2 cytokine that promotes alternative activation (M2 polarization) in primary human monocytes. Our studies have characterized the functional IL-13 receptor complex and the downstream signaling events in response to IL-13 stimulation in alternatively activated monocytes/macrophages. In this report, we present evidence that IL-13 induces the activation of a Src family tyrosine kinase, which is required for IL-13 induction of M2 gene expression, including 15-lipoxygenase (15-LO). Our data show that Src kinase activity regulates IL-13-induced p38 MAPK tyrosine phosphorylation via the upstream kinases MKK3 or MKK6. Our findings also reveal that the IL-13 receptor-associated tyrosine kinase Jak2 is required for the activation of both Src kinase as well as p38 MAPK. Further, we found that Src tyrosine kinase-mediated activation of p38 MAPK is required for Stat1 and Stat3 serine 727 phosphorylation in alternatively activated monocytes/macrophages. Additional studies identify Hck as the specific Src family member, stimulated by IL-13 and involved in regulating both p38 MAPK activation and p38 MAPK-mediated 15-LO expression. Finally we show that the Hck regulates the expression of other alternative state (M2)-specific genes (Mannose receptor, MAO-A, and CD36) and therefore conclude that Hck acts as a key regulator controlling gene expression in alternatively activated monocytes/macrophages.

A vitellogenic-like carboxypeptidase expressed by human macrophages is localized in endoplasmic reticulum and membrane ruffles

PubMed Central

Harris, James; Schwinn, Nicole; Mahoney, James A; Lin, Hsi-Hsien; Shaw, Michael; Howard, Chris J; da Silva, Rosangela P; Gordon, Siamon

2006-01-01

Carboxypeptidase, vitellogenic-like (CPVL) is a serine carboxypeptidase of unknown function that was first characterized in human macrophages. Initial studies suggested that CPVL is largely restricted to the monocytic lineage, although it may also be expressed by cells outside the immune system. Here, we use a new monoclonal antibody to characterize the properties and localization of CPVL in human macrophages to elucidate a possible function for the protease. CPVL is up-regulated during the maturation of monocytes (MO) to macrophages, although the protein can be seen in both. In primary macrophages, CPVL is glycosylated with high mannose residues and colocalizes with markers for endoplasmic reticulum, while in MO it is more disperse and less clearly associated with endoplasmic reticulum. CPVL is highly expressed in lamellipodia and membrane ruffles, which also concentrate markers of the secretory pathway (MIP-1α and tumour necrosis factor-α) and major histocompatibility complex (MHC) class I and II molecules. CPVL can be seen on early latex bead and Candida albicans phagosomes, but it is not retained in the maturing phagosome, unlike MHC class I/II. CPVL has a mixed cytosolic and membrane-associated localization but is not detectable on the outer plasma membrane. We propose that CPVL may be involved in antigen processing, the secretory pathway and/or in actin remodelling and lamellipodium formation. PMID:16436111

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

PubMed

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.

Cyclic AMP Pathway Suppress Autoimmune Neuroinflammation by Inhibiting Functions of Encephalitogenic CD4 T Cells and Enhancing M2 Macrophage Polarization at the Site of Inflammation

PubMed Central

Veremeyko, Tatyana; Yung, Amanda W. Y.; Dukhinova, Marina; Kuznetsova, Inna S.; Pomytkin, Igor; Lyundup, Alexey; Strekalova, Tatyana; Barteneva, Natasha S.; Ponomarev, Eugene D.

2018-01-01

Although it has been demonstrated that cAMP pathway affect both adaptive and innate cell functions, the role of this pathway in the regulation of T-cell-mediated central nervous system (CNS) autoimmune inflammation, such as in experimental autoimmune encephalomyelitis (EAE), remains unclear. It is also unclear how cAMP pathway affects the function of CD4 T cells in vivo at the site of inflammation. We found that adenylyl cyclase activator Forskolin besides inhibition of functions autoimmune CD4 T cells also upregulated microRNA (miR)-124 in the CNS during EAE, which is associated with M2 phenotype of microglia/macrophages. Our study further established that in addition to direct influence of cAMP pathway on CD4 T cells, stimulation of this pathway promoted macrophage polarization toward M2 leading to indirect inhibition of function of T cells in the CNS. We demonstrated that Forskolin together with IL-4 or with Forskolin together with IL-4 and IFNγ effectively stimulated M2 phenotype of macrophages indicating high potency of this pathway in reprogramming of macrophage polarization in Th2- and even in Th1/Th2-mixed inflammatory conditions such as EAE. Mechanistically, Forskolin and/or IL-4 activated ERK pathway in macrophages resulting in the upregulation of M2-associated molecules miR-124, arginase (Arg)1, and Mannose receptor C-type 1 (Mrc1), which was reversed by ERK inhibitors. Administration of Forskolin after the onset of EAE substantially upregulated M2 markers Arg1, Mrc1, Fizz1, and Ym1 and inhibited M1 markers nitric oxide synthetase 2 and CD86 in the CNS during EAE resulting in decrease in macrophage/microglia activation, lymphocyte and CD4 T cell infiltration, and the recovery from the disease. Forskolin inhibited proliferation and IFNγ production by CD4 T cells in the CNS but had rather weak direct effect on proliferation of autoimmune T cells in the periphery and in vitro, suggesting prevalence of indirect effect of Forskolin on differentiation and functions of autoimmune CD4 T cells in vivo. Thus, our data indicate that Forskolin has potency to skew balance toward M2 affecting ERK pathway in macrophages and indirectly inhibit pathogenic CD4 T cells in the CNS leading to the suppression of autoimmune inflammation. These data may have also implications for future therapeutic approaches to inhibit autoimmune Th1 cells at the site of tissue inflammation. PMID:29422898

Cyclic AMP Pathway Suppress Autoimmune Neuroinflammation by Inhibiting Functions of Encephalitogenic CD4 T Cells and Enhancing M2 Macrophage Polarization at the Site of Inflammation.

PubMed

Veremeyko, Tatyana; Yung, Amanda W Y; Dukhinova, Marina; Kuznetsova, Inna S; Pomytkin, Igor; Lyundup, Alexey; Strekalova, Tatyana; Barteneva, Natasha S; Ponomarev, Eugene D

2018-01-01

Although it has been demonstrated that cAMP pathway affect both adaptive and innate cell functions, the role of this pathway in the regulation of T-cell-mediated central nervous system (CNS) autoimmune inflammation, such as in experimental autoimmune encephalomyelitis (EAE), remains unclear. It is also unclear how cAMP pathway affects the function of CD4 T cells in vivo at the site of inflammation. We found that adenylyl cyclase activator Forskolin besides inhibition of functions autoimmune CD4 T cells also upregulated microRNA (miR)-124 in the CNS during EAE, which is associated with M2 phenotype of microglia/macrophages. Our study further established that in addition to direct influence of cAMP pathway on CD4 T cells, stimulation of this pathway promoted macrophage polarization toward M2 leading to indirect inhibition of function of T cells in the CNS. We demonstrated that Forskolin together with IL-4 or with Forskolin together with IL-4 and IFNγ effectively stimulated M2 phenotype of macrophages indicating high potency of this pathway in reprogramming of macrophage polarization in Th2- and even in Th1/Th2-mixed inflammatory conditions such as EAE. Mechanistically, Forskolin and/or IL-4 activated ERK pathway in macrophages resulting in the upregulation of M2-associated molecules miR-124, arginase (Arg)1, and Mannose receptor C-type 1 (Mrc1), which was reversed by ERK inhibitors. Administration of Forskolin after the onset of EAE substantially upregulated M2 markers Arg1, Mrc1, Fizz1, and Ym1 and inhibited M1 markers nitric oxide synthetase 2 and CD86 in the CNS during EAE resulting in decrease in macrophage/microglia activation, lymphocyte and CD4 T cell infiltration, and the recovery from the disease. Forskolin inhibited proliferation and IFNγ production by CD4 T cells in the CNS but had rather weak direct effect on proliferation of autoimmune T cells in the periphery and in vitro , suggesting prevalence of indirect effect of Forskolin on differentiation and functions of autoimmune CD4 T cells in vivo . Thus, our data indicate that Forskolin has potency to skew balance toward M2 affecting ERK pathway in macrophages and indirectly inhibit pathogenic CD4 T cells in the CNS leading to the suppression of autoimmune inflammation. These data may have also implications for future therapeutic approaches to inhibit autoimmune Th1 cells at the site of tissue inflammation.

Differential regulation of IL-23 production in M1 macrophages by TIR8/SIGIRR through TLR4- or TLR7/8-mediated signaling.

PubMed

Yamaguchi, Rui; Sakamoto, Arisa; Yamamoto, Takatoshi; Narahara, Shinji; Sugiuchi, Hiroyuki; Yamaguchi, Yasuo

2017-11-01

Cross-talks between toll-like receptors (TLRs) including various negative regulatory mechanisms are many unknown. We investigated the differential mechanism of IL-23 production in M1 macrophages by single immunoglobulin interleukin-1 receptor-related (SIGIRR) molecule through TLR4 or TLR7/8. IL-12p40 production by M1 macrophages pretreated with human neutrophil elastase (HNE) was synergistically enhanced IL-12p40, but not IL-23 production, after exposure to lipopolysaccharide (LPS). LPS (a TLR4 agonist) induced a slight increase of IL-23 production, while Resiquimod (a TLR7/8 agonist) significantly enhanced IL-23 production. Expression of SIGIRR protein, a negative regulator of TLR4, was higher in M1 macrophages than in monocytes. Interestingly, SIGIRR siRNA induced a slight increment of IL-23 production after exposure of macrophages to LPS, while IL-23 production in response to Resiquimod was significantly upregulated by SIGIRR siRNA. Silencing SIGIRR enhanced IRF4 protein level determined by western blotting or ELISA. IRF4 siRNA dramatically restored IL-23 production after exposure to Resiquimod in macrophages transfected with SIGIRR siRNA. In conclusion, production of IL-23 is differentially regulated in M1 macrophages by SIGIRR through TLR4- or TLR7/8-mediated signaling. SIGIRR is both a negative regulator of TLR4 and a positive regulator of TLR7/8. Copyright © 2017 Elsevier Ltd. All rights reserved.

An Essential Role for ECSIT in Mitochondrial Complex I Assembly and Mitophagy in Macrophages.

PubMed

Carneiro, Flávia R G; Lepelley, Alice; Seeley, John J; Hayden, Matthew S; Ghosh, Sankar

2018-03-06

ECSIT is a mitochondrial complex I (CI)-associated protein that has been shown to regulate the production of mitochondrial reactive oxygen species (mROS) following engagement of Toll-like receptors (TLRs). We have generated an Ecsit conditional knockout (CKO) mouse strain to study the in vivo role of ECSIT. ECSIT deletion results in profound alteration of macrophage metabolism, leading to a striking shift to reliance on glycolysis, complete disruption of CI activity, and loss of the CI holoenzyme and multiple subassemblies. An increase in constitutive mROS production in ECSIT-deleted macrophages prevents further TLR-induced mROS production. Surprisingly, ECSIT-deleted cells accumulate damaged mitochondria because of defective mitophagy. ECSIT associates with the mitophagy regulator PINK1 and exhibits Parkin-dependent ubiquitination. However, upon ECSIT deletion, we observed increased mitochondrial Parkin without the expected increase in mitophagy. Taken together, these results demonstrate a key role of ECSIT in CI function, mROS production, and mitophagy-dependent mitochondrial quality control. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Establishment of S100A8 Transgenic Rats to Understand Innate Property of S100A8 and Its Immunological Role.

PubMed

Okada, Kohki; Itoh, Hiroshi; Kamikubo, Yasuhiko; Adachi, Souichi; Ikemoto, Masaki

2018-02-01

The innate properties of S100A8 as a regulator in acute inflammation have not yet been elucidated in detail. Our aims are to newly establish S100A8 transgenic rats (Tg-S100A8) and to elucidate the immunological functions of S100A8. Following the treatment with 5% dextran sulfate sodium for 1 week, the body weight in Tg-S100A8 weakly decreased after the start; however, that in Japanese Wistar rats (WT) significantly decreased in the end. The serum level of CRP in Tg-S100A8 was significantly lower than that in WT, although the concentration of CRP apparently increased in both Tg-S100A8 and WT. The dynamic mobility of S100A8 and S100A9 in macrophages was microscopically observed using fluorescent immunological staining, in which the S100A9 was dominantly expressed in many macrophages in the rectal tissue of WT. As determined by PCR and real-time PCR, the levels of S100A8 messenger RNA (mRNA) in several organ tissues of the Tg-S100A8, such as heart and small intestine, were apparently higher than those of WT, respectively. The expression of IL-6 and TNF-α mRNAs was negatively regulated in main organ tissues of the large colon of Tg-S100A8 followed by down-regulation of IL-6 protein. An important result was that the expression of S100A8 mRNA was strongly induced in many macrophages of Tg-S100A8, whereas that of some inflammatory cytokine mRNAs described above were significantly reduced. Tg-S100A8 has potential as a useful experimental model rat not only for investigating the innate properties of S100A8 as a regulator, but also for clarifying its functional role in immune cells from a myeloid origin, particularly macrophages.

miR-20a inhibition using locked nucleic acid (LNA) technology and its effects on apoptosis of human macrophages infected by Toxoplasma gondii RH strain.

PubMed

Rezaei, Fatemeh; Daryani, Ahmad; Sharifi, Mohammadreza; Sarvi, Shahabeddin; Jafari, Narjes; Pagheh, Abdol Sattar; Hashemi, Nooshin; Hejazi, Seyed Hossein

2018-05-22

Toxoplasma gondii is a ubiquitous and infectious parasite that multiplies in any nucleated cell of warm-blooded animals and humans worldwide. This parasite has intricate mechanisms to reciprocate host-cell apoptosis to exist in the host cell. So far, the details of the parasite interactions with host cells are not well known. MicroRNAs (miRNAs) are one of the small noncoding RNAs that are now considered as a key mechanism of gene regulation. They are important in physiological and pathological processes such as apoptosis. In this study a Real Time quantitative PCR technique was used to evaluate the levels of miR-20a of miRNAs family in human macrophage during T. gondii infection to determine the role of miR-20a in apoptosis. Then, the inhibition of miR-20a function through interaction with transfection of Locked Nucleic Acid (LNA) antisense oligomer was studied. Furthermore, it was examined whether miR-20a is involved in apoptosis of human macrophages with T. gondii infected cells using flow cytometry. We found that miR-20a expression is up-regulated in human macrophages following T. gondii infection. After LNA anti miR-20a oligomer transfection, miR-20a inhibition was evaluated by quantitative reverse transcriptase polymerase chain reaction. Flow cytometry results showed that LNA anti-miR20a oligomer increased apoptosis. In agreement with this result, we found that specific LNA oligonucleotides prevent the functional activity of miR-20a and promotion of human macrophages apoptosis with T. gondii infection by inhibition of this miRNAs gene. Also, the results support the concept that LNA oligomer antisense may be used as a therapeutic implement for blocking detrimental miRNAs overexpressed in infections. Copyright © 2018 Elsevier Ltd. All rights reserved.

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

PubMed

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

2014-08-01

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

Tissue damage drives co-localization of NF-κB, Smad3, and Nrf2 to direct Rev-erb sensitive wound repair in mouse macrophages

PubMed Central

Eichenfield, Dawn Z; Troutman, Ty Dale; Link, Verena M; Lam, Michael T; Cho, Han; Gosselin, David; Spann, Nathanael J; Lesch, Hanna P; Tao, Jenhan; Muto, Jun; Gallo, Richard L; Evans, Ronald M; Glass, Christopher K

2016-01-01

Although macrophages can be polarized to distinct phenotypes in vitro with individual ligands, in vivo they encounter multiple signals that control their varied functions in homeostasis, immunity, and disease. Here, we identify roles of Rev-erb nuclear receptors in regulating responses of mouse macrophages to complex tissue damage signals and wound repair. Rather than reinforcing a specific program of macrophage polarization, Rev-erbs repress subsets of genes that are activated by TLR ligands, IL4, TGFβ, and damage-associated molecular patterns (DAMPS). Unexpectedly, a complex damage signal promotes co-localization of NF-κB, Smad3, and Nrf2 at Rev-erb-sensitive enhancers and drives expression of genes characteristic of multiple polarization states in the same cells. Rev-erb-sensitive enhancers thereby integrate multiple damage-activated signaling pathways to promote a wound repair phenotype. DOI: http://dx.doi.org/10.7554/eLife.13024.001 PMID:27462873

Redox-signals and macrophage biology (for the upcoming issue of molecular aspects of medicine on signaling by reactive oxygen species).

PubMed

Weigert, Andreas; von Knethen, Andreas; Fuhrmann, Dominik; Dehne, Nathalie; Brüne, Bernhard

2018-01-11

Macrophages are known for their versatile role in biology. They sense and clear structures that contain exogenous or endogenous pathogen-associated molecular patterns. This process is tightly linked to the production of a mixture of potentially harmful oxidants and cytokines. Their inherent destructive behavior is directed against foreign material or structures of 'altered self', which explains the role of macrophages during innate immune reactions and inflammation. However, there is also another side of macrophages when they turn into a tissue regenerative, pro-resolving, and healing phenotype. Phenotype changes of macrophages are termed macrophage polarization, representing a continuum between classical and alternative activation. Macrophages as the dominating producers of superoxide/hydrogen peroxide and nitric oxide are not only prone to oxidative modifications but also to more subtle signaling properties of redox-active molecules conveying redox regulation. We review basic concepts of the enzymatic nitric oxide and superoxide production within macrophages, refer to their unique chemical reactions and outline biological consequences not only for macrophage biology but also for their communication with cells in the microenvironment. These considerations link hypoxia to the NO system, addressing feedforward as well as feedback circuits. Moreover, we summarize the role of redox-signaling affecting epigenetics and reflect the central role of mitochondrial-derived oxygen species in inflammation. To better understand the diverse functions of macrophages during initiation as well as resolution of inflammation and to decode their versatile roles during innate and adaptive immunity with the entire spectrum of cell protective towards cell destructive activities we need to appreciate the signaling properties of redox-active species. Herein we discuss macrophage responses in terms of nitric oxide and superoxide formation with the modulating impact of hypoxia. Copyright © 2018. Published by Elsevier Ltd.

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

PubMed Central

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

Functional plasticity of macrophages: in situ reprogramming of tumor-associated macrophages

PubMed Central

Stout, Robert D.; Watkins, Stephanie K.; Suttles, Jill

2009-01-01

The extent to which the functional heterogeneity of Mϕs is dependent on the differentiation of functional sublineages remains unresolved. One alternative hypothesis proposes that Mϕs are functionally plastic cells, which are capable of altering their functional activities progressively in response to progressively changing signaling molecules generated in their microenvironment. This “functional plasticity” hypothesis predicts that the functionally polarized Mϕs in chronic pathologies do not represent Mϕ sublineages but rather, are mutable phenotypes sustained by chronic signaling from the pathological environment. Solid TAMϕs are chronically polarized to provide activities that support tumor growth and metastasis and suppress adaptive immune responses. In support of the functional plasticity hypothesis, administration of slow-release microsphere-encapsulated IL-12 successfully reprogrammed TAMϕs in situ, reducing Mϕ support of tumor growth and metastasis and enhancing Mϕ proimmunogenic activities. Increased knowledge of how Mϕ function is regulated and how polarized Mϕs can be reprogrammed in situ will increase our ability to control Mϕ function in a variety of pathological states, including cancer and chronic inflammatory disease. PMID:19605698

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

PubMed

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.

Receptor for advanced glycation endproducts (RAGE) maintains pulmonary structure and regulates the response to cigarette smoke.

PubMed

Wolf, Lisa; Herr, Christian; Niederstraßer, Julia; Beisswenger, Christoph; Bals, Robert

2017-01-01

The receptor for advanced glycation endproducts (RAGE) is highly expressed in the lung but its physiological functions in this organ is still not completely understood. To determine the contribution of RAGE to physiological functions of the lung, we analyzed pulmonary mechanics and structure of wildtype and RAGE deficient (RAGE-/-) mice. RAGE deficiency spontaneously resulted in a loss of lung structure shown by an increased mean chord length, increased respiratory system compliance, decreased respiratory system elastance and increased concentrations of serum protein albumin in bronchoalveolar lavage fluids. Pulmonary expression of RAGE was mainly localized on alveolar epithelial cells and alveolar macrophages. Primary murine alveolar epithelial cells isolated from RAGE-/- mice revealed an altered differentiation and defective barrier formation under in vitro conditions. Stimulation of interferone-y (IFNy)-activated alveolar macrophages deficient for RAGE with Toll-like receptor (TLR) ligands resulted in significantly decreased release of proinflammatory cytokines and chemokines. Exposure to chronic cigarette smoke did not affect emphysema-like changes in lung parenchyma in RAGE-/- mice. Acute cigarette smoke exposure revealed a modified inflammatory response in RAGE-/- mice that was characterized by an influx of macrophages and a decreased keratinocyte-derived chemokine (KC) release. Our data suggest that RAGE regulates the differentiation of alveolar epithelial cells and impacts on the development and maintenance of pulmonary structure. In cigarette smoke-induced lung pathology, RAGE mediates inflammation that contributes to lung damage.

Impact of neutrophil-secreted myeloid related proteins 8 and 14 (MRP 8/14) on leishmaniasis progression.

PubMed

Contreras, Irazú; Shio, Marina T; Cesaro, Annabelle; Tessier, Philippe A; Olivier, Martin

2013-01-01

The myeloid-related proteins (MRPs) 8/14 are small proteins mainly produced by neutrophils, which have been reported to induce NO production in macrophages. On the other hand, Leishmania survives and multiplies within phagocytes by inactivating several of their microbicidal functions. Whereas MRPs are rapidly released during the innate immune response, their role in the regulation of Leishmaniasis is still unknown. In vitro experiments revealed that Leishmania infection alters MRP-induced signaling, leading to inhibition of macrophage functions (NO, TNF-α). In contrast, MRP-primed cells showed normal signaling activation and NO production in response to Leishmania infection. Using a murine air-pouch model, we observed that infection with L. major induced leukocyte recruitment and MRP secretion comparable to LPS-treated mice. Depletion of MRPs significantly reduced these inflammatory events and augmented both parasite load and footpad swelling during the first 8 weeks post-infection, as also observed in MRP KO mice. On the contrary, mouse treatment with recombinant MRPs (rMRPs) had the opposite effect. Collectively, our results suggest that rapid secretion of MRPs by neutrophils at the site of infection may protect uninfected macrophages and favor a more efficient innate inflammatory response against Leishmania infection. In summary, our study reveals the critical role played by MRPs in the regulation of Leishmania infection and how this pathogen can subvert its action.

Impact of Neutrophil-Secreted Myeloid Related Proteins 8 and 14 (MRP 8/14) on Leishmaniasis Progression

PubMed Central

Contreras, Irazú; Shio, Marina T.; Cesaro, Annabelle; Tessier, Philippe A.; Olivier, Martin

2013-01-01

The myeloid-related proteins (MRPs) 8/14 are small proteins mainly produced by neutrophils, which have been reported to induce NO production in macrophages. On the other hand, Leishmania survives and multiplies within phagocytes by inactivating several of their microbicidal functions. Whereas MRPs are rapidly released during the innate immune response, their role in the regulation of Leishmaniasis is still unknown. In vitro experiments revealed that Leishmania infection alters MRP-induced signaling, leading to inhibition of macrophage functions (NO, TNF-α). In contrast, MRP-primed cells showed normal signaling activation and NO production in response to Leishmania infection. Using a murine air-pouch model, we observed that infection with L. major induced leukocyte recruitment and MRP secretion comparable to LPS-treated mice. Depletion of MRPs significantly reduced these inflammatory events and augmented both parasite load and footpad swelling during the first 8 weeks post-infection, as also observed in MRP KO mice. On the contrary, mouse treatment with recombinant MRPs (rMRPs) had the opposite effect. Collectively, our results suggest that rapid secretion of MRPs by neutrophils at the site of infection may protect uninfected macrophages and favor a more efficient innate inflammatory response against Leishmania infection. In summary, our study reveals the critical role played by MRPs in the regulation of Leishmania infection and how this pathogen can subvert its action. PMID:24086787

The Phagocytic Function of Macrophage-Enforcing Innate Immunity and Tissue Homeostasis.

PubMed

Hirayama, Daisuke; Iida, Tomoya; Nakase, Hiroshi

2017-12-29

Macrophages are effector cells of the innate immune system that phagocytose bacteria and secrete both pro-inflammatory and antimicrobial mediators. In addition, macrophages play an important role in eliminating diseased and damaged cells through their programmed cell death. Generally, macrophages ingest and degrade dead cells, debris, tumor cells, and foreign materials. They promote homeostasis by responding to internal and external changes within the body, not only as phagocytes, but also through trophic, regulatory, and repair functions. Recent studies demonstrated that macrophages differentiate from hematopoietic stem cell-derived monocytes and embryonic yolk sac macrophages. The latter mainly give rise to tissue macrophages. Macrophages exist in all vertebrate tissues and have dual functions in host protection and tissue injury, which are maintained at a fine balance. Tissue macrophages have heterogeneous phenotypes in different tissue environments. In this review, we focused on the phagocytic function of macrophage-enforcing innate immunity and tissue homeostasis for a better understanding of the role of tissue macrophages in several pathological conditions.

HIF1α-dependent glycolysis promotes macrophage functional activities in protecting against bacterial and fungal infection.

PubMed

Li, Chunxiao; Wang, Yu; Li, Yan; Yu, Qing; Jin, Xi; Wang, Xiao; Jia, Anna; Hu, Ying; Han, Linian; Wang, Jian; Yang, Hui; Yan, Dapeng; Bi, Yujing; Liu, Guangwei

2018-02-26

Macrophages are important innate immune defense system cells in the fight against bacterial and fungal pathogenic infections. They exhibit significant plasticity, particularly with their ability to undergo functional differentiation. Additionally, HIF1α is critically involved in the functional differentiation of macrophages during inflammation. However, the role of macrophage HIF1α in protecting against different pathogenic infections remains unclear. In this study, we investigated and compared the roles of HIF1α in different macrophage functional effects of bacterial and fungal infections in vitro and in vivo. We found that bacterial and fungal infections produced similar effects on macrophage functional differentiation. HIF1α deficiency inhibited pro-inflammatory macrophage functional activities when cells were stimulated with LPS or curdlan in vitro or when mice were infected with L. monocytogenes or C. albicans in vivo, thus decreasing pro-inflammatory TNFα and IL-6 secretion associated with pathogenic microorganism survival. Alteration of glycolytic pathway activation was required for the functional differentiation of pro-inflammatory macrophages in protecting against bacterial and fungal infections. Thus, the HIF1α-dependent glycolytic pathway is essential for pro-inflammatory macrophage functional differentiation in protecting against bacterial and fungal infections.

Identification of Caspase-6 as a New Regulator of Alternatively Activated Macrophages*

PubMed Central

Yao, Yongfang; Shi, Qian; Chen, Bing; Wang, Qingsong; Li, Xinda; Li, Long; Huang, Yahong; Ji, Jianguo; Shen, Pingping

2016-01-01

Alternatively activated macrophages (AAMs) play essential roles in the promotion of tissue remodeling, vasculogenesis, and tumor progression; however, the detailed mechanisms underlying the activation of AAMs remain largely unknown. Here, by using quantitative proteomic analysis, we identified 62 proteins that were up-regulated in IL-4-induced macrophages. Among these, Caspase-6 was increased significantly. Caspase-6 is important in the apoptotic signaling pathway; however, its role in non-apoptosis is also reported. Here, we first examined the non-apoptotic role of Caspase-6 in the alternative activation of macrophages after administration of IL-4, 4T1 tumor conditional medium, or co-culture with 4T1 cells. Both treatments promoted alternative activation of RAW264.7 cells and primary macrophages, whereas disruption of caspase-6 expression and activity could markedly suppress the biomarker levels of AAMs. Overexpression of Caspase-6 could significantly promote the activation of AAMs. Importantly, we further present evidence that caspase-6 could regulate breast cancer cell invasion by modulating MMP-2 and MMP-9 expression in 4T1 tumor-associated macrophages, as ablation of protein levels or activity of caspase-6 suppressed tumor cell invasion in vitro. In conclusion, the observed results markedly expanded our views of the dynamic changes in protein composition during alternative activation of macrophages, and they revealed a critical new role of caspase-6 in regulating this cellular biological process, which suggested that caspase-6 might be a key nod molecule to regulate immunological steady-state and be a therapeutic candidate for tumor immunotherapy. PMID:27325699

Classical and alternative macrophage activation in the lung following ozone-induced oxidative stress

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

Sunil, Vasanthi R., E-mail: sunilva@pharmacy.rutgers.edu; Patel-Vayas, Kinal; Shen, Jianliang

Ozone is a pulmonary irritant known to cause oxidative stress, inflammation and tissue injury. Evidence suggests that macrophages play a role in the pathogenic response; however, their contribution depends on the mediators they encounter in the lung which dictate their function. In these studies we analyzed the effects of ozone-induced oxidative stress on the phenotype of alveolar macrophages (AM). Exposure of rats to ozone (2 ppm, 3 h) resulted in increased expression of 8-hydroxy-2′-deoxyguanosine (8-OHdG), as well as heme oxygenase-1 (HO-1) in AM. Whereas 8-OHdG was maximum at 24 h, expression of HO-1 was biphasic increasing after 3 h andmore » 48–72 h. Cleaved caspase-9 and beclin-1, markers of apoptosis and autophagy, were also induced in AM 24 h post-ozone. This was associated with increased bronchoalveolar lavage protein and cells, as well as matrix metalloproteinase (MMP)-2 and MMP-9, demonstrating alveolar epithelial injury. Ozone intoxication resulted in biphasic activation of the transcription factor, NFκB. This correlated with expression of monocyte chemotactic protein‐1, inducible nitric oxide synthase and cyclooxygenase‐2, markers of proinflammatory macrophages. Increases in arginase-1, Ym1 and galectin-3 positive anti-inflammatory/wound repair macrophages were also observed in the lung after ozone inhalation, beginning at 24 h (arginase-1, Ym1), and persisting for 72 h (galectin-3). This was associated with increased expression of pro-surfactant protein-C, a marker of Type II cell proliferation and activation, important steps in wound repair. These data suggest that both proinflammatory/cytotoxic and anti-inflammatory/wound repair macrophages are activated early in the response to ozone-induced oxidative stress and tissue injury. -- Highlights: ► Lung macrophages are highly sensitive to ozone induced oxidative stress. ► Ozone induces autophagy and apoptosis in lung macrophages. ► Proinflammatory and wound repair macrophages are activated early after ozone. ► Oxidative stress may contribute to regulating macrophage phenotype and function.« less

Forkhead-associated (FHA) Domain Containing ABC Transporter Rv1747 Is Positively Regulated by Ser/Thr Phosphorylation in Mycobacterium tuberculosis*

PubMed Central

Spivey, Vicky L.; Molle, Virginie; Whalan, Rachael H.; Rodgers, Angela; Leiba, Jade; Stach, Lasse; Walker, K. Barry; Smerdon, Stephen J.; Buxton, Roger S.

2011-01-01

One major signaling method employed by Mycobacterium tuberculosis, the causative agent of tuberculosis, is through reversible phosphorylation of proteins mediated by protein kinases and phosphatases. This study concerns one of these enzymes, the serine/threonine protein kinase PknF, that is encoded in an operon with Rv1747, an ABC transporter that is necessary for growth of M. tuberculosis in vivo and contains two forkhead-associated (FHA) domains. FHA domains are phosphopeptide recognition motifs that specifically recognize phosphothreonine-containing epitopes. Experiments to determine how PknF regulates the function of Rv1747 demonstrated that phosphorylation occurs on two specific threonine residues, Thr-150 and Thr-208. To determine the in vivo consequences of phosphorylation, infection experiments were performed in bone marrow-derived macrophages and in mice using threonine-to-alanine mutants of Rv1747 that prevent specific phosphorylation and revealed that phosphorylation positively modulates Rv1747 function in vivo. The role of the FHA domains in this regulation was further demonstrated by isothermal titration calorimetry, using peptides containing both phosphothreonine residues. FHA-1 domain mutation resulted in attenuation in macrophages highlighting the critical role of this domain in Rv1747 function. A mutant deleted for pknF did not, however, have a growth phenotype in an infection, suggesting that other kinases can fulfill its role when it is absent. This study provides the first information on the molecular mechanism(s) regulating Rv1747 through PknF-dependent phosphorylation but also indicates that phosphorylation activates Rv1747, which may have important consequences in regulating growth of M. tuberculosis. PMID:21622570

Adipose Genes Down-Regulated During Experimental Endotoxemia Are Also Suppressed in Obesity

PubMed Central

Hinkle, Christine C.; Haris, Lalarukh; Shah, Rhia; Mehta, Nehal N.; Putt, Mary E.; Reilly, Muredach P.

2012-01-01

Context: Adipose inflammation is a crucial link between obesity and its metabolic complications. Human experimental endotoxemia is a controlled model for the study of inflammatory cardiometabolic responses in vivo. Objective: We hypothesized that adipose genes down-regulated during endotoxemia would approximate changes observed with obesity-related inflammation and reveal novel candidates in cardiometabolic disease. Design, Subjects, and Intervention: Healthy volunteers (n = 14) underwent a 3 ng/kg endotoxin challenge; adipose biopsies were taken at 0, 4, 12, and 24 h for mRNA microarray. A priority list of highly down-regulated and biologically relevant genes was validated by RT-PCR in an independent sample of adipose from healthy subjects (n = 7) undergoing a subclinical 0.6 ng/kg endotoxemia protocol. Expression of validated genes was screened in adipose of lean and severely obese individuals (n = 11 per group), and cellular source was probed in cultured adipocytes and macrophages. Results: Endotoxemia (3 ng/kg) suppressed expression of 353 genes (to <67% of baseline; P < 1 × 10−5) of which 68 candidates were prioritized for validation. In low-dose (0.6 ng/kg) endotoxin validation, 22 (32%) of these 68 genes were confirmed. Functional classification revealed that many of these genes are involved in cell development and differentiation. Of validated genes, 59% (13 of 22) were down-regulated more than 1.5-fold in primary human adipocytes after treatment with endotoxin. In human macrophages, 59% (13 of 22) were up-regulated during differentiation to inflammatory M1 macrophages whereas 64% (14 of 22) were down-regulated during transition to homeostatic M2 macrophages. Finally, in obese vs. lean adipose, 91% (20 of 22) tended to have reduced expression (χ2 = 10.72, P < 0.01) with 50% (11 of 22) reaching P < 0.05 (χ2 = 9.28, P < 0.01). Conclusions: Exploration of down-regulated mRNA in adipose during human endotoxemia revealed suppression of genes involved in cell development and differentiation. A majority of candidates were also suppressed in endogenous human obesity, suggesting a potential pathophysiological role in human obesity-related adipose inflammation. PMID:22893715

Adipose genes down-regulated during experimental endotoxemia are also suppressed in obesity.

PubMed

Shah, Rachana; Hinkle, Christine C; Haris, Lalarukh; Shah, Rhia; Mehta, Nehal N; Putt, Mary E; Reilly, Muredach P

2012-11-01

Adipose inflammation is a crucial link between obesity and its metabolic complications. Human experimental endotoxemia is a controlled model for the study of inflammatory cardiometabolic responses in vivo. We hypothesized that adipose genes down-regulated during endotoxemia would approximate changes observed with obesity-related inflammation and reveal novel candidates in cardiometabolic disease. Healthy volunteers (n = 14) underwent a 3 ng/kg endotoxin challenge; adipose biopsies were taken at 0, 4, 12, and 24 h for mRNA microarray. A priority list of highly down-regulated and biologically relevant genes was validated by RT-PCR in an independent sample of adipose from healthy subjects (n = 7) undergoing a subclinical 0.6 ng/kg endotoxemia protocol. Expression of validated genes was screened in adipose of lean and severely obese individuals (n = 11 per group), and cellular source was probed in cultured adipocytes and macrophages. Endotoxemia (3 ng/kg) suppressed expression of 353 genes (to <67% of baseline; P < 1 × 10(-5)) of which 68 candidates were prioritized for validation. In low-dose (0.6 ng/kg) endotoxin validation, 22 (32%) of these 68 genes were confirmed. Functional classification revealed that many of these genes are involved in cell development and differentiation. Of validated genes, 59% (13 of 22) were down-regulated more than 1.5-fold in primary human adipocytes after treatment with endotoxin. In human macrophages, 59% (13 of 22) were up-regulated during differentiation to inflammatory M1 macrophages whereas 64% (14 of 22) were down-regulated during transition to homeostatic M2 macrophages. Finally, in obese vs. lean adipose, 91% (20 of 22) tended to have reduced expression (χ(2) = 10.72, P < 0.01) with 50% (11 of 22) reaching P < 0.05 (χ(2) = 9.28, P < 0.01). Exploration of down-regulated mRNA in adipose during human endotoxemia revealed suppression of genes involved in cell development and differentiation. A majority of candidates were also suppressed in endogenous human obesity, suggesting a potential pathophysiological role in human obesity-related adipose inflammation.

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

PubMed Central

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

Macrophage polarization in response to ECM coated polypropylene mesh

PubMed Central

Wolf, MT; Dearth, CL; Ranallo, CA; LoPresti, S; Carey, LE; Daly, KA; Brown, BN; Badylak, SF

2015-01-01

The host response to implanted biomaterials is a highly regulated process that influences device functionality and clinical outcome. Non-degradable biomaterials, such as knitted polypropylene mesh, frequently elicit a chronic foreign body reaction with resultant fibrosis. Previous studies have shown that an extracellular matrix (ECM) hydrogel coating of polypropylene mesh reduces the intensity of the foreign body reaction, though the mode of action is unknown. Macrophage participation plays a key role in the development of the foreign body reaction to biomaterials, and therefore the present study investigated macrophage polarization following mesh implantation. Spatiotemporal analysis of macrophage polarization was conducted in response to uncoated polypropylene mesh and mesh coated with hydrated and dry forms of ECM hydrogels derived from either dermis or urinary bladder. Pro-inflammatory M1 macrophages (CD86+/CD68+), alternatively activated M2 macrophages (CD206+/CD68+), and foreign body giant cells were quantified between 3-35 days. Uncoated polypropylene mesh elicited a dominant M1 response at the mesh fiber surface, which was decreased by each ECM coating type beginning at 7 days. The diminished M1 response was accompanied by a reduction in the number of foreign body giant cells at 14 and 35 days, though there was a minimal effect upon the number of M2 macrophages at any time. These results show that ECM coatings attenuate the M1 macrophage response and increase the M2/M1 ratio to polypropylene mesh in vivo. PMID:24856104

Resident alveolar macrophages are master regulators of arrested alveolarization in experimental bronchopulmonary dysplasia.

PubMed

Kalymbetova, Tatiana V; Selvakumar, Balachandar; Rodríguez-Castillo, José Alberto; Gunjak, Miša; Malainou, Christina; Heindl, Miriam Ruth; Moiseenko, Alena; Chao, Cho-Ming; Vadász, István; Mayer, Konstantin; Lohmeyer, Jürgen; Bellusci, Saverio; Böttcher-Friebertshäuser, Eva; Seeger, Werner; Herold, Susanne; Morty, Rory E

2018-06-01

Trophic functions for macrophages are emerging as key mediators of developmental processes, including bone, vessel, and mammary gland development. Yolk sac-derived macrophages mature in the distal lung shortly after birth. Myeloid-lineage macrophages are recruited to the lung and are activated under pathological conditions. These pathological conditions include bronchopulmonary dysplasia (BPD), a common complication of preterm birth characterized by stunted lung development, where the formation of alveoli is blocked. No study has addressed causal roles for immune cells in lung alveolarization. We employed antibody-based and transgenic death receptor-based depletion approaches to deplete or prevent lung recruitment of immune cell populations in a hyperoxia-based mouse model of BPD. Neither neutrophils nor exudate macrophages (which might include lung interstitial macrophages) contributed to structural perturbations to the lung that were provoked by hyperoxia; however, cells of the Csf1r-expressing monocyte/macrophage lineage were implicated as causal mediators of stunted lung development. We propose that resident alveolar macrophages differentiate into a population of CD45 + CD11c + SiglecF + CD11b + CD68 + MHCII + cells, which are activated by hyperoxia, and contribute to disturbances to the structural development of the immature lung. This is the first report that causally implicates immune cells in pathological disturbances to postnatal lung organogenesis. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Glycogen synthase kinase 3β promotes liver innate immune activation by restraining AMP-activated protein kinase activation.

PubMed

Zhou, Haoming; Wang, Han; Ni, Ming; Yue, Shi; Xia, Yongxiang; Busuttil, Ronald W; Kupiec-Weglinski, Jerzy W; Lu, Ling; Wang, Xuehao; Zhai, Yuan

2018-07-01

Glycogen synthase kinase 3β (Gsk3β [Gsk3b]) is a ubiquitously expressed kinase with distinctive functions in different types of cells. Although its roles in regulating innate immune activation and ischaemia and reperfusion injuries (IRIs) have been well documented, the underlying mechanisms remain ambiguous, in part because of the lack of cell-specific tools in vivo. We created a myeloid-specific Gsk3b knockout (KO) strain to study the function of Gsk3β in macrophages in a murine liver partial warm ischaemia model. Compared with controls, myeloid Gsk3b KO mice were protected from IRI, with diminished proinflammatory but enhanced anti-inflammatory immune responses in livers. In bone marrow-derived macrophages, Gsk3β deficiency resulted in an early reduction of Tnf gene transcription but sustained increase of Il10 gene transcription on Toll-like receptor 4 stimulation in vitro. These effects were associated with enhanced AMP-activated protein kinase (AMPK) activation, which led to an accelerated and higher level of induction of the novel innate immune negative regulator small heterodimer partner (SHP [Nr0b2]). The regulatory function of Gsk3β on AMPK activation and SHP induction was confirmed in wild-type bone marrow-derived macrophages with a Gsk3 inhibitor. Furthermore, we found that this immune regulatory mechanism was independent of Gsk3β Ser9 phosphorylation and the phosphoinositide 3-kinase-Akt signalling pathway. In vivo, myeloid Gsk3β deficiency facilitated SHP upregulation by ischaemia-reperfusion in liver macrophages. Treatment of Gsk3b KO mice with either AMPK inhibitor or SHP small interfering RNA before the onset of liver ischaemia restored liver proinflammatory immune activation and IRI in these otherwise protected hosts. Additionally, pharmacological activation of AMPK protected wild-type mice from liver IRI, with reduced proinflammatory immune activation. Inhibition of the AMPK-SHP pathway by liver ischaemia was demonstrated in tumour resection patients. Gsk3β promotes innate proinflammatory immune activation by restraining AMPK activation. Glycogen synthase kinase 3β promotes macrophage inflammatory activation by inhibiting the immune regulatory signalling of AMP-activated protein kinase and the induction of small heterodimer partner. Therefore, therapeutic targeting of glycogen synthase kinase 3β enhances innate immune regulation and protects liver from ischaemia and reperfusion injury. Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Expression of CXCR1 (IL-8 receptor A) in splenic, peritoneal macrophages and resident bone marrow cells after acute live or heat killed Staphylococcus aureus stimulation in mice.

PubMed

Bishayi, Biswadev; Nandi, Ajeya; Dey, Rajen; Adhikary, Rana

2017-08-01

Literature reveals that interaction with live Staphylococcus aureus (S. aureus) or heat killed S. aureus (HKSA) promotes secretion of CXCL-8 or interleukin-8 (IL-8) from leukocytes, however, the expressions of CXCR1 in murine splenic (SPM), peritoneal macrophages (PM) and resident fresh bone marrow cells (FBMC) have not been identified. Currently, very few studies are available on the functional characterization of CXCR1 in mouse macrophage subtypes and its modulation in relation to acute S. aureus infection. SPM, PM and FBMCs were infected with viable S. aureus or stimulated with HKSA in presence and absence of anti-CXCR1 antibody in this study. We reported here that CXCR1 was not constitutively expressed by macrophage subtypes and the receptor was induced only after S. aureus stimulation. The CXCR1 band was found specific as we compared with human polymorphonuclear neutrophils (PMNs) as a positive control (data not shown). Although, we did not show that secreted IL-8 from S. aureus-infected macrophages promotes migration of PMNs. Blocking of cell surface CXCR1 decreases the macrophage's ability to clear staphylococcal infection, attenuates proinflammatory cytokine production and the increased catalase and decreased superoxide dismutase (SOD) enzymes of the bacteria might indicate their role in scavenging macrophage derived hydrogen peroxide (H 2 O 2 ). The decreased levels of cytokines due to CXCR1 blockade before S. aureus infection appear to regulate the killing of bacteria by destroying H 2 O 2 and nitric oxide (NO). Moreover, functional importance of macrophage subpopulation heterogeneity might be important in designing new effective approaches to limit S. aureus infection induced inflammation and cytotoxicity. Copyright © 2017 Elsevier Ltd. All rights reserved.

MicroRNA-155 Is Required for Mycobacterium bovis BCG-Mediated Apoptosis of Macrophages

PubMed Central

Ghorpade, Devram Sampat; Leyland, Rebecca; Kurowska-Stolarska, Mariola; Patil, Shripad A.

2012-01-01

Pathogenic mycobacteria, including Mycobacterium tuberculosis and Mycobacterium bovis, cause significant morbidity and mortality worldwide. However, the vaccine strain Mycobacterium bovis BCG, unlike virulent strains, triggers extensive apoptosis of infected macrophages, a step necessary for the elicitation of robust protective immunity. We here demonstrate that M. bovis BCG triggers Toll-like receptor 2 (TLR2)-dependent microRNA-155 (miR-155) expression, which involves signaling cross talk among phosphatidylinositol 3-kinase (PI3K), protein kinase Cδ (PKCδ), and mitogen-activated protein kinases (MAPKs) and recruitment of NF-κB and c-ETS to miR-155 promoter. Genetic and signaling perturbations presented the evidence that miR-155 regulates PKA signaling by directly targeting a negative regulator of PKA, protein kinase inhibitor alpha (PKI-α). Enhanced activation of PKA signaling resulted in the generation of PKA C-α; phosphorylation of MSK1, cyclic AMP response element binding protein (CREB), and histone H3; and recruitment of phospho-CREB to the apoptotic gene promoters. The miR-155-triggered activation of caspase-3, BAK1, and cytochrome c translocation involved signaling integration of MAPKs and epigenetic or posttranslational modification of histones or CREB. Importantly, M. bovis BCG infection-induced apoptosis was severely compromised in macrophages derived from miR-155 knockout mice. Gain-of-function and loss-of-function studies validated the requirement of miR-155 for M. bovis BCG's ability to trigger apoptosis. Overall, M. bovis BCG-driven miR-155 dictates cell fate decisions of infected macrophages, strongly implicating a novel role for miR-155 in orchestrating cellular reprogramming during immune responses to mycobacterial infection. PMID:22473996

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

PubMed

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 metabolic enzyme fructose-1,6-bisphosphate aldolase acts as a transcriptional regulator in pathogenic Francisella.

PubMed

Ziveri, Jason; Tros, Fabiola; Guerrera, Ida Chiara; Chhuon, Cerina; Audry, Mathilde; Dupuis, Marion; Barel, Monique; Korniotis, Sarantis; Fillatreau, Simon; Gales, Lara; Cahoreau, Edern; Charbit, Alain

2017-10-11

The enzyme fructose-bisphosphate aldolase occupies a central position in glycolysis and gluconeogenesis pathways. Beyond its housekeeping role in metabolism, fructose-bisphosphate aldolase has been involved in additional functions and is considered as a potential target for drug development against pathogenic bacteria. Here, we address the role of fructose-bisphosphate aldolase in the bacterial pathogen Francisella novicida. We demonstrate that fructose-bisphosphate aldolase is important for bacterial multiplication in macrophages in the presence of gluconeogenic substrates. In addition, we unravel a direct role of this metabolic enzyme in transcription regulation of genes katG and rpoA, encoding catalase and an RNA polymerase subunit, respectively. We propose a model in which fructose-bisphosphate aldolase participates in the control of host redox homeostasis and the inflammatory immune response.The enzyme fructose-bisphosphate aldolase (FBA) plays central roles in glycolysis and gluconeogenesis. Here, Ziveri et al. show that FBA of the pathogen Francisella novicida acts, in addition, as a transcriptional regulator and is important for bacterial multiplication in macrophages.

[THE INCONSISTENCIES OF REGULATION OF METABOLISM IN PHYLOGENESIS AT THREE LEVELS OF "RELATIVE BIOLOGICAL PERFECTION": ETIOLOGY OF METABOLIC PANDEMICS].

PubMed

Titov, V N

2015-11-01

The regulation of metabolism in vivo can be comprehended by considering stages of becoming inphylogenesis of humoral, hormonal, vegetative regulators separately: at the level of cells; in paracrin-regulated cenosises of cells; organs and systems under open blood circulation and closed system of blood flow. The levels of regulations formed at different stages of phylogenesis. Their completion occurred at achievement of "relative biological perfection". Only this way need of cells in functional, structural interaction and forming of multicellular developed. The development of organs and systems of organs also completed at the level of "relative biological perfection". From the same level the third stage of becoming of regulation of metabolism at the level of organism started. When three conditions of "relative biological perfection" achieved consequently at level in vivo are considered in species Homo sapiens using system approach it is detected that "relative biological perfection" in vivo is accompanied by different inconsistencies of regulation of metabolism. They are etiologic factors of "metabolic pandemics ". The inconsistencies (etiological factors) are consider as exemplified by local (at the level of paracrin-regulated cenosises of cells) and system (at the level of organism) regulation of biological reaction metabolism-microcirculation that results in dysfunction of target organs and development of pathogenesis of essential metabolic arterial hypertension. The article describes phylogenetic difference between visceral fatty cells and adpocytes, regulation of metabolism by phylogenetically late insulin, reaction of albumin at increasing of content of unesterified fatty acids in blood plasma, difference of function of resident macrophage and monocytes-macrophages in pathogenesis of atherosclerosis, metabolic syndrome, insulin resistance, obesity, under diabetes mellitus and essential metabolic arterial hypertension.

The thyroid hormone triiodothyronine controls macrophage maturation and functions: protective role during inflammation.

PubMed

Perrotta, Cristiana; Buldorini, Marcella; Assi, Emma; Cazzato, Denise; De Palma, Clara; Clementi, Emilio; Cervia, Davide

2014-01-01

The endocrine system participates in regulating macrophage maturation, although little is known about the modulating role of the thyroid hormones. In vitro results demonstrate a negative role of one such hormone, triiodothyronine (T3), in triggering the differentiation of bone marrow-derived monocytes into unpolarized macrophages. T3-induced macrophages displayed a classically activated (M1) signature. A T3-induced M1-priming effect was also observed on polarized macrophages because T3 reverses alternatively activated (M2) activation, whereas it enhances that of M1 cells. In vivo, circulating T3 increased the content of the resident macrophages in the peritoneal cavity, whereas it reduced the content of the recruited monocyte-derived cells. Of interest, T3 significantly protected mice against endotoxemia induced by lipopolysaccharide i.p. injection; in these damaged animals, decreased T3 levels increased the recruited (potentially damaging) cells, whereas restoring T3 levels decreased recruited and increased resident (potentially beneficial) cells. These data suggest that the anti-inflammatory effect of T3 is coupled to the modulation of peritoneal macrophage content, in a context not fully explained by the M1/M2 framework. Thyroid hormone receptor expression analysis and the use of different thyroid hormone receptor antagonists suggest thyroid hormone receptor β1 as the major player mediating T3 effects on macrophages. The novel homeostatic link between thyroid hormones and the pathophysiological role of macrophages opens new perspectives on the interactions between the endocrine and immune systems. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Trichomonas vaginalis Induces Production of Proinflammatory Cytokines in Mouse Macrophages Through Activation of MAPK and NF-κB Pathways Partially Mediated by TLR2

PubMed Central

Li, Ling; Li, Xin; Gong, Pengtao; Zhang, Xichen; Yang, Zhengtao; Yang, Ju; Li, Jianhua

2018-01-01

Trichomoniasis, caused by Trichomonas vaginalis infection, is the most prevalent sexually transmitted disease in female and male globally. However, the mechanisms by innate immunity against T. vaginalis infection have not been fully elucidated. Toll-like receptor2 (TLR2) has been shown to be involved in pathogen recognition, innate immunity activation, and inflammatory response to the pathogens. Nonetheless, the function of TLR2 against T. vaginalis remains unclear. In the present study, we investigated the role of TLR2 in mouse macrophages against T. vaginalis. RT-qPCR analysis revealed that T. vaginalis stimulation increased the gene expression of TLR2 in wild-type (WT) mouse macrophages. T. vaginalis also induced the secretion of IL-6, TNF-α, and IFN-γ in WT mouse macrophages, and the expression of these cytokines significantly decreased in TLR2-/- mouse macrophages and in WT mouse macrophages pretreated with MAPK inhibitors SB203580 (p38) and PD98059 (ERK). Western blot analysis demonstrated that T. vaginalis stimulation induced the activation of p38, ERK, and p65 NF-κB signal pathways in WT mouse macrophages, and the phosphorylation of p38, ERK, and p65 NF-κB significantly decreased in TLR2-/- mouse macrophages. Taken together, our data suggested that T. vaginalis may regulates proinflammatory cytokines production by activation of p38, ERK, and NF-κB p65 signal pathways via TLR2 in mouse macrophages. TLR2 might be involved in the defense and elimination of T. vaginalis infection. PMID:29692771

Trichomonas vaginalis Induces Production of Proinflammatory Cytokines in Mouse Macrophages Through Activation of MAPK and NF-κB Pathways Partially Mediated by TLR2.

PubMed

Li, Ling; Li, Xin; Gong, Pengtao; Zhang, Xichen; Yang, Zhengtao; Yang, Ju; Li, Jianhua

2018-01-01

Trichomoniasis, caused by Trichomonas vaginalis infection, is the most prevalent sexually transmitted disease in female and male globally. However, the mechanisms by innate immunity against T. vaginalis infection have not been fully elucidated. Toll-like receptor2 (TLR2) has been shown to be involved in pathogen recognition, innate immunity activation, and inflammatory response to the pathogens. Nonetheless, the function of TLR2 against T. vaginalis remains unclear. In the present study, we investigated the role of TLR2 in mouse macrophages against T. vaginalis . RT-qPCR analysis revealed that T. vaginalis stimulation increased the gene expression of TLR2 in wild-type (WT) mouse macrophages. T. vaginalis also induced the secretion of IL-6, TNF-α, and IFN-γ in WT mouse macrophages, and the expression of these cytokines significantly decreased in TLR 2-/- mouse macrophages and in WT mouse macrophages pretreated with MAPK inhibitors SB203580 (p38) and PD98059 (ERK). Western blot analysis demonstrated that T. vaginalis stimulation induced the activation of p38, ERK, and p65 NF-κB signal pathways in WT mouse macrophages, and the phosphorylation of p38, ERK, and p65 NF-κB significantly decreased in TLR2 -/- mouse macrophages. Taken together, our data suggested that T. vaginalis may regulates proinflammatory cytokines production by activation of p38, ERK, and NF-κB p65 signal pathways via TLR2 in mouse macrophages. TLR2 might be involved in the defense and elimination of T. vaginalis infection.

Regulation of iNOS function and cellular redox state by macrophage Gch1 reveals specific requirements for tetrahydrobiopterin in NRF2 activation

PubMed Central

McNeill, Eileen; Crabtree, Mark J.; Sahgal, Natasha; Patel, Jyoti; Chuaiphichai, Surawee; Iqbal, Asif J.; Hale, Ashley B.; Greaves, David R.; Channon, Keith M.

2015-01-01

Inducible nitric oxide synthase (iNOS) is a key enzyme in the macrophage inflammatory response, which is the source of nitric oxide (NO) that is potently induced in response to proinflammatory stimuli. However, the specific role of NO production, as distinct from iNOS induction, in macrophage inflammatory responses remains unproven. We have generated a novel mouse model with conditional deletion of Gch1, encoding GTP cyclohydrolase 1 (GTPCH), an essential enzyme in the biosynthesis of tetrahydrobiopterin (BH4) that is a required cofactor for iNOS NO production. Mice with a floxed Gch1 allele (Gch1fl/fl) were crossed with Tie2cre transgenic mice, causing Gch1 deletion in leukocytes (Gch1fl/flTie2cre). Macrophages from Gch1fl/flTie2cre mice lacked GTPCH protein and de novo biopterin biosynthesis. When activated with LPS and IFNγ, macrophages from Gch1fl/flTie2cre mice induced iNOS protein in a manner indistinguishable from wild-type controls, but produced no detectable NO, as judged by L-citrulline production, EPR spin trapping of NO, and by nitrite accumulation. Incubation of Gch1fl/flTie2cre macrophages with dihydroethidium revealed significantly increased production of superoxide in the presence of iNOS expression, and an iNOS-independent, BH4-dependent increase in other ROS species. Normal BH4 levels, nitric oxide production, and cellular redox state were restored by sepiapterin, a precursor of BH4 production by the salvage pathway, demonstrating that the effects of BH4 deficiency were reversible. Gch1fl/flTie2cre macrophages showed only minor alterations in cytokine production and normal cell migration, and minimal changes in basal gene expression. However, gene expression analysis after iNOS induction identified 78 genes that were altered between wild-type and Gch1fl/flTie2cre macrophages. Pathway analysis identified decreased NRF2 activation, with reduced induction of archetypal NRF2 genes (gclm, prdx1, gsta3, nqo1, and catalase) in BH4-deficient Gch1fl/flTie2cre macrophages. These findings identify BH4-dependent iNOS regulation and NO generation as specific requirements for NRF2-dependent responses in macrophage inflammatory activation. PMID:25451639

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

PubMed

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

2015-09-08

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

Bioelectric modulation of macrophage polarization

NASA Astrophysics Data System (ADS)

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

2016-02-01

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

Tissue-Resident Macrophages in Fungal Infections.

PubMed

Xu, Shengjie; Shinohara, Mari L

2017-01-01

Invasive fungal infections result in high morbidity and mortality. Host organs targeted by fungal pathogens vary depending on the route of infection and fungal species encountered. Cryptococcus neoformans infects the respiratory tract and disseminates throughout the central nervous system. Candida albicans infects mucosal tissues and the skin, and systemic Candida infection in rodents has a tropism to the kidney. Aspergillus fumigatus reaches distal areas of the lung once inhaled by the host. Across different tissues in naïve hosts, tissue-resident macrophages (TRMs) are one of the most populous cells of the innate immune system. Although they function to maintain homeostasis in a tissue-specific manner during steady state, TRMs may function as the first line of defense against invading pathogens and may regulate host immune responses. Thus, in any organs, TRMs are uniquely positioned and specifically programmed to function. This article reviews the current understanding of the roles of TRMs during major fungal infections.

CXCL10 Controls Inflammatory Pain via Opioid Peptide-Containing Macrophages in Electroacupuncture

PubMed Central

Wang, Ying; Gehringer, Rebekka; Mousa, Shaaban A.; Hackel, Dagmar; Brack, Alexander; Rittner, Heike L.

2014-01-01

Acupuncture is widely used for pain treatment in patients with osteoarthritis or low back pain, but molecular mechanisms remain largely enigmatic. In the early phase of inflammation neutrophilic chemokines direct opioid-containing neutrophils in the inflamed tissue and stimulate opioid peptide release and antinociception. In this study the molecular pathway and neuroimmune connections in complete Freund's adjuvant (CFA)-induced hind paw inflammation and electroacupuncture for peripheral pain control were analyzed. Free moving Wistar rats with hind paw inflammation were treated twice with electroacupuncture at GB30 (Huan Tiao - gall bladder meridian) (day 0 and 1) and analyzed for mechanical and thermal nociceptive thresholds. The cytokine profiles as well as the expression of opioid peptides were quantified in the inflamed paw. Electroacupuncture elicited long-term antinociception blocked by local injection of anti-opioid peptide antibodies (beta-endorphin, met-enkephalin, dynorphin A). The treatment altered the cytokine profile towards an anti-inflammatory pattern but augmented interferon (IFN)-gamma and the chemokine CXCL10 (IP-10: interferon gamma-inducible protein) protein and mRNA expression with concomitant increased numbers of opioid peptide-containing CXCR3+ macrophages. In rats with CFA hind paw inflammation without acupuncture repeated injection of CXCL10 triggered opioid-mediated antinociception and increase opioid-containing macrophages. Conversely, neutralization of CXCL10 time-dependently decreased electroacupuncture-induced antinociception and the number of infiltrating opioid peptide-expressing CXCR3+ macrophages. In summary, we describe a novel function of the chemokine CXCL10 - as a regulator for an increase of opioid-containing macrophages and antinociceptive mediator in inflammatory pain and as a key chemokine regulated by electroacupuncture. PMID:24732949

Angiogenic Deficiency and Adipose Tissue Dysfunction Are Associated with Macrophage Malfunction in SIRT1−/− Mice

PubMed Central

Xu, Fen; Burk, David; Gao, Zhanguo; Yin, Jun; Zhang, Xia

2012-01-01

The histone deacetylase sirtuin 1 (SIRT1) inhibits adipocyte differentiation and suppresses inflammation by targeting the transcription factors peroxisome proliferator-activated receptor γ and nuclear factor κB. Although this suggests that adiposity and inflammation should be enhanced when SIRT1 activity is inactivated in the body, this hypothesis has not been tested in SIRT1 null (SIRT1−/−) mice. In this study, we addressed this issue by investigating the adipose tissue in SIRT1−/− mice. Compared with their wild-type littermates, SIRT1 null mice exhibited a significant reduction in body weight. In adipose tissue, the average size of adipocytes was smaller, the content of extracellular matrix was lower, adiponectin and leptin were expressed at 60% of normal level, and adipocyte differentiation was reduced. All of these changes were observed with a 50% reduction in capillary density that was determined using a three-dimensional imaging technique. Except for vascular endothelial growth factor, the expression of several angiogenic factors (Pdgf, Hgf, endothelin, apelin, and Tgf-β) was reduced by about 50%. Macrophage infiltration and inflammatory cytokine expression were 70% less in the adipose tissue of null mice and macrophage differentiation was significantly inhibited in SIRT1−/− mouse embryonic fibroblasts in vitro. In wild-type mice, macrophage deletion led to a reduction in vascular density. These data suggest that SIRT1 controls adipose tissue function through regulation of angiogenesis, whose deficiency is associated with macrophage malfunction in SIRT1−/− mice. The study supports the concept that inflammation regulates angiogenesis in the adipose tissue. PMID:22315447

Endogenous hydrogen sulfide regulates histone demethylase JMJD3-mediated inflammatory response in LPS-stimulated macrophages and in a mouse model of LPS-induced septic shock.

PubMed

Liu, Siyu; Wang, Xiling; Pan, Lilong; Wu, Weijun; Yang, Di; Qin, Ming; Jia, Wanwan; Xiao, Chenxi; Long, Fen; Ge, Junbo; Liu, Xinhua; Zhu, YiZhun

2018-03-01

Overproduction of inflammatory mediators contributes to uncontrolled inflammation during endotoxin shock. Cystathionine-γ-lyase (CSE), an enzyme involved in hydrogen sulfide (H 2 S) biosynthesis, has potential anti-inflammatory activity in a variety of inflammatory diseases. Jumonji domain-containing protein 3 (JMJD3), a histone 3 Lys27 (H3K27) demethylase, has been implicated in macrophage activation, but its function in CSE-mediated anti-inflammatory activities remains unknown. In the present study CSE was found to be upregulated in macrophages and mouse lipopolysaccharide (LPS) challenge models. LPS stimulation also enhanced the activation of JMJD3 and decreased H3K27me3 levels. JMJD3 knockdown upregulated H3K27me3 levels and attenuated the LPS-mediated inflammatory response. CSE knockout amplified the inflammatory cascade by increasing JMJD3 expression in septic mice. Similarly, enhanced production of inflammatory mediators by macrophages was mitigated by CSE overexpression via inhibition of JMJD3 expression. This is the first report indicating that inflammation enhanced CSE/H 2 S system biosynthesis, that in turn attenuated the LPS-triggered inflammatory response by regulating JMJD3 expression. Thus, the CSE/H 2 S system represents an epigenetic-based modification mechanism to prevent uncontrolled inflammation. Copyright © 2017 Elsevier Inc. All rights reserved.

Circadian Clearance of a Fungal Pathogen from the Lung Is Not Based on Cell-intrinsic Macrophage Rhythms.

PubMed

Chen, Shan; Fuller, Kevin K; Dunlap, Jay C; Loros, Jennifer J

2018-02-01

Circadian rhythms govern immune cell function, giving rise to time-of-day variation in the recognition and clearance of bacterial or viral pathogens; to date, however, no such regulation of the host-fungal interaction has been described. In this report, we use murine models to explore circadian control of either fungal-macrophage interactions in vitro or pathogen clearance from the lung in vivo. First, we show that expression of the important fungal pattern recognition receptor Dectin-1 ( clec7a), from either bone marrow-derived or peritoneum-derived macrophages, is not under circadian regulation at either the level of transcript or cell surface protein expression. Consistent with this finding, the phagocytic activity of macrophages in culture against spores of the pathogen Aspergillus fumigatus also did not vary over time. To account for the multiple cell types and processes that may be coordinated in a circadian fashion in vivo, we examined the clearance of A. fumigatus from the lungs of immunocompetent mice. Interestingly, animals inoculated at night demonstrated a 2-fold enhancement in clearance compared with animals inoculated in the morning. Taken together, our data suggest that while molecular recognition of fungi by immune cells may not be circadian, other processes in vivo may still allow for time-of-day differences in fungal clearance from the lung.

Gremlin-1 inhibits macrophage migration inhibitory factor-dependent monocyte function and survival.

PubMed

Müller, Iris I; Chatterjee, Madhumita; Schneider, Martina; Borst, Oliver; Seizer, Peter; Schönberger, Tanja; Vogel, Sebastian; Müller, Karin A L; Geisler, Tobias; Lang, Florian; Langer, Harald; Gawaz, Meinrad

2014-10-20

Monocyte migration and their differentiation into macrophages critically regulate vascular inflammation and atherogenesis and are governed by macrophage migration inhibitory factor (MIF). Gremlin-1 binds to MIF. Current experimental evidences present Gremlin-1 as a potential physiological agent that might counter-regulate the inflammatory attributes of MIF. We found that Gremlin-1 inhibited MIF-dependent monocyte migration and adhesion to activated endothelial cells in flow chamber perfusion assay in vitro and to the injured carotid artery of WT and ApoE-/- mice in vivo as deciphered by intravital microscopy. Intravenous administration of Gremlin-1, but not of control protein, significantly reduced leukocyte recruitment towards the inflamed carotid artery of ApoE-/- mice. Besides, leukocytes from MIF-/- when administered into ApoE-/- mice showed lesser adhesion as compared to wild type. In the presence of Gremlin-1 however, adhesion of wild type, but not of MIF-/- leukocytes, to the carotid artery was significantly inhibited as compared to control. Gremlin-1 also inhibited the MIF-induced differentiation of monocytes into macrophages. Gremlin-1 substantially inhibited the anti-apoptotic impact of MIF on monocytes against BH3 mimetic ABT-737-induced apoptosis as verified by Annexin V-binding, caspase 3 activity, and mitochondrial depolarization. Therefore Gremlin-1 can modulate MIF dependent monocyte adhesion, migration, differentiation and survival. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Interferon-γ acts as a regulator in the trade-off between phagocytosis and production performance in dwarf chickens.

PubMed

Yuan, Yitong; Liu, Shunqi; Zhao, Yue; Lian, Ling; Lian, Zhengxing

2018-01-01

Interferon-γ (IFN-γ) is critical for innate and adaptive immunity against viral and bacterial infections. IFN-γ reportedly affects the phagocytic ability of monocytes and macrophages as well as regulates pituitary function in humans and mice. The present study analyzed the impact of IFN-γ on monocyte and macrophage phagocytosis, production performance, and pituitary function in vivo and in vitro (in dwarf chickens). IFN-γ was injected into dwarf chickens through a vein, and then, the laying rate, average egg weight, and levels of follicle-stimulating hormone (FSH) and IFN-γ were measured in treatment and control groups. For the in vitro experiment, the pituitary tissues were supplemented with IFN-γ, and the mRNA expression levels of follicle-stimulating hormone beta subunit ( FSH-β ), interferon gamma receptor 1 ( IFNGR 1), and interferon gamma receptor 2 ( IFNGR 2) in the pituitary were assessed. Monocyte and macrophage phagocytosis product (PP) was decreased by IFN-γ treatment in a dose-dependent manner in vitro. In the in vivo experiment, the level of IFN-γ in the treatment group was higher than that in the control group at 7 d ( P  < 0.05), 14 d ( P  < 0.01), and 21 d ( P  < 0.01) post-injection. Compared with the control group, monocyte and macrophage PP was lower in the treatment group after injection ( P  < 0.01). The laying rate was higher in the treatment group than in the control group at 2 and 3 wk post-injection ( P  < 0.05). There was a significant difference between the treatment and control groups in the levels of FSH at 1, 3, 7, and 14 d post-injection ( P  < 0.01). In the in vitro experiment, increased mRNA expression levels of FSH-β , IFNGR 1, and IFNGR 2 were observed in the treatment group after stimulation with 100 U/mL IFN-γ for 24 h compared to those in the control group ( P  < 0.05). IFN-γ inhibited the phagocytosis of monocytes and macrophages; up-regulated the mRNA expression levels of the FSH-β , IFNGR 1, and IFNGR 2; enhanced the secretion of FSH; and improved the laying rate. IFN-γ might be an important regulator in the trade-off between the immune effect and production performance in dwarf chickens.

Androgen-androgen receptor system improves chronic inflammatory conditions by suppressing monocyte chemoattractant protein-1 gene expression in adipocytes via transcriptional regulation

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

Morooka, Nobukatsu, E-mail: amorooka@gunma-u.ac.jp; Ueguri, Kei; Yee, Karen Kar Lye

Age-related decreases in sex hormones are closely related to chronic inflammation in obesity and metabolic diseases. Particularly, the molecular basis of androgen activity in regulating inflammation and controlling metabolism remains largely unknown. Obese adipocytes secrete monocyte chemoattractant protein-1 (MCP-1), a key chemokine that promotes the infiltration of monocytes/macrophages into adipose tissue, thereby leading to metabolic disorders. Here, we studied the role of androgen-androgen receptor (AR) action in regulating MCP-1 expression in adipose tissue. We observed the induction of Mcp-1 expression in 3T3-L1 adipocytes co-cultured with RAW264.7 macrophages. Additionally, Mcp-1 expression was upregulated by culturing in conditioned medium derived from inflammatorymore » macrophages (M1-Mφ) containing tumor necrosis factor-alpha (TNF-α). We found that sex hormones downregulated TNF-α-induced Mcp-1 and interleukin (Il)-6 expression in 3T3-L1 adipocytes. Furthermore, luciferase-reporter analysis indicated that MCP-1 promoter activity was predominantly suppressed by dihydrotestosterone (DHT)-AR interactions through functional canonical nuclear factor-kappa B (NF-κB) sites, whereas non-canonical NF-κB site containing important flanking sequences exhibited minor contributions to DHT-AR transcriptional repression. These findings suggested that androgen-AR suppressed obesity-induced chronic inflammation in adipose tissue. - Highlights: • DHT, non-aromatizable androgen suppresses Mcp-1 expression in adipocytes. • Mcp-1 transcription was negatively regulated by DHT-AR action. • DHT-AR selectively regulates Mcp-1 transcription through distinct NF-κB sites.« less

Mutant monocyte chemoattractant protein 1 protein attenuates migration of and inflammatory cytokine release by macrophages exposed to orthopedic implant wear particles.

PubMed

Yao, Zhenyu; Keeney, Michael; Lin, Tzu-Hua; Pajarinen, Jukka; Barcay, Katherine; Waters, Heather; Egashira, Kensuke; Yang, Fan; Goodman, Stuart

2014-09-01

Wear particles generated from total joint replacements can stimulate macrophages to release chemokines, such as monocyte chemoattractant protein 1 (MCP-1), which is the most important chemokine regulating systemic and local cell trafficking and infiltration of monocyte/macrophages in chronic inflammation. One possible strategy to curtail the adverse events associated with wear particles is to mitigate migration and activation of monocyte/macrophages. The purpose of this study is to modulate the adverse effects of particulate biomaterials and inflammatory stimuli such as endotoxin by interfering with the biological effects of the chemokine MCP-1. In the current study, the function of MCP-1 was inhibited by the mutant MCP-1 protein called 7ND, which blocks its receptor, the C-C chemokine receptor type 2 (CCR2) on macrophages. Addition of 7ND decreased MCP-1-induced migration of THP-1 cells in cell migration experiments in a dose-dependent manner. Conditioned media from murine macrophages exposed to clinically relevant polymethylmethacrylate (PMMA) particles with/without endotoxin [lipopolysaccharide (LPS)] had a chemotactic effect on human macrophages, which was decreased dramatically by 7ND. 7ND demonstrated no adverse effects on the viability of macrophages, and the capability of mesenchymal stem cells (MSCs) to form bone at the doses tested. Finally, proinflammatory cytokine production was mitigated when macrophages were exposed to PMMA particles with/without LPS in the presence of 7ND. Our studies confirm that the MCP-1 mutant protein 7ND can decrease macrophage migration and inflammatory cytokine release without adverse effects at the doses tested. Local delivery of 7ND at the implant site may provide a therapeutic strategy to diminish particle-associated periprosthetic inflammation and osteolysis. © 2013 Wiley Periodicals, Inc.

HIV-1 Vpr modulates macrophage metabolic pathways: a SILAC-based quantitative analysis.

PubMed

Barrero, Carlos A; Datta, Prasun K; Sen, Satarupa; Deshmane, Satish; Amini, Shohreh; Khalili, Kamel; Merali, Salim

2013-01-01

Human immunodeficiency virus type 1 encoded viral protein Vpr is essential for infection of macrophages by HIV-1. Furthermore, these macrophages are resistant to cell death and are viral reservoir. However, the impact of Vpr on the macrophage proteome is yet to be comprehended. The goal of the present study was to use a stable-isotope labeling by amino acids in cell culture (SILAC) coupled with mass spectrometry-based proteomics approach to characterize the Vpr response in macrophages. Cultured human monocytic cells, U937, were differentiated into macrophages and transduced with adenovirus construct harboring the Vpr gene. More than 600 proteins were quantified in SILAC coupled with LC-MS/MS approach, among which 136 were significantly altered upon Vpr overexpression in macrophages. Quantified proteins were selected and clustered by biological functions, pathway and network analysis using Ingenuity computational pathway analysis. The proteomic data illustrating increase in abundance of enzymes in the glycolytic pathway (pentose phosphate and pyruvate metabolism) was further validated by western blot analysis. In addition, the proteomic data demonstrate down regulation of some key mitochondrial enzymes such as glutamate dehydrogenase 2 (GLUD2), adenylate kinase 2 (AK2) and transketolase (TKT). Based on these observations we postulate that HIV-1 hijacks the macrophage glucose metabolism pathway via the Vpr-hypoxia inducible factor 1 alpha (HIF-1 alpha) axis to induce expression of hexokinase (HK), glucose-6-phosphate dehyrogenase (G6PD) and pyruvate kinase muscle type 2 (PKM2) that facilitates viral replication and biogenesis, and long-term survival of macrophages. Furthermore, dysregulation of mitochondrial glutamate metabolism in macrophages can contribute to neurodegeneration via neuroexcitotoxic mechanisms in the context of NeuroAIDS.

Primary mouse lung fibroblasts help macrophages to tackle Mycobacterium tuberculosis more efficiently and differentiate into myofibroblasts up on bacterial stimulation.

PubMed

Verma, Subash Chand; Agarwal, Pooja; Krishnan, Manju Y

2016-03-01

Keeping with their classical role in wound healing, fibroblasts of the lung take part in the resolution of tubercular granulomas. They are totally absent in nascent granulomas, but surround necrotizing granulomas, and are the majority of cells in healed granulomas. Lung fibroblasts may become infected with Mycobacterium tuberculosis (Mtb). Two previous studies suggested an immunomodulatory effect of fibroblasts on infected macrophages. In the present study, we looked at the role of primary mouse lung fibroblasts on naive or activated mouse bone marrow macrophages infected with Mtb and the effect of infection on fibroblast properties. We observed that with fibroblasts in the vicinity, infected naive macrophages restricted the bacterial growth, while activated macrophages turned more bactericidal with concomitant increase in nitrite production. Neutralizing IL-1α in fibroblast supernatant reduced the nitrite production by infected macrophages. Secretion of IL-6 and MCP-1 was down-regulated, while TNF-α was up-regulated in infected naive macrophages. In infected activated macrophages, the secretion of IL-6 was up-regulated, while that of MCP-1 and TNF-α was unaffected. The 'fibroblast effects' were enhanced when the fibroblasts too were infected. Mtb induced IL-1 secretion and pro-fibrotic responses by fibroblasts. Mtb-induced myofibroblast conversion was blocked by rapamycin suggesting cell signalling via mTOR. Copyright © 2015 Elsevier Ltd. All rights reserved.

Neutrophil-derived alpha defensins control inflammation by inhibiting macrophage mRNA translation

PubMed Central

Tomlinson, Gareth H.; Miles, Katherine; Smith, Richard W. P.; Rossi, Adriano G.; Hiemstra, Pieter S.; van ’t Wout, Emily F. A.; Dean, Jonathan L. E.; Gray, Nicola K.; Lu, Wuyuan; Gray, Mohini

2016-01-01

Neutrophils are the first and most numerous cells to arrive at the site of an inflammatory insult and are among the first to die. We previously reported that alpha defensins, released from apoptotic human neutrophils, augmented the antimicrobial capacity of macrophages while also inhibiting the biosynthesis of proinflammatory cytokines. In vivo, alpha defensin administration protected mice from inflammation, induced by thioglychollate-induced peritonitis or following infection with Salmonella enterica serovar Typhimurium. We have now dissected the antiinflammatory mechanism of action of the most abundant neutrophil alpha defensin, Human Neutrophil Peptide 1 (HNP1). Herein we show that HNP1 enters macrophages and inhibits protein translation without inducing the unfolded-protein response or affecting mRNA stability. In a cell-free in vitro translation system, HNP1 powerfully inhibited both cap-dependent and cap-independent mRNA translation while maintaining mRNA polysomal association. This is, to our knowledge, the first demonstration of a peptide released from one cell type (neutrophils) directly regulating mRNA translation in another (macrophages). By preventing protein translation, HNP1 functions as a “molecular brake” on macrophage-driven inflammation, ensuring both pathogen clearance and the resolution of inflammation with minimal bystander tissue damage. PMID:27044108

Characterization of the receptors for mycobacterial cord factor in Guinea pig.

PubMed

Toyonaga, Kenji; Miyake, Yasunobu; Yamasaki, Sho

2014-01-01

Guinea pig is a widely used animal for research and development of tuberculosis vaccines, since its pathological disease process is similar to that present in humans. We have previously reported that two C-type lectin receptors, Mincle (macrophage inducible C-type lectin, also called Clec4e) and MCL (macrophage C-type lectin, also called Clec4d), recognize the mycobacterial cord factor, trehalose-6,6'-dimycolate (TDM). Here, we characterized the function of the guinea pig homologue of Mincle (gpMincle) and MCL (gpMCL). gpMincle directly bound to TDM and transduced an activating signal through ITAM-bearing adaptor molecule, FcRγ. Whereas, gpMCL lacked C-terminus and failed to bind to TDM. mRNA expression of gpMincle was detected in the spleen, lymph nodes and peritoneal macrophages and it was strongly up-regulated upon stimulation of zymosan and TDM. The surface expression of gpMincle was detected on activated macrophages by a newly established monoclonal antibody that also possesses a blocking activity. This antibody potently suppressed TNF production in BCG-infected macrophages. Collectively, gpMincle is the TDM receptor in the guinea pig and TDM-Mincle axis is involved in host immune responses against mycobacteria.

Effects of co-administration of fluoxetine and risperidone on properties of peritoneal and pleural macrophages in rats subjected to the forced swimming test.

PubMed

Roman, Adam; Kuśmierczyk, Justyna; Klimek, Ewa; Rogóż, Zofia; Nalepa, Irena

2012-01-01

Literature data show that administration of atypical antipsychotic drug, risperidone (RIS), enhances antidepressive action of fluoxetine (FLU). As antidepressive treatments also regulate immune functions, we examined whether combined administration of FLU and RIS to rats subsequently subjected to a forced swimming test (FST) modifies parameters of macrophage activity that are directly related to their immunomodulatory functions, i.e., arginase (ARG) activity and nitric oxide (NO) synthesis. Antidepressive action of the drugs was assessed with FST. Peritoneal and pleural cells were eluted and selected parameters of immunoreactivity were assessed colorimetrically. We found that the concomitant administration of FLU (10 mg/kg) and RIS (0.1 mg/kg) produced antidepressive-like effects in the FST,whereas the drugs were ineffective if administered separately. Stress related to the FST affected immune cell redistribution and changed some of the metabolic and immunomodulatory properties of macrophages. FLU administered to rats at a suboptimal dose for antidepressive action potently influenced macrophage immunomodulatory properties and redirected their activity toward anti-inflammatory M2 functional phenotype, as manifested by changes in the ARG/NO ratio. These effects resulted from a direct cellular influence of the drug, as well as its action via neuroendocrine pathways, as evidenced in peritoneal and pleural cells. Addition of RIS did not augment immunomodulatory action of FLU, though the combination showed antidepressant-like activity in the FST. Our results suggest that when the drugs were administered together, FLU was potent enough to redirect macrophages toward M2 activity. It is also postulated that drug-induced changes in the immune system are not so closely related to antidepressant-like effects or might be secondary to those produced in the neuroendocrine system.

A systems-based approach to analyse the host response in murine lung macrophages challenged with respiratory syncytial virus

PubMed Central

2013-01-01

Background Respiratory syncytial virus (RSV) is an important cause of lower respiratory tract infection in young children. The degree of disease severity is determined by the host response to infection. Lung macrophages play an important early role in the host response to infection and we have used a systems-based approach to examine the host response in RSV-infected lung-derived macrophage cells. Results Lung macrophage cells could be efficiently infected (>95%) with RSV in vitro, and the expression of several virus structural proteins could be detected. Although we failed to detect significant levels of virus particle production, virus antigen could be detected up until 96 hours post-infection (hpi). Microarray analysis indicated that 20,086 annotated genes were expressed in the macrophage cells, and RSV infection induced an 8.9% and 11.3% change in the global gene transcriptome at 4 hpi and 24 hpi respectively. Genes showing up-regulated expression were more numerous and exhibited higher changes in expression compared to genes showing down-regulated expression. Based on gene ontology, genes with cytokine, antiviral, cell death, and signal transduction functions showed the highest increases in expression, while signalling transduction, RNA binding and protein kinase genes showed the greatest reduction in expression levels. Analysis of the global gene expression profile using pathway enrichment analysis confirmed that up-regulated expression of pathways related to pathogen recognition, interferon signalling and antigen presentation occurred in the lung macrophage cells challenged with RSV. Conclusion Our data provided a comprehensive analysis of RSV-induced gene expression changes in lung macrophages. Although virus gene expression was detected, our data was consistent with an abortive infection and this correlated with the activation of several antivirus signalling pathways such as interferon type I signalling and cell death signalling. RSV infection induced a relatively large increase in pro-inflammatory cytokine expression, however the maintenance of this pro-inflammatory response was not dependent on the production of infectious virus particles. The sustained pro-inflammatory response even in the absence of a productive infection suggests that drugs that control the pro-inflammatory response may be useful in the treatment of patients with severe RSV infection. PMID:23506210

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

NASA Astrophysics Data System (ADS)

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

2010-07-01

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

Antiviral Regulation in Porcine Monocytic Cells at Different Activation States

PubMed Central

Rowland, Raymond R. R.

2014-01-01

ABSTRACT Monocytic cells, including macrophages and dendritic cells, exist in different activation states that are critical to the regulation of antimicrobial immunity. Many pandemic viruses are monocytotropic, including porcine reproductive and respiratory syndrome virus (PRRSV), which directly infects subsets of monocytic cells and interferes with antiviral responses. To study antiviral responses in PRRSV-infected monocytic cells, we characterized inflammatory cytokine responses and genome-wide profiled signature genes to investigate response pathways in uninfected and PRRSV-infected monocytic cells at different activation states. Our findings showed suppressed interferon (IFN) production in macrophages in non-antiviral states and an arrest of lipid metabolic pathways in macrophages at antiviral states. Importantly, porcine monocytic cells at different activation states were susceptible to PRRSV and responded differently to viral infection. Based on Gene Ontology (GO) analysis, two approaches were used to potentiate antiviral activity: (i) pharmaceutical modulation of cellular lipid metabolism and (ii) in situ PRRSV replication-competent expression of interferon alpha (IFN-α). Both approaches significantly suppressed exogenous viral infection in monocytic cells. In particular, the engineered IFN-expressing PRRSV strain eliminated exogenous virus infection and sustained cell viability at 4 days postinfection in macrophages. These findings suggest an intricate interaction of viral infection with the activation status of porcine monocytic cells. An understanding and integration of antiviral infection with activation status of monocytic cells may provide a means of potentiating antiviral immunity. IMPORTANCE Activation statuses of monocytic cells, including monocytes, macrophages (Mϕs), and dendritic cells (DCs), are critically important for antiviral immunity. Unfortunately, the activation status of porcine monocytic cells or how cell activation status functionally interacts with antiviral immunity remains largely unknown. This is a significant omission because many economically important porcine viruses are monocytotropic, including our focus, PRRSV, which alone causes nearly $800 million economic loss annually in the U.S. swine industries. PRRSV is ideal for deciphering how monocytic cell activation statuses interact with antiviral immunity, because it directly infects subsets of monocytic cells and subverts overall immune responses. In this study, we systematically investigate the activation status of porcine monocytic cells to determine the intricate interaction of viral infection with activation statuses and functionally regulate antiviral immunity within the framework of the activation paradigm. Our findings may provide a means of potentiating antiviral immunity and leading to novel vaccines for PRRS prevention. PMID:25056886

Three-dimensional organotypic co-culture model of intestinal epithelial cells and macrophages to study Salmonella enterica colonization patterns.

PubMed

Barrila, Jennifer; Yang, Jiseon; Crabbé, Aurélie; Sarker, Shameema F; Liu, Yulong; Ott, C Mark; Nelman-Gonzalez, Mayra A; Clemett, Simon J; Nydam, Seth D; Forsyth, Rebecca J; Davis, Richard R; Crucian, Brian E; Quiriarte, Heather; Roland, Kenneth L; Brenneman, Karen; Sams, Clarence; Loscher, Christine; Nickerson, Cheryl A

2017-01-01

Three-dimensional models of human intestinal epithelium mimic the differentiated form and function of parental tissues often not exhibited by two-dimensional monolayers and respond to Salmonella in key ways that reflect in vivo infections. To further enhance the physiological relevance of three-dimensional models to more closely approximate in vivo intestinal microenvironments encountered by Salmonella , we developed and validated a novel three-dimensional co-culture infection model of colonic epithelial cells and macrophages using the NASA Rotating Wall Vessel bioreactor. First, U937 cells were activated upon collagen-coated scaffolds. HT-29 epithelial cells were then added and the three-dimensional model was cultured in the bioreactor until optimal differentiation was reached, as assessed by immunohistochemical profiling and bead uptake assays. The new co-culture model exhibited in vivo-like structural and phenotypic characteristics, including three-dimensional architecture, apical-basolateral polarity, well-formed tight/adherens junctions, mucin, multiple epithelial cell types, and functional macrophages. Phagocytic activity of macrophages was confirmed by uptake of inert, bacteria-sized beads. Contribution of macrophages to infection was assessed by colonization studies of Salmonella pathovars with different host adaptations and disease phenotypes (Typhimurium ST19 strain SL1344 and ST313 strain D23580; Typhi Ty2). In addition, Salmonella were cultured aerobically or microaerobically, recapitulating environments encountered prior to and during intestinal infection, respectively. All Salmonella strains exhibited decreased colonization in co-culture (HT-29-U937) relative to epithelial (HT-29) models, indicating antimicrobial function of macrophages. Interestingly, D23580 exhibited enhanced replication/survival in both models following invasion. Pathovar-specific differences in colonization and intracellular co-localization patterns were observed. These findings emphasize the power of incorporating a series of related three-dimensional models within a study to identify microenvironmental factors important for regulating infection.

FurA contributes to the oxidative stress response regulation of Mycobacterium avium ssp. paratuberculosis

PubMed Central

Eckelt, Elke; Meißner, Thorsten; Meens, Jochen; Laarmann, Kristin; Nerlich, Andreas; Jarek, Michael; Weiss, Siegfried; Gerlach, Gerald-F.; Goethe, Ralph

2015-01-01

The ferric uptake regulator A (FurA) is known to be involved in iron homeostasis and stress response in many bacteria. In mycobacteria the precise role of FurA is still unclear. In the presented study, we addressed the functional role of FurA in the ruminant pathogen Mycobacterium avium ssp. paratuberculosis (MAP) by construction of a furA deletion strain (MAPΔfurA). RNA deep sequencing revealed that the FurA regulon consists of repressed and activated genes associated to stress response or intracellular survival. Not a single gene related to metal homeostasis was affected by furA deletion. A decisive role of FurA during intracellular survival in macrophages was shown by significantly enhanced survival of MAPΔfurA compared to the wildtype, indicating that a principal task of mycobacterial FurA is oxidative stress response regulation in macrophages. This resistance was not associated with altered survival of mice after long term infection with MAP. Our results demonstrate for the first time, that mycobacterial FurA is not involved in the regulation of iron homeostasis. However, they provide strong evidence that FurA contributes to intracellular survival as an oxidative stress sensing regulator. PMID:25705205

FurA contributes to the oxidative stress response regulation of Mycobacterium avium ssp. paratuberculosis.

PubMed

Eckelt, Elke; Meißner, Thorsten; Meens, Jochen; Laarmann, Kristin; Nerlich, Andreas; Jarek, Michael; Weiss, Siegfried; Gerlach, Gerald-F; Goethe, Ralph

2015-01-01

The ferric uptake regulator A (FurA) is known to be involved in iron homeostasis and stress response in many bacteria. In mycobacteria the precise role of FurA is still unclear. In the presented study, we addressed the functional role of FurA in the ruminant pathogen Mycobacterium avium ssp. paratuberculosis (MAP) by construction of a furA deletion strain (MAPΔfurA). RNA deep sequencing revealed that the FurA regulon consists of repressed and activated genes associated to stress response or intracellular survival. Not a single gene related to metal homeostasis was affected by furA deletion. A decisive role of FurA during intracellular survival in macrophages was shown by significantly enhanced survival of MAPΔfurA compared to the wildtype, indicating that a principal task of mycobacterial FurA is oxidative stress response regulation in macrophages. This resistance was not associated with altered survival of mice after long term infection with MAP. Our results demonstrate for the first time, that mycobacterial FurA is not involved in the regulation of iron homeostasis. However, they provide strong evidence that FurA contributes to intracellular survival as an oxidative stress sensing regulator.

Chicken macrophages infected with Salmonella (S.) Enteritidis or S. heidelberg produce differential responses in immune and metabolic signaling pathways

USDA-ARS?s Scientific Manuscript database

Protein kinases act in coordination with phosphatases to control protein phosphorylation and regulate signaling pathways and cellular processes involved in nearly every functions of cell life. Salmonella are known to manipulate the host kinase network to gain entrance and survive inside host cells....

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

PubMed

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.

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

PubMed

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

2017-08-19

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

Loss of autophagy enhances MIF/macrophage migration inhibitory factor release by macrophages.

PubMed

Lee, Jacinta P W; Foote, Andrew; Fan, Huapeng; Peral de Castro, Celia; Lang, Tali; Jones, Sarah A; Gavrilescu, Nichita; Mills, Kingston H G; Leech, Michelle; Morand, Eric F; Harris, James

2016-06-02

MIF (macrophage migration inhibitory factor [glycosylation-inhibiting factor]) is a pro-inflammatory cytokine expressed in multiple cells types, including macrophages. MIF plays a pathogenic role in a number of inflammatory diseases and has been linked to tumor progression in some cancers. Previous work has demonstrated that loss of autophagy in macrophages enhances secretion of IL1 family cytokines. Here, we demonstrate that loss of autophagy, by pharmacological inhibition or siRNA silencing of Atg5, enhances MIF secretion by monocytes and macrophages. We further demonstrate that this is dependent on mitochondrial reactive oxygen species (ROS). Induction of autophagy with MTOR inhibitors had no effect on MIF secretion, but amino acid starvation increased secretion. This was unaffected by Atg5 siRNA but was again dependent on mitochondrial ROS. Our data demonstrate that autophagic regulation of mitochondrial ROS plays a pivotal role in the regulation of inflammatory cytokine secretion in macrophages, with potential implications for the pathogenesis of inflammatory diseases and cancers.

Estrogen receptor-alpha promotes alternative macrophage activation during cutaneous repair.

PubMed

Campbell, Laura; Emmerson, Elaine; Williams, Helen; Saville, Charis R; Krust, Andrée; Chambon, Pierre; Mace, Kimberly A; Hardman, Matthew J

2014-09-01

Efficient local monocyte/macrophage recruitment is critical for tissue repair. Recruited macrophages are polarized toward classical (proinflammatory) or alternative (prohealing) activation in response to cytokines, with tight temporal regulation crucial for efficient wound repair. Estrogen acts as a potent anti-inflammatory regulator of cutaneous healing. However, an understanding of estrogen/estrogen receptor (ER) contribution to macrophage polarization and subsequent local effects on wound healing is lacking. Here we identify, to our knowledge previously unreported, a role whereby estrogen receptor α (ERα) signaling preferentially polarizes macrophages from a range of sources to an alternative phenotype. Cell-specific ER ablation studies confirm an in vivo role for inflammatory cell ERα, but not ERβ, in poor healing associated with an altered cytokine profile and fewer alternatively activated macrophages. Furthermore, we reveal intrinsic changes in ERα-deficient macrophages, which are unable to respond to alternative activation signals in vitro. Collectively, our data reveal that inflammatory cell-expressed ERα promotes alternative macrophage polarization, which is beneficial for timely healing. Given the diverse physiological roles of ERs, these findings will likely be of relevance to many pathologies involving excessive inflammation.

Response of macrophages in rat skeletal muscle after eccentric exercise.

PubMed

Zuo, Qun; Wang, Shu-Chen; Yu, Xin-Kai; Chao, Wei-Wei

2018-04-01

Macrophages are known to be important for healing numerous injured tissues depending on their functional phenotypes in response to different stimuli. The objective of this study was to reveal macrophage phenotypic changes involved in exercise-induced skeletal muscle injury and regeneration. Adult male Sprague-Dawley rats experienced one session of downhill running (16° decline, 16 m/min) for 90 min. After exercise the blood and soleus muscles were collected at 0 h, 6 h, 12 h, 1 d, 2 d, 3 d, 1 w and 2 w after exercise, separately. It was showed that CD68 + M1 macrophages mainly infiltrated into muscle necrotic sites at 1-3 d, while CD163 + M2 macrophages were present in muscles from 0 h to 2 weeks after exercise. Using transmission electron microscopy, we observed activated satellite cells 1 d after exercise. Th1-associated transcripts of iNOS and Ccl2 were inhibited post exercise, while COX-2 mRNA was dramatically increased 12 h after running (p < 0.01). M2 phenotype marker Arg-1 increased 12 h and 3 d (p < 0.05, p < 0.01) after exercise, and Clec10a and Mrc2 were up-regulated in muscles 12 h following exercise (p < 0.05, p < 0.05). The data demonstrate the dynamic patterns of macrophage phenotype in skeletal muscle upon eccentric exercise stimuli, and M1 and M2 phenotypes perform different functions during exercise-induced skeletal muscle injury and recovery. Copyright © 2018 Daping Hospital and the Research Institute of Surgery of the Third Military Medical University. Production and hosting by Elsevier B.V. All rights reserved.

S1PR3 Signaling Drives Bacterial Killing and Is Required for Survival in Bacterial Sepsis.

PubMed

Hou, JinChao; Chen, QiXing; Wu, XiaoLiang; Zhao, DongYan; Reuveni, Hadas; Licht, Tamar; Xu, MengLong; Hu, Hu; Hoeft, Andreas; Ben-Sasson, Shmuel A; Shu, Qiang; Fang, XiangMing

2017-12-15

Efficient elimination of pathogenic bacteria is a critical determinant in the outcome of sepsis. Sphingosine-1-phosphate receptor 3 (S1PR3) mediates multiple aspects of the inflammatory response during sepsis, but whether S1PR3 signaling is necessary for eliminating the invading pathogens remains unknown. To investigate the role of S1PR3 in antibacterial immunity during sepsis. Loss- and gain-of-function experiments were performed using cell and murine models. S1PR3 levels were determined in patients with sepsis and healthy volunteers. S1PR3 protein levels were up-regulated in macrophages upon bacterial stimulation. S1pr3 -/- mice showed increased mortality and increased bacterial burden in multiple models of sepsis. The transfer of wild-type bone marrow-derived macrophages rescued S1pr3 -/- mice from lethal sepsis. S1PR3-overexpressing macrophages further ameliorated the mortality rate of sepsis. Loss of S1PR3 led to markedly decreased bacterial killing in macrophages. Enhancing endogenous S1PR3 activity using a peptide agonist potentiated the macrophage bactericidal function and improved survival rates in multiple models of sepsis. Mechanically, the reactive oxygen species levels were decreased and phagosome maturation was delayed in S1pr3 -/- macrophages due to impaired recruitment of vacuolar protein-sorting 34 to the phagosomes. In addition, S1RP3 expression levels were elevated in monocytes from patients with sepsis. Higher levels of monocytic S1PR3 were associated with efficient intracellular bactericidal activity, better immune status, and preferable outcomes. S1PR3 signaling drives bacterial killing and is essential for survival in bacterial sepsis. Interventions targeting S1PR3 signaling could have translational implications for manipulating the innate immune response to combat pathogens.

Combined effect of BCG vaccination and enriched environment promote neurogenesis and spatial cognition via a shift in meningeal macrophage M2 polarization.

PubMed

Qi, Fangfang; Zuo, Zejie; Yang, Junhua; Hu, Saisai; Yang, Yang; Yuan, Qunfang; Zou, Juntao; Guo, Kaihua; Yao, Zhibin

2017-02-10

The spatial learning abilities of developing mice benefit from extrinsic cues, such as an enriched environment, with concomitant enhancement in cognitive functions. Interestingly, such enhancements can be further increased through intrinsic Bacillus Calmette-Guérin (BCG) vaccination. Here, we first report that combined neonatal BCG vaccination and exposure to an enriched environment (Enr) induced combined neurobeneficial effects, including hippocampal long-term potentiation, and increased neurogenesis and spatial learning and memory, in mice exposed to the Enr and vaccinated with BCG relative to those in the Enr that did not receive BCG vaccination. Neonatal BCG vaccination markedly induced anti-inflammatory meningeal macrophage polarization both in regular and Enr breeding mice. The meninges are composed of the pia mater, dura mater, and choroid plexus. Alternatively, this anti-inflammatory activity of the meninges occurred simultaneously with increased expression of the neurotrophic factors BDNF/IGF-1 and the M2 microglial phenotype in the hippocampus. Our results reveal a critical role for BCG vaccination in the regulation of neurogenesis and spatial cognition through meningeal macrophage M2 polarization and neurotrophic factor expression; these effects were completely or partially prevented by minocycline or anti-IL-10 antibody treatment, respectively. Together, we first claim that immunological factor and environmental factor induce a combined effect on neurogenesis and cognition via a common pathway-meningeal macrophage M2 polarization. We also present a novel functional association between peripheral T lymphocytes and meningeal macrophages after evoking adaptive immune responses in the periphery whereby T lymphocytes are recruited to the meninges in response to systemic IFN-γ signaling. This leads to meningeal macrophage M2 polarization, subsequent to microglial M2 activation and neurotrophic factor expression, and eventually promotes a positive behavior.

The effect of RU 486 and related compounds on cultured macrophage differentiation and function.

PubMed

Roberts, C P; Murphy, A A; Santanam, N; Parthasarathy, S

1996-08-01

Our purpose was to examine RU 486 and related compounds on macrophage scavenger receptors and cellular adhesion. THP-1 cells were activated with phorbol myristate acetate and treated with dexamethasone, levonorgestrel, and RU 486 alone or in combination. Scavenger receptor activity was determined by counting adhered cells. In addition, fluorescently labeled acetyl low density lipoprotein uptake was determined. Both dexamethasone and RU 486 significantly decreased activated macrophages (81% and 26% of control). Levonorgestrel stimulated adherent cells in activated monocytes (130% of control). RU 486 and dexamethasone were antagonistic when combined (p < 0.001). In contrast, dexamethasone could not overcome the stimulatory effect of levonorgestrel (p < 0.001). Fluorescent studies yielded similar results. RU 486 is a known antiglucocorticoid with novel antioxidant properties. Levonorgestrel has antiglucocorticoid but no antioxidant activity. Glucocorticoids decrease scavenger receptors and antioxidants regulate inflammatory cytokines. RU 486 antagonized the inhibitory effect of dexamethasone on scavenger receptors, whereas levonorgestrel was stimulatory. It is therapeutically important to up-regulate scavenger receptor activity by antiglucocorticoids in the peritoneal cavity of women with endometriosis. However, because these mechanisms also induce inflammatory cytokines, a balance of antioxidants and antiglucocorticoids may prove beneficial.

Microvasculature remodeling in the mouse lower gut during inflammaging

PubMed Central

Jeong, Jae-Ho; Kim, KwangSoo; Lim, Daejin; Kim, Kun-Hee; Kim, Hyung-Seok; Lee, Sungsu; Song, Joo-Hye; Moon, Byoung-Gon; Choy, Hyon E.; Park, Sang Chul

2017-01-01

Inflammaging is defined as low-grade, chronic, systemic inflammation in aging, in the absence of overt infection. Age-associated deterioration of gastrointestinal function could be ascribed to the inflammaging, although evidence is yet to emerge. Here we show that microvessels in aging mouse intestine were progressively deprived of supportive structures, microvessel-associated pericytes and adherens junction protein vascular endothelial (VE)-cadherin, and became leaky. This alteration was ascribed to up-regulation of angiopoetin-2 in microvascular endothelial cells. Up-regulation of the angiopoietin-2 was by TNF-α, originated from M2-like residential CD206+ macrophages, proportion of which increases as animal ages. It was concluded that antigenic burdens encountered in intestine throughout life create the condition of chronic stage of inflammation, which accumulates M2-like macrophages expressing TNF-α. The TNF-α induces vascular leakage to facilitate recruitment of immune cells into intestine under the chronic inflammatory setting. PMID:28045067

Mouse CD23 regulates monocyte activation through an interaction with the adhesion molecule CD11b/CD18.

PubMed

Lecoanet-Henchoz, S; Plater-Zyberk, C; Graber, P; Gretener, D; Aubry, J P; Conrad, D H; Bonnefoy, J Y

1997-09-01

CD23 is expressed on a variety of hemopoietic cells. Recently, we have reported that blocking CD23 interactions in a murine model of arthritis resulted in a marked improvement of disease severity. Here, we demonstrate that CD11b, the alpha chain of the beta 2 integrin adhesion molecule complex CD11b/CD18 expressed on monocytes interacts with CD23. Using a recombinant fusion protein (ZZ-CD23), murine CD23 was shown to bind to peritoneal macrophages and peripheral blood cells isolated from mice as well as the murine macrophage cell line, RAW. The interactions between mouse ZZ-CD23 and CD11b/CD18-expressing cells were significantly inhibited by anti-CD11b monoclonal antibodies. A functional consequence was then demonstrated by inducing an up-regulation of interleukin-6 (IL-6) production following ZZ-CD23 incubation with monocytes. The addition of Fab fragments generated from the monoclonal antibody CD11b impaired this cytokine production by 50%. Interestingly, a positive autocrine loop was identified as IL-6 was shown to increase CD23 binding to macrophages. These results demonstrate that similar to findings using human cells, murine CD23 binds to the surface adhesion molecule, CD11b, and these interactions regulate biological activities of murine myeloid cells.

Suppression of PLCβ2 by Endotoxin Plays a Role in the Adenosine A2A Receptor-Mediated Switch of Macrophages from an Inflammatory to an Angiogenic Phenotype

PubMed Central

Grinberg, Stan; Hasko, Gyorgy; Wu, Dianqing; Leibovich, Samuel Joseph

2009-01-01

Toll-like receptor (TLR) 2, 4, 7, and 9 agonists, together with adenosine A2A receptor (A2AR) agonists, switch macrophages from an inflammatory (M1) to an angiogenic (M2-like) phenotype. This switch involves induction of A2ARs by TLR agonists, down-regulation of tumor necrosis factor α (TNFα) and interleukin-12, and up-regulation of vascular endothelial growth factor (VEGF) and interleukin-10 expression. We show here that the TLR4 agonist lipopolysaccharide (LPS) induces rapid and specific post-transcriptional down-regulation of phospholipase C(PLC)β1 and β2 expression in macrophages by de-stabilizing their mRNAs. The PLCβ inhibitor U73122 down-regulates TNFα expression by macrophages, and in the presence of A2AR agonists, up-regulates VEGF, mimicking the synergistic action of LPS with A2AR agonists. Selective down-regulation of PLCβ2, but not PLCβ1, using small-interfering RNA resulted in increased VEGF expression in response to A2AR agonists, but did not suppress TNFα expression. Macrophages from PLCβ2−/− mice also expressed increased VEGF in response to A2AR agonists. LPS-mediated suppression of PLCβ1 and β2 is MyD88-dependent. In a model of endotoxic shock, LPS (35 μg/mouse, i.p.) suppressed PLCβ1 and β2 expression in spleen, liver, and lung of wild-type but not MyD88−/− mice. These studies indicate that LPS suppresses PLCβ1 and β2 expression in macrophages in vitro and in several tissues in vivo. These results suggest that suppression of PLCβ2 plays an important role in switching M1 macrophages into an M2-like state. PMID:19850892

Innate defense regulator IDR-1018 activates human mast cells through G protein-, phospholipase C-, MAPK- and NF-ĸB-sensitive pathways.

PubMed

Yanashima, Kensuke; Chieosilapatham, Panjit; Yoshimoto, Eri; Okumura, Ko; Ogawa, Hideoki; Niyonsaba, François

2017-08-01

Host defense (antimicrobial) peptides not only display antimicrobial activities against numerous pathogens but also exert a broader spectrum of immune-modulating functions. Innate defense regulators (IDRs) are a class of host defense peptides synthetically developed from natural or endogenous cationic host defense peptides. Of the IDRs developed to date, IDR-1018 is more efficient not only in killing bacteria but also in regulating the various functions of macrophages and neutrophils and accelerating the wound healing process. Because mast cells intimately participate in wound healing and a number of host defense peptides involved in wound healing are also known to activate mast cells, this study aimed to investigate the effects of IDR-1018 on mast cell activation. Here, we showed that IDR-1018 induced the degranulation of LAD2 human mast cells and caused their production of leukotrienes, prostaglandins and various cytokines and chemokines, including granulocyte-macrophage colony-stimulating factor, interleukin-8, monocyte chemoattractant protein-1 and -3, macrophage-inflammatory protein-1α and -1β, and tumor necrosis factor-α. Furthermore, IDR-1018 increased intracellular calcium mobilization and induced mast cell chemotaxis. The mast cell activation was markedly suppressed by pertussis toxin, U-73122, U0126, SB203580, JNK inhibitor II, and NF-κB activation inhibitor II, suggesting the involvement of G-protein, phospholipase C, ERK, p38, JNK and NF-κB pathways, respectively, in IDR-1018-induced mast cell activation. Notably, we confirmed that IDR-1018 caused the phosphorylation of MAPKs and IκB. Altogether, the current study suggests a novel immunomodulatory role of IDR-1018 through its ability to recruit and activate human mast cells at the sites of inflammation and wounds. We report that IDR-1018 stimulates various functions of human mast cells. IDR-1018-induced mast cell activation is mediated through G protein, PLC, MAPK and NF-κB pathways. IDR-1018 will be a useful therapeutic agent for wound healing.

Hypericin-mediated sonodynamic therapy induces autophagy and decreases lipids in THP-1 macrophage by promoting ROS-dependent nuclear translocation of TFEB.

PubMed

Li, Xuesong; Zhang, Xin; Zheng, Longbin; Kou, Jiayuan; Zhong, Zhaoyu; Jiang, Yueqing; Wang, Wei; Dong, Zengxiang; Liu, Zhongni; Han, Xiaobo; Li, Jing; Tian, Ye; Zhao, Yajun; Yang, Liming

2016-12-22

Lipid catabolism disorder is the primary cause of atherosclerosis. Transcription factor EB (TFEB) prevents atherosclerosis by activating macrophage autophagy to promote lipid degradation. Hypericin-mediated sonodynamic therapy (HY-SDT) has been proved non-invasively inducing THP-1-derived macrophage apoptosis; however, it is unknown whether macrophage autophagy could be triggered by HY-SDT to influence cellular lipid catabolism via regulating TFEB. Here, we report that HY-SDT resulted in the time-dependent THP-1-derived macrophage autophagy activation through AMPK/AKT/mTOR pathway. Besides, TFEB nuclear translocation in macrophage was triggered by HY-SDT to promote autophagy activation and lysosome regeneration which enhanced lipid degradation in response to atherogenic lipid stressors. Moreover, following HY-SDT, the ABCA1 expression level was increased to promote lipid efflux in macrophage, and the expression levels of CD36 and SR-A were decreased to inhibit lipid uptake, both of which were prevented by TFEB knockdown. These results indicated that TFEB nuclear translocation activated by HY-SDT was not only the key regulator of autophagy activation and lysosome regeneration in macrophage to promote lipolysis, but also had a crucial role in reverse cholesterol transporters to decrease lipid uptake and increase lipid efflux. Reactive oxygen species (ROS) were adequately generated in macrophage by HY-SDT. Further, ROS scavenger N-acetyl-l-cysteine abolished HY-SDT-induced TFEB nuclear translocation and autophagy activation, implying that ROS were the primary upstream factors responsible for these effects during HY-SDT. In summary, our data indicate that HY-SDT decreases lipid content in macrophage by promoting ROS-dependent nuclear translocation of TFEB to influence consequent autophagy activation and cholesterol transporters. Thus, HY-SDT may be beneficial for atherosclerosis via TFEB regulation to ameliorate lipid overload in atherosclerotic plaques.

Resveratrol Prevents Tumor Growth and Metastasis by Inhibiting Lymphangiogenesis and M2 Macrophage Activation and Differentiation in Tumor-associated Macrophages.

PubMed

Kimura, Yoshiyuki; Sumiyoshi, Maho

2016-01-01

Antitumor and antimetastatic effects of resveratrol on tumor-induced lymphangiogenesis through the regulation of M2 macrophages in tumor-associated macrophages currently remain unknown. Therefore, we herein examined the effects of resveratrol on M2 macrophage activation and differentiation, and those of resveratrol-treated condition medium (CM) in M2 macrophages on vascular endothelial cell growth factor (VEGF)-C-induced migration, invasion, and tube formation by human lymphatic endothelial cells (HLECs). Resveratrol (50 μM or 5-50 μM) inhibited the production of interleukin-10 and monocyte chemoattractant protein-1 in M2 macrophages, whereas it promoted that of transforming growth factor-β1. Resveratrol (25 and 50 μM) inhibited the phosphorylation of signal transducer and activator of transcript 3 without affecting its expression in the differentiation process of M2 macrophages. Furthermore, resveratrol-treated CM of M2 macrophages inhibited VEGF-C-induced HLEC migration, invasion, and lymphangiogenesis. Resveratrol (25 mg/kg, twice daily) inhibited tumor growth and metastasis to the lung and also reduced the area of lymphatic endothelial cells in tumors (in vivo). These results suggest that the antitumor and antimetastatic effects of resveratrol were partly due to antilymphangiogenesis through the regulation of M2 macrophage activation and differentiation.

The liver in regulation of iron homeostasis.

PubMed

Rishi, Gautam; Subramaniam, V Nathan

2017-09-01

The liver is one of the largest and most functionally diverse organs in the human body. In addition to roles in detoxification of xenobiotics, digestion, synthesis of important plasma proteins, gluconeogenesis, lipid metabolism, and storage, the liver also plays a significant role in iron homeostasis. Apart from being the storage site for excess body iron, it also plays a vital role in regulating the amount of iron released into the blood by enterocytes and macrophages. Since iron is essential for many important physiological and molecular processes, it increases the importance of liver in the proper functioning of the body's metabolism. This hepatic iron-regulatory function can be attributed to the expression of many liver-specific or liver-enriched proteins, all of which play an important role in the regulation of iron homeostasis. This review focuses on these proteins and their known roles in the regulation of body iron metabolism. Copyright © 2017 the American Physiological Society.

Metabolic and Epigenetic Coordination of T Cell and Macrophage Immunity.

PubMed

Phan, Anthony T; Goldrath, Ananda W; Glass, Christopher K

2017-05-16

Recognition of pathogens by innate and adaptive immune cells instructs rapid alterations of cellular processes to promote effective resolution of infection. To accommodate increased bioenergetic and biosynthetic demands, metabolic pathways are harnessed to maximize proliferation and effector molecule production. In parallel, activation initiates context-specific gene-expression programs that drive effector functions and cell fates that correlate with changes in epigenetic landscapes. Many chromatin- and DNA-modifying enzymes make use of substrates and cofactors that are intermediates of metabolic pathways, providing potential cross talk between metabolism and epigenetic regulation of gene expression. In this review, we discuss recent studies of T cells and macrophages supporting a role for metabolic activity in integrating environmental signals with activation-induced gene-expression programs through modulation of the epigenome and speculate as to how this may influence context-specific macrophage and T cell responses to infection. Copyright © 2017 Elsevier Inc. All rights reserved.

Metabolic and epigenetic coordination of T cell and Macrophage immunity

PubMed Central

Phan, Anthony T.; Goldrath, Ananda W.; Glass, Christopher K.

2017-01-01

Recognition of pathogens by innate and adaptive immune cells instructs rapid alterations of cellular processes to promote effective resolution of infection. To accommodate increased bioenergetic and biosynthetic demands, metabolic pathways are harnessed to maximize proliferation and effector molecule production. In parallel, activation initiates context-specific gene-expression programs that drive effector functions and cell fates that correlate with changes in epigenetic landscapes. Many chromatin- and DNA-modifying enzymes make use of substrates and cofactors that are intermediates of metabolic pathways, providing potential cross talk between metabolism and epigenetic regulation of gene expression. In this review, we discuss recent studies of T cells and macrophages supporting a role for metabolic activity in integrating environmental signals with activation-induced gene-expression programs through modulation of the epigenome and speculate as to how this may influence context-specific macrophage and T cell responses to infection. PMID:28514673

MafB antagonizes phenotypic alteration induced by GM-CSF in microglia

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

Koshida, Ryusuke, E-mail: rkoshida-myz@umin.ac.jp; Oishi, Hisashi, E-mail: hoishi@md.tsukuba.ac.jp; Hamada, Michito

2015-07-17

Microglia are tissue-resident macrophages which are distributed throughout the central nervous system (CNS). Recent studies suggest that microglia are a unique myeloid population distinct from peripheral macrophages in terms of origin and gene expression signature. Granulocyte-macrophage colony-stimulating factor (GM-CSF), a pleiotropic cytokine regulating myeloid development, has been shown to stimulate proliferation and alter phenotype of microglia in vitro. However, how its signaling is modulated in microglia is poorly characterized. MafB, a bZip transcriptional factor, is highly expressed in monocyte-macrophage lineage cells including microglia, although its role in microglia is largely unknown. We investigated the crosstalk between GM-CSF signaling and MafB bymore » analyzing primary microglia. We found that Mafb-deficient microglia grew more rapidly than wild-type microglia in response to GM-CSF. Moreover, the expression of genes associated with microglial differentiation was more downregulated in Mafb-deficient microglia cultured with GM-CSF. Notably, such differences between the genotypes were not observed in the presence of M-CSF. In addition, we found that Mafb-deficient microglia cultured with GM-CSF barely extended their membrane protrusions, probably due to abnormal activation of RhoA, a key regulator of cytoskeletal remodeling. Altogether, our study reveals that MafB is a negative regulator of GM-CSF signaling in microglia. These findings could provide new insight into the modulation of cytokine signaling by transcription factors in microglia. - Highlights: • GM-CSF alters the phenotype of microglia in vitro more potently than M-CSF. • Transcription factor MafB antagonizes the effect of GM-CSF on microglia in vitro. • MafB deficiency leads to RhoA activation in microglia in response to GM-CSF. • We show for the first time the function of MafB in microglia.« less

Nitro-oleic acid inhibits vascular endothelial inflammatory responses and the endothelial-mesenchymal transition.

PubMed

Ambrozova, Gabriela; Fidlerova, Tana; Verescakova, Hana; Koudelka, Adolf; Rudolph, Tanja K; Woodcock, Steven R; Freeman, Bruce A; Kubala, Lukas; Pekarova, Michaela

2016-11-01

Inflammatory-mediated pathological processes in the endothelium arise as a consequence of the dysregulation of vascular homeostasis. Of particular importance are mediators produced by stimulated monocytes/macrophages inducing activation of endothelial cells (ECs). This is manifested by excessive soluble pro-inflammatory mediator production and cell surface adhesion molecule expression. Nitro-fatty acids are endogenous products of metabolic and inflammatory reactions that display immuno-regulatory potential and may represent a novel therapeutic strategy to treat inflammatory diseases. The purpose of our study was to characterize the effects of nitro-oleic acid (OA-NO2) on inflammatory responses and the endothelial-mesenchymal transition (EndMT) in ECs that is a consequence of the altered healing phase of the immune response. The effect of OA-NO2 on inflammatory responses and EndMT was determined in murine macrophages and murine and human ECs using Western blotting, ELISA, immunostaining, and functional assays. OA-NO2 limited the activation of macrophages and ECs by reducing pro-inflammatory cytokine production and adhesion molecule expression through its modulation of STAT, MAPK and NF-κB-regulated signaling. OA-NO2 also decreased transforming growth factor-β-stimulated EndMT and pro-fibrotic phenotype of ECs. These effects are related to the downregulation of Smad2/3. The study shows the pleiotropic effect of OA-NO2 on regulating EC-macrophage interactions during the immune response and suggests a role for OA-NO2 in the regulation of vascular endothelial immune and fibrotic responses arising during chronic inflammation. These findings propose the OA-NO2 may be useful as a novel therapeutic agent for treatment of cardiovascular disorders associated with dysregulation of the endothelial immune response. Copyright © 2016 Elsevier B.V. All rights reserved.

Macrophage Pro-Inflammatory Response to Francisella novicida Infection Is Regulated by SHIP

PubMed Central

Parsa, Kishore V. L; Ganesan, Latha P; Rajaram, Murugesan V. S; Gavrilin, Mikhail A; Balagopal, Ashwin; Mohapatra, Nrusingh P; Wewers, Mark D; Schlesinger, Larry S; Gunn, John S; Tridandapani, Susheela

2006-01-01

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 NFκB. Consistently, macrophages lacking SHIP displayed enhanced NFκB-driven gene transcription, whereas overexpression of SHIP led to decreased NFκB activation. Thus, we propose that SHIP negatively regulates F. novicida–induced inflammatory cytokine response by antagonizing the PI3K/Akt pathway and suppressing NFκB-mediated gene transcription. A detailed analysis of phosphoinositide signaling may provide valuable clues for better understanding the pathogenesis of tularemia. PMID:16848641

The Inositol Phosphatase SHIP Controls Salmonella enterica Serovar Typhimurium Infection In Vivo▿

PubMed Central

Bishop, Jennifer L.; Sly, Laura M.; Krystal, Gerald; Finlay, B. Brett

2008-01-01

The SH2 domain-containing inositol 5′-phosphatase, SHIP, negatively regulates various hematopoietic cell functions and is critical for maintaining immune homeostasis. However, whether SHIP plays a role in controlling bacterial infections in vivo remains unknown. Salmonella enterica causes human salmonellosis, a disease that ranges in severity from mild gastroenteritis to severe systemic illness, resulting in significant morbidity and mortality worldwide. The susceptibility of ship+/+and ship−/− mice and bone marrow-derived macrophages to S. enterica serovar Typhimurium infection was compared. ship−/− mice displayed an increased susceptibility to both oral and intraperitoneal serovar Typhimurium infection and had significantly higher bacterial loads in intestinal and systemic sites than ship+/+mice, indicating a role for SHIP in the gut-associated and systemic pathogenesis of serovar Typhimurium in vivo. Cytokine analysis of serum from orally infected mice showed that ship−/− mice produce lower levels of Th1 cytokines than do ship+/+ animals at 2 days postinfection, and in vitro analysis of supernatants taken from infected bone marrow-derived macrophages derived to mimic the in vivo ship−/− alternatively activated (M2) macrophage phenotype correlated with these data. M2 macrophages were the predominant population in vivo in both oral and intraperitoneal infections, since tissue macrophages within the small intestine and peritoneal macrophages from ship−/− mice showed elevated levels of the M2 macrophage markers Ym1 and Arginase 1 compared to ship+/+ cells. Based on these data, we propose that M2 macrophage skewing in ship−/− mice contributes to ineffective clearance of Salmonella in vivo. PMID:18426884

Adenosine A2A Receptor Activation and Macrophage-mediated Experimental Glomerulonephritis

PubMed Central

Garcia, Gabriela E.; Truong, Luan D.; Li, Ping; Zhang, Ping; Du, Jie; Chen, Jiang-Fan; Feng, Lili

2010-01-01

In immune-induced inflammation, leukocytes are key mediators of tissue damage. Since A2A adenosine receptors (A2AR) are endogenous suppressors of inflammation, we examined cellular and molecular mechanisms of kidney damage to determine whether selective activation of A2AR will suppress inflammation in a rat model of glomerulonephritis. Activation of A2AR reduced the degree of kidney injury in both the acute inflammatory phase and the progressive phase of glomerulonephritis. This protection against acute and chronic inflammation was associated with suppression of the glomerular expression of the MDC/CCL22 chemokine and down-regulation of MIP-1α/CCL3, RANTES/CCL5, MIP-1β/CCL4, and MCP-1/CCL2 chemokines. The expression of anti-inflammatory cytokines, IL-4 and IL-10, also increased. The mechanism for these anti-inflammatory responses to the A2AR agonist was suppression of macrophages function. A2AR expression was increased in macrophages, macrophage-derived chemokines were reduced in response to the A2AR agonist, and chemokines not expressed in macrophages did not respond to A2AR activation. Thus, activation of the A2AR on macrophages inhibits immune-associated inflammation. In glomerulonephritis, A2AR activation modulates inflammation and tissue damage even in the progressive phase of glomerulonephritis. Accordingly, pharmacological activation of A2AR could be developed into a novel treatment for glomerulonephritis and other macrophage-related inflammatory diseases. PMID:17898087

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

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

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

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

Paeonia japonica, Houttuynia cordata, and Aster scaber water extracts induce nitric oxide and cytokine production by lipopolysaccharide-activated macrophages.

PubMed

Kim, Jin; Park, Chang-Shin; Lim, Yunsook; Kim, Hyun-Sook

2009-04-01

Natural products are increasingly recognized as potential targets for drug discovery and development. We previously reported that Paeonia japonica, Houttuynia cordata, and Aster scaber enhanced macrophage activation both in vitro and in vivo. In the present study we investigated the immunomodulating effects of these plants on lipopolysacharide (LPS)-stimulated macrophages. An aqueous extract of each plant was administered to female BALB/c mice every other day for 4 weeks. Peritoneal macrophages were then collected and incubated to examine the immunoreactivity of macrophages against LPS at different time points. The expression levels of inducible nitric oxide (NO) synthetase (iNOS), cyclooxygenase (COX)-2, and inhibitory factor kappaB alpha (IkappaBalpha) proteins and the production of NO metabolite (nitrite), prostaglandin (PG) E(2), and the pro-inflammatory cytokines interleukin-1beta, interleukin-6, and tumor necrosis factor-alpha were determined in the activated macrophages treated with extracts from each plant individually or combined. High levels of pro-inflammatory cytokines were produced by A. scaber-, P. japonica-, and H. cordata-treated macrophages following 24 hours of LPS stimulation. P. japonica, H. cordata, and A. scaber treatment also induced the production of nitrate by LPS-treated macrophages. Induction of iNOS mRNA and protein was also different in each group. PGE(2) secretion was up-regulated by all extract-treated macrophages at early time points; however, no significant differences were observed between the groups by 8 hours post-LPS stimulation. Treatment with A. scaber extract resulted in the highest levels of IkappaBalpha degradation. Our findings illustrate that the natural plant products P. japonica, H. cordata, and A. scaber may enhance immune function by modulating ex vivo pro-inflammatory cytokine and NO production as well as the expression of iNOS and COX-2.

Vitronectin and fibronectin function as glucan binding proteins augmenting macrophage responses to Pneumocystis carinii.

PubMed

Vassallo, R; Kottom, T J; Standing, J E; Limper, A H

2001-08-01

beta-glucans represent major structural components of fungal cell walls. We recently reported that Pneumocystis carinii beta-glucans stimulate alveolar macrophages to release proinflammatory cytokines. Macrophage activation by beta-glucan is augmented by serum, implying the presence of circulating factors that interact with beta-glucans and enhance their ability to stimulate macrophages. Using beta-glucan-enriched cell wall fractions from P. carinii and Saccharomyces cerevisiae, two prominent proteins were precipitated from serum and demonstrated to be vitronectin (VN) and fibronectin (FN) by immune analysis. Preincubation of beta-glucan with VN or FN enhanced macrophage activation in response to this cell wall component. Because VN and FN accumulate in the lungs during P. carinii pneumonia, we further investigated hepatic and pulmonary expression of VN and FN messenger RNA during infection. P. carinii pneumonia in rodents is associated with increased hepatic expression of VN and FN as well as increased local expression of FN in the lung. Because interleukin (IL)-6 represents the major regulator of VN and FN expression during inflammatory conditions, we measured macrophage IL-6 release in response to stimulation with P. carinii beta-glucan. Stimulation of macrophages with P. carinii beta-glucan induced significant release of IL-6. Elevated concentrations of IL-6 were noted in the blood of infected animals compared with uninfected control animals. These studies indicate that VN and FN bind to beta-glucan components of P. carinii and augment macrophage inflammatory responses. P. carinii cell wall beta-glucan stimulates secretion of IL-6 by macrophages, thereby enhancing hepatic synthesis of both VN and FN, and lung synthesis of FN during pneumonia.

Type conversion of secretomes in a 3D TAM2 and HCC cell co-culture system and functional importance of CXCL2 in HCC

PubMed Central

Lu, Yu; Li, Shan; Ma, Liping; Li, Yan; Zhang, Xiaolian; Peng, Qiliu; Mo, Cuiju; Huang, Li; Qin, Xue; Liu, Yinkun

2016-01-01

Macrophages play important roles in the tumor microenvironment, driving cancer progression and metastasis, particularly in hepatocellular carcinoma (HCC). However, few studies have assessed the exact secretome composition in HCC. In the present study, the impact of different phenotype of macrophages on HCC cells was investigated. Alternatively activated macrophages (M2) were found to significantly increase the proliferation, migration, and invasion abilities of SMMC7721 cells (all P < 0.05). M2 were then co-cultured with SMMC7721 cells to reconstruct the tumor microenvironment. Conditioned medium from 3D single cultures of M2, SMMC7721 cells, and their co-culture system were analyzed using quantitative proteomics via iTRAQ labeling combined with mass spectrometric analysis. Secretome analysis revealed a total of 159 differential secreted proteins in the co-culture system compared to the single culture systems, with 63 being up-regulated (>1.3-fold) and 96 down-regulated (<0.7-fold). CXCL2 was confirmed to have higher expression in the co-culture system and HCC tissues, and was selected for further investigation. Functional effects data suggested that recombinant human CXCL2 significantly enhanced the migration, invasion ability of SMMC7721 cells, and weakened adhesion ability. While CXCL2 neutralization and CXCR2 blockage significantly inhibited the effects of CXCL2 on SMMC7721 cells, indicating that CXCL2 may play pivotal role in HCC metastasis. PMID:27117207

Type conversion of secretomes in a 3D TAM2 and HCC cell co-culture system and functional importance of CXCL2 in HCC.

PubMed

Lu, Yu; Li, Shan; Ma, Liping; Li, Yan; Zhang, Xiaolian; Peng, Qiliu; Mo, Cuiju; Huang, Li; Qin, Xue; Liu, Yinkun

2016-04-27

Macrophages play important roles in the tumor microenvironment, driving cancer progression and metastasis, particularly in hepatocellular carcinoma (HCC). However, few studies have assessed the exact secretome composition in HCC. In the present study, the impact of different phenotype of macrophages on HCC cells was investigated. Alternatively activated macrophages (M2) were found to significantly increase the proliferation, migration, and invasion abilities of SMMC7721 cells (all P < 0.05). M2 were then co-cultured with SMMC7721 cells to reconstruct the tumor microenvironment. Conditioned medium from 3D single cultures of M2, SMMC7721 cells, and their co-culture system were analyzed using quantitative proteomics via iTRAQ labeling combined with mass spectrometric analysis. Secretome analysis revealed a total of 159 differential secreted proteins in the co-culture system compared to the single culture systems, with 63 being up-regulated (>1.3-fold) and 96 down-regulated (<0.7-fold). CXCL2 was confirmed to have higher expression in the co-culture system and HCC tissues, and was selected for further investigation. Functional effects data suggested that recombinant human CXCL2 significantly enhanced the migration, invasion ability of SMMC7721 cells, and weakened adhesion ability. While CXCL2 neutralization and CXCR2 blockage significantly inhibited the effects of CXCL2 on SMMC7721 cells, indicating that CXCL2 may play pivotal role in HCC metastasis.

Mimicking the tumor microenvironment to regulate macrophage phenotype and assessing chemotherapeutic efficacy in embedded cancer cell/macrophage spheroid models.

PubMed

Tevis, Kristie M; Cecchi, Ryan J; Colson, Yolonda L; Grinstaff, Mark W

2017-03-01

Tumor associated macrophages (TAMs) are critical stromal components intimately involved with the progression, invasion, and metastasis of cancer cells. To address the need for an in vitro system that mimics the clinical observations of TAM localizations and subsequent functional performance, a cancer cell/macrophage spheroid model is described. The central component of the model is a triple negative breast cancer spheroid embedded in a three-dimensional collagen gel. Macrophages are incorporated in two different ways. The first is a heterospheroid, a spheroid containing both tumor cells and macrophages. The heterospheroid mimics the population of TAMs infiltrated into the tumor mass, thus being exposed to hypoxia and metabolic gradients. In the second model, macrophages are diffusely seeded in the collagen surrounding the spheroid, thus modeling TAMs in the cancer stroma. The inclusion of macrophages as a heterospheroid changes the metabolic profile, indicative of synergistic growth. In contrast, macrophages diffusely seeded in the collagen bear the same profile regardless of the presence of a tumor cell spheroid. The macrophages in the heterospheroid secrete EGF, a cytokine critical to tumor/macrophage co-migration, and an EGF inhibitor decreases the metabolic activity of the heterospheroid, which is not observed in the other systems. The increased secretion of IL-10 indicates that the heterospheroid macrophages follow an M2/TAM differentiation pathway. Lastly, the heterospheroid exhibits resistance to paclitaxel. In summary, the collagen embedded heterospheroid model promotes TAM-like characteristics, and will be of utility in cancer biology and drug discovery. Two in vitro collagen-embedded multicellular spheroid models are described that mimic the clinical observations of macrophage localization within a tumor. Incorporation of macrophages within a breast cancer spheroid emphasizes cell-cell interactions with subsequent differentiation toward a tumor-promoting TAM phenotype. In contrast, macrophages seeded around the tumor spheroid display decreased interaction with cancer cells and no indication of a TAM phenotype. Finally, the presence of macrophages in the heterospheroid increases resistance to paclitaxel. This study demonstrates that cell-cell interactions and 3D collagen matrix direct macrophage activity, and, thus, highlights the important role the local environment itself plays in macrophage behavior. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The contribution of inflammasome components on macrophage response to surface nanotopography and chemistry

NASA Astrophysics Data System (ADS)

Christo, Susan; Bachhuka, Akash; Diener, Kerrilyn R.; Vasilev, Krasimir; Hayball, John D.

2016-05-01

Implantable devices have become an established part of medical practice. However, often a negative inflammatory host response can impede the integration and functionality of the device. In this paper, we interrogate the role of surface nanotopography and chemistry on the potential molecular role of the inflammasome in controlling macrophage responses. To achieve this goal we engineered model substrata having precisely controlled nanotopography of predetermined height and tailored outermost surface chemistry. Bone marrow derived macrophages (BMDM) were harvested from genetically engineered mice deficient in the inflammasome components ASC, NLRP3 and AIM2. These cells were then cultured on these nanoengineered substrata and assessed for their capacity to attach and express pro-inflammatory cytokines. Our data provide evidence that the inflammasome components ASC, NLRP3 and AIM2 play a role in regulating macrophage adhesion and activation in response to surface nanotopography and chemistry. The findings of this paper are important for understanding the inflammatory consequences caused by biomaterials and pave the way to the rational design of future implantable devices having controlled and predictable inflammatory outcomes.

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

PubMed Central

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

2015-01-01

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

Foxp3-positive macrophages display immunosuppressive properties and promote tumor growth

PubMed Central

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

2011-01-01

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

Loss of the co-repressor GPS2 sensitizes macrophage activation upon metabolic stress induced by obesity and type 2 diabetes.

PubMed

Fan, Rongrong; Toubal, Amine; Goñi, Saioa; Drareni, Karima; Huang, Zhiqiang; Alzaid, Fawaz; Ballaire, Raphaelle; Ancel, Patricia; Liang, Ning; Damdimopoulos, Anastasios; Hainault, Isabelle; Soprani, Antoine; Aron-Wisnewsky, Judith; Foufelle, Fabienne; Lawrence, Toby; Gautier, Jean-Francois; Venteclef, Nicolas; Treuter, Eckardt

2016-07-01

Humans with obesity differ in their susceptibility to developing insulin resistance and type 2 diabetes (T2D). This variation may relate to the extent of adipose tissue (AT) inflammation that develops as their obesity progresses. The state of macrophage activation has a central role in determining the degree of AT inflammation and thus its dysfunction, and these states are driven by epigenomic alterations linked to gene expression. The underlying mechanisms that regulate these alterations, however, are poorly defined. Here we demonstrate that a co-repressor complex containing G protein pathway suppressor 2 (GPS2) crucially controls the macrophage epigenome during activation by metabolic stress. The study of AT from humans with and without obesity revealed correlations between reduced GPS2 expression in macrophages, elevated systemic and AT inflammation, and diabetic status. The causality of this relationship was confirmed by using macrophage-specific Gps2-knockout (KO) mice, in which inappropriate co-repressor complex function caused enhancer activation, pro-inflammatory gene expression and hypersensitivity toward metabolic-stress signals. By contrast, transplantation of GPS2-overexpressing bone marrow into two mouse models of obesity (ob/ob and diet-induced obesity) reduced inflammation and improved insulin sensitivity. Thus, our data reveal a potentially reversible disease mechanism that links co-repressor-dependent epigenomic alterations in macrophages to AT inflammation and the development of T2D.

Macrophage-Mediated Glial Cell Elimination in the Postnatal Mouse Cochlea

PubMed Central

Brown, LaShardai N.; Xing, Yazhi; Noble, Kenyaria V.; Barth, Jeremy L.; Panganiban, Clarisse H.; Smythe, Nancy M.; Bridges, Mary C.; Zhu, Juhong; Lang, Hainan

2017-01-01

Hearing relies on the transmission of auditory information from sensory hair cells (HCs) to the brain through the auditory nerve. This relay of information requires HCs to be innervated by spiral ganglion neurons (SGNs) in an exclusive manner and SGNs to be ensheathed by myelinating and non-myelinating glial cells. In the developing auditory nerve, mistargeted SGN axons are retracted or pruned and excessive cells are cleared in a process referred to as nerve refinement. Whether auditory glial cells are eliminated during auditory nerve refinement is unknown. Using early postnatal mice of either sex, we show that glial cell numbers decrease after the first postnatal week, corresponding temporally with nerve refinement in the developing auditory nerve. Additionally, expression of immune-related genes was upregulated and macrophage numbers increase in a manner coinciding with the reduction of glial cell numbers. Transient depletion of macrophages during early auditory nerve development, using transgenic CD11bDTR/EGFP mice, resulted in the appearance of excessive glial cells. Macrophage depletion caused abnormalities in myelin formation and transient edema of the stria vascularis. Macrophage-depleted mice also showed auditory function impairment that partially recovered in adulthood. These findings demonstrate that macrophages contribute to the regulation of glial cell number during postnatal development of the cochlea and that glial cells play a critical role in hearing onset and auditory nerve maturation. PMID:29375297

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

Ye, Dan, E-mail: y.dan@lacdr.leidenuniv.nl; Meurs, Illiana; Ohigashi, Megumi

Objectives: To determine the role of macrophage ATP-binding cassette transporter A5 (ABCA5) in cellular cholesterol homeostasis and atherosclerotic lesion development. Methods and results: Chimeras with dysfunctional macrophage ABCA5 (ABCA5{sup -M/-M}) were generated by transplantation of bone marrow from ABCA5 knockout (ABCA5{sup -/-}) mice into irradiated LDLr{sup -/-} mice. In vitro, bone marrow-derived macrophages from ABCA5{sup -M/-M} chimeras exhibited a 29% (P < 0.001) decrease in cholesterol efflux to HDL, whereas a 21% (P = 0.07) increase in cholesterol efflux to apoA-I was observed. Interestingly, expression of ABCA1, but not ABCG1, was up-regulated in absence of functional ABCA5 in macrophages. Tomore » induce atherosclerosis, the transplanted LDLr{sup -/-} mice were fed a high-cholesterol Western-type diet (WTD) for 6, 10, or 18 weeks, allowing analysis of effects on initial as well as advanced lesion development. Atherosclerosis development was not affected in male ABCA5{sup -M/-M} chimeras after 6, 10, and 18 weeks WTD feeding. However, female ABCA5{sup -M/-M} chimeras did develop significantly (P < 0.05) larger aortic root lesions as compared with female controls after 6 and 10 weeks WTD feeding. Conclusions: ABCA5 influences macrophage cholesterol efflux, and selective disruption of ABCA5 in macrophages leads to increased atherosclerotic lesion development in female LDLr{sup -/-} mice.« less

Hematopoietic-to-mesenchymal transition of adipose tissue macrophages is regulated by integrin β1 and fabricated fibrin matrices

PubMed Central

Majka, Susan M.; Kohrt, Wendy M.; Miller, Heidi L.; Sullivan, Timothy M.; Klemm, Dwight J.

2017-01-01

ABSTRACT Some bona fide adult adipocytes arise de novo from a bone marrow-derived myeloid lineage. These studies further demonstrate that adipose tissue stroma contains a resident population of myeloid cells capable of adipocyte and multilineage mesenchymal differentiation. These resident myeloid cells lack hematopoietic markers and express mesenchymal and progenitor cell markers. Because bone marrow mesenchymal progenitor cells have not been shown to enter the circulation, we hypothesized that myeloid cells acquire mesenchymal differentiation capacity in adipose tissue. We fabricated a 3-dimensional fibrin matrix culture system to define the adipose differentiation potential of adipose tissue-resident myeloid subpopulations, including macrophages, granulocytes and dendritic cells. Our data show that multilineage mesenchymal potential was limited to adipose tissue macrophages, characterized by the acquisition of adipocyte, osteoblast, chondrocyte and skeletal muscle myocyte phenotypes. Fibrin hydrogel matrices stimulated macrophage loss of hematopoietic cell lineage determinants and the expression of mesenchymal and progenitor cell markers, including integrin β1. Ablation of integrin β1 in macrophages inhibited adipocyte specification. Therefore, some bona fide adipocytes are specifically derived from adipose tissue-resident macrophages via an integrin β1-dependent hematopoietic-to-mesenchymal transition, whereby they become capable of multipotent mesenchymal differentiation. The requirement for integrin β1 highlights this molecule as a potential target for controlling the production of marrow-derived adipocytes and their contribution to adipose tissue development and function. PMID:28441086

CD163-L1 is an endocytic macrophage protein strongly regulated by mediators in the inflammatory response.

PubMed

Moeller, Jesper B; Nielsen, Marianne J; Reichhardt, Martin P; Schlosser, Anders; Sorensen, Grith L; Nielsen, Ole; Tornøe, Ida; Grønlund, Jørn; Nielsen, Maria E; Jørgensen, Jan S; Jensen, Ole N; Mollenhauer, Jan; Moestrup, Søren K; Holmskov, Uffe

2012-03-01

CD163-L1 belongs to the group B scavenger receptor cysteine-rich family of proteins, where the CD163-L1 gene arose by duplication of the gene encoding the hemoglobin scavenger receptor CD163 in late evolution. The current data demonstrate that CD163-L1 is highly expressed and colocalizes with CD163 on large subsets of macrophages, but in contrast to CD163 the expression is low or absent in monocytes and in alveolar macrophages, glia, and Kupffer cells. The expression of CD163-L1 increases when cultured monocytes are M-CSF stimulated to macrophages, and the expression is further increased by the acute-phase mediator IL-6 and the anti-inflammatory mediator IL-10 but is suppressed by the proinflammatory mediators IL-4, IL-13, TNF-α, and LPS/IFN-γ. Furthermore, we show that CD163-L1 is an endocytic receptor, which internalizes independently of cross-linking through a clathrin-mediated pathway. Two cytoplasmic splice variants of CD163-L1 are differentially expressed and have different subcellular distribution patterns. Despite its many similarities to CD163, CD163-L1 does not possess measurable affinity for CD163 ligands such as the haptoglobin-hemoglobin complex or various bacteria. In conclusion, CD163-L1 exhibits similarity to CD163 in terms of structure and regulated expression in cultured monocytes but shows clear differences compared with the known CD163 ligand preferences and expression pattern in the pool of tissue macrophages. We postulate that CD163-L1 functions as a scavenger receptor for one or several ligands that might have a role in resolution of inflammation.

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

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

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

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

Macrophage-derived LIF and IL1B regulate alpha(1,2)fucosyltransferase 2 (Fut2) expression in mouse uterine epithelial cells during early pregnancy.

PubMed

Jasper, Melinda J; Care, Alison S; Sullivan, Brad; Ingman, Wendy V; Aplin, John D; Robertson, Sarah A

2011-01-01

Macrophages accumulate within stromal tissue subjacent to the luminal epithelium in the mouse uterus during early pregnancy after seminal fluid exposure at coitus. To investigate their role in regulating epithelial cell expression of fucosylated structures required for embryo attachment and implantation, fucosyltransferase enzymes Fut1, Fut2 (Enzyme Commission number [EC] 2.4.1.69), and Fut4 (EC 2.4.1.214) and Muc1 and Muc4 mRNAs were quantified by quantitative real-time PCR in uterine epithelial cells after laser capture microdissection in situ or after epithelial cell coculture with macrophages or macrophage-secreted factors. When uterine macrophage recruitment was impaired by mating with seminal plasma-deficient males, epithelial cell Fut2 expression on Day 3.5 postcoitus (pc) was reduced compared to intact-mated controls. Epithelial cell Fut2 was upregulated in vitro by coculture with macrophages or macrophage-conditioned medium (MCM). Macrophage-derived cytokines LIF, IL1B, and IL12 replicated the effect of MCM on Fut2 mRNA expression, and MCM-stimulated expression was inhibited by anti-LIF and anti-IL1B neutralizing antibodies. The effects of acute macrophage depletion on fucosylated structures detected with lectins Ulex europaeus 1 (UEA-1) and Lotus tetragonolobus purpureas (LTP), or LewisX immunoreactivity, were quantified in vivo in Cd11b-dtr transgenic mice. Depletion of macrophages caused a 30% reduction in luminal epithelial UEA-1 staining and a 67% reduction in LewisX staining in uterine tissues of mice hormonally treated to mimic early pregnancy. Together, these data demonstrate that uterine epithelial Fut2 mRNA expression and terminal fucosylation of embryo attachment ligands is regulated in preparation for implantation by factors including LIF and IL1B secreted from macrophages recruited during the inflammatory response to insemination.

Dan-Lou Prescription Inhibits Foam Cell Formation Induced by ox-LDL via the TLR4/NF-κB and PPARγ Signaling Pathways.

PubMed

Gao, Li-Na; Zhou, Xin; Lu, Yu-Ren; Li, Kefeng; Gao, Shan; Yu, Chun-Quan; Cui, Yuan-Lu

2018-01-01

Atherosclerosis is the major worldwide cause of mortality for patients with coronary heart disease. Many traditional Chinese medicine compound prescriptions for atherosclerosis treatment have been tried in patients. Dan-Lou prescription, which is improved from Gualou-Xiebai-Banxia decoction, has been used to treat chest discomfort (coronary atherosclerosis) for approximately 2,000 years in China. Although the anti-inflammatory activities of Dan-Lou prescription have been proposed previously, the mechanism remains to be explored. Based on the interaction between inflammation and atherosclerosis, we further investigated the effect of Dan-Lou prescription on macrophage-derived foam cell formation and disclosed the underlying mechanisms. In the oxidative low-density lipoprotein (ox-LDL) induced foam cells model using murine macrophage RAW 264.7 cells, the ethanol extract from Dan-Lou prescription (EEDL) reduced ox-LDL uptake and lipid deposition by inhibiting the protein and mRNA expression of Toll-like receptor (TLR)4 and scavenger receptor (SR)B1. After stimulation with ox-LDL, the metabolic profile of macrophages was also changed, while the intervention of the EEDL mainly regulated the metabolism of isovalerylcarnitine, arachidonic acid, cholesterol, aspartic acid, arginine, lysine, L-glutamine and phosphatidylethanolamine (36:3), which participated in the regulation of the inflammatory response, lipid accumulation and cell apoptosis. In total, 27 inflammation-related gene targets were screened, and the biological mechanisms, pathways and biological functions of the EEDL on macrophage-derived foam cells were systemically analyzed by Ingenuity Pathway Analysis system (IPA). After verification, we found that EEDL alleviated ox-LDL induced macrophage foam cell formation by antagonizing the mRNA and protein over-expression of PPARγ, blocking the phosphorylation of IKKα/β, IκBα and NF-κB p65 and maintaining the expression balance between Bax and Bcl-2. In conclusion, we provided evidences that Dan-Lou prescription effectively attenuated macrophage foam cell formation via the TLR4/NF-κB and PPARγ signaling pathways.

Phosphorylation of SNAP-23 at Ser95 causes a structural alteration and negatively regulates Fc receptor-mediated phagosome formation and maturation in macrophages.

PubMed

Sakurai, Chiye; Itakura, Makoto; Kinoshita, Daiki; Arai, Seisuke; Hashimoto, Hitoshi; Wada, Ikuo; Hatsuzawa, Kiyotaka

2018-05-17

SNAP-23 is a plasma membrane-localized SNARE protein involved in Fc receptor (FcR)-mediated phagocytosis. However, the regulatory mechanism underlying its function remains elusive. Using phosphorylation specific-antibodies, SNAP-23 was found to be phosphorylated at Ser95 in macrophages. To understand the role of this phosphorylation, we established macrophage lines overexpressing the non-phosphorylatable S95A or the phospho-mimicking S95D mutation. The efficiency of phagosome formation and maturation was severely reduced in SNAP-23-S95D-overexpressing cells. To examine whether phosphorylation at Ser95 affected SNAP-23 structure, we constructed intramolecular Förster resonance energy transfer (FRET) probes of SNAP-23 designed to evaluate the approximation of the N-termini of the two SNARE motifs. Interestingly, a high FRET efficiency was detected on the membrane when the S95D probe was used, indicating that phosphorylation at Ser95 caused a dynamic structural shift to the closed form. Co-expression of IκB kinase (IKK) 2 enhanced the FRET efficiency of the wild-type probe on the phagosome membrane. Furthermore, the enhanced phagosomal FRET signal in interferon-γ-activated macrophages was largely dependent on IKK2, and this kinase mediated a delay in phagosome-lysosome fusion. These results suggested that SNAP-23 phosphorylation at Ser95 played an important role in the regulation of SNARE-dependent membrane fusion during FcR-mediated phagocytosis.

Periodontitis promotes the diabetic development of obese rat via miR-147 induced classical macrophage activation.

PubMed

Xu, Ran; Zeng, Guang; Wang, Shuyong; Tao, Hong; Ren, Le; Zhang, Zhe; Zhang, Qingna; Zhao, Jinxiu; Gao, Jing; Li, Daxu

2016-10-01

Emerging evidence has indicated the bad effect of periodontal inflammation on diabetes control. However, the exact regulatory mechanisms within the association between periodontitis and diabetic development remain unclear. This study aims to investigate the function of microRNAs in regulating periodontitis-induced inflammation in an obese rat model. Experimental periodontitis was introduced into OLETF and LETO rat. Intraperitoneal glucose tolerance test was performed to detect diabetic development. Serum cytokines levels and microRNAs expression were detected by ELISA and RT-PCR analysis respectively. And, macrophages were isolated for gain- and loss-of-function studies, to investigate the regulatory mechanism of miR-147 in periodontitis-induced inflammation. Periodontitis induced proinflammatory response with classical activated macrophages in both rats, but distinctively aggravated the impaired glucose tolerance of OLETF rat with spontaneous type 2 diabetes. Analysis for serum microRNAs expression showed the distinctive and synergistic upregulation of miR-147 with periodontitis-induced effects in rats, while further experiments demonstrated the positive regulatory mechanism of miR-147 on classical activated macrophages with overexpressed proinflammatory markers, showing M1 phenotype. This study provided new evidence for the positive effect of periodontal inflammation on diabetic development, while the regulatory mechanism of miR-147 on classical macrophage activation, was verified, and presumed to contribute to the impaired glucose tolerance aggravated by periodontitis in obese rats. Besides, this study indicated the application of miR-147 for therapeutic approach in the treatment of diabetes with periodontitis. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

The decidua of preeclamptic-like BPH/5 mice exhibits an exaggerated inflammatory response during early pregnancy

PubMed Central

Heyward, CY; Sones, JL; Lob, HE; Yuen, LC; Abbott, KE; Huang, W; Begun, ZR; Butler, SD; August, A; Leifer, CA; Davisson, RL

2017-01-01

Preeclampsia is a devastating complication of pregnancy characterized by late-gestation hypertension and proteinuria. Because the only definitive treatment is delivery of the fetus and placenta, preeclampsia contributes to increased morbidity and mortality of both mother and fetus. The BPH/5 mouse model, which spontaneously develops a syndrome strikingly similar to preeclampsia, displays excessive inflammation and suppression of inflammation improves pregnancy outcomes. During early pregnancy, decidual macrophages play an important role in promoting maternal tolerance to fetal antigens and regulating tissue remodeling, two functions that are critical for normal placental development. BPH/5 pregnancies are characterized by abnormal placentation; therefore, we hypothesized that macrophage localization and/or function is altered during early pregnancy at the site of placental formation (the decidua) compared to C57BL/6 controls. At early gestation time points, before the onset of maternal hypertension or proteinuria, there was a reduction in the number of macrophages in BPH/5 decidua and a concomitant increase in activated T cells compared to C57BL/6. BPH/5 decidua also exhibited decreased expression of the immunosuppressive cytokine, IL-10, and increased expression of pro-inflammatory, inducible nitric oxide synthase (iNOS). Together, these data suggest that a reduction in decidual macrophages during pregnancy is associated with immune activation in BPH/5 mice, inadequate placental development and may contribute to adverse pregnancy outcomes in this model. PMID:28432903

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

PubMed

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.

Huaier extract suppresses breast cancer via regulating tumor-associated macrophages

PubMed Central

Li, Yaming; Qi, Wenwen; Song, Xiaojin; Lv, Shangge; Zhang, Hanwen; Yang, Qifeng

2016-01-01

Macrophages in tumor microenvironment are mostly M2-polarized - and have been reported to promote tumorigenesis, which are also defined as tumor-associated macrophages (TAMs). Here, we examined the regulatory effects of Huaier extract on TAMs using RAW264.7 murine macrophage cell line. Our data demonstrated that Huaier extract could inhibit the infiltration of macrophages into tumor microenvironment in a dose-dependent manner. By performing RT-PCR, immunofluorescence and phagocytosis assay, we were able to find that Huaier extract could regulate the polarization of macrophages, with decreased M2-polarization and increased phagocytosis of RAW264.7 cells. Moreover, we identified that Huaier extract could suppress macrophages-induced angiogenesis by using HUVEC migration assay, tube formation and chorioallantoic membrane assay. Additionally, western blotting showed decreased expression of MMP2, MMP9 and VEGF with the use of Huaier extract. Finally, we found that Huaier extract could inhibit M2-macrophages infiltration and angiogenesis through treating 4T1 tumor bearing mice with Huaier extract. Our study revealed a novel mechanism of the anti-tumor effect of Huaier extract which inhibited angiogenesis by targeting TAMs. These findings provided that Huaier was a promising drug for clinical treatment of breast cancer. PMID:26831282

Genetic and environmental risk factors for atherosclerosis regulate transcription of phosphatase and actin regulating gene PHACTR1.

PubMed

Reschen, Michael E; Lin, Da; Chalisey, Anil; Soilleux, Elizabeth J; O'Callaghan, Christopher A

2016-07-01

Coronary artery disease (CAD) risk is associated with non-coding genetic variants at the phosphatase and actin regulating protein 1(PHACTR1) gene locus. The PHACTR1 gene encodes an actin-binding protein with phosphatase regulating activity. The mechanism whereby PHACTR1 influences CAD risk is unknown. We hypothesized that PHACTR1 would be expressed in human cell types relevant to CAD and regulated by atherogenic or genetic factors. Using immunohistochemistry, we demonstrate that PHACTR1 protein is expressed strongly in human atherosclerotic plaque macrophages, lipid-laden foam cells, adventitial lymphocytes and endothelial cells. Using a combination of genomic analysis and molecular techniques, we demonstrate that PHACTR1 is expressed as multiple previously uncharacterized transcripts in macrophages, foam cells, lymphocytes and endothelial cells. Immunoblotting confirmed a total absence of PHACTR1 in vascular smooth muscle cells. Real-time quantitative PCR showed that PHACTR1 is regulated by atherogenic and inflammatory stimuli. In aortic endothelial cells, oxLDL and TNF-alpha both upregulated an intermediate length transcript. A short transcript expressed only in immune cells was upregulated in macrophages by oxidized low-density lipoprotein, and oxidized phospholipids but suppressed by lipopolysaccharide or TNF-alpha. In primary human macrophages, we identified a novel expression quantitative trait locus (eQTL) specific for this short transcript, whereby the risk allele at CAD risk SNP rs9349379 is associated with reduced PHACTR1 expression, similar to the effect of an inflammatory stimulus. Our data demonstrate that PHACTR1 is a key atherosclerosis candidate gene since it is regulated by atherogenic stimuli in macrophages and endothelial cells and we identify an effect of the genetic risk variant on PHACTR1 expression in macrophages that is similar to that of an inflammatory stimulus. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Niacin and its metabolites as master regulators of macrophage activation.

PubMed

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.

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

PubMed Central

Campbell, Sharon M.; Duncan, Sheelagh; Hewitson, James P.; Barr, Tom A.; Jackson-Jones, Lucy H.; Maizels, Rick M.

2017-01-01

Rapid reprogramming of the macrophage activation phenotype is considered important in the defense against consecutive infection with diverse infectious agents. However, in the setting of persistent, chronic infection the functional importance of macrophage-intrinsic adaptation to changing environments vs. recruitment of new macrophages remains unclear. Here we show that resident peritoneal macrophages expanded by infection with the nematode Heligmosomoides polygyrus bakeri altered their activation phenotype in response to infection with Salmonella enterica ser. Typhimurium in vitro and in vivo. The nematode-expanded resident F4/80high macrophages efficiently upregulated bacterial induced effector molecules (e.g. MHC-II, NOS2) similarly to newly recruited monocyte-derived macrophages. Nonetheless, recruitment of blood monocyte-derived macrophages to Salmonella infection occurred with equal magnitude in co-infected animals and caused displacement of the nematode-expanded, tissue resident-derived macrophages from the peritoneal cavity. Global gene expression analysis revealed that although nematode-expanded resident F4/80high macrophages made an anti-bacterial response, this was muted as compared to newly recruited F4/80low macrophages. However, the F4/80high macrophages adopted unique functional characteristics that included enhanced neutrophil-stimulating chemokine production. Thus, our data provide important evidence that plastic adaptation of MΦ activation does occur in vivo, but that cellular plasticity is outweighed by functional capabilities specific to the tissue origin of the cell. PMID:28334040

Impairment of Immunoproteasome Function by Cigarette Smoke and in Chronic Obstructive Pulmonary Disease.

PubMed

Kammerl, Ilona E; Dann, Angela; Mossina, Alessandra; Brech, Dorothee; Lukas, Christina; Vosyka, Oliver; Nathan, Petra; Conlon, Thomas M; Wagner, Darcy E; Overkleeft, Hermen S; Prasse, Antje; Rosas, Ivan O; Straub, Tobias; Krauss-Etschmann, Susanne; Königshoff, Melanie; Preissler, Gerhard; Winter, Hauke; Lindner, Michael; Hatz, Rudolf; Behr, Jürgen; Heinzelmann, Katharina; Yildirim, Ali Ö; Noessner, Elfriede; Eickelberg, Oliver; Meiners, Silke

2016-06-01

Patients with chronic obstructive pulmonary disease (COPD) and in particular smokers are more susceptible to respiratory infections contributing to acute exacerbations of disease. The immunoproteasome is a specialized type of proteasome destined to improve major histocompatibility complex (MHC) class I-mediated antigen presentation for the resolution of intracellular infections. To characterize immunoproteasome function in COPD and its regulation by cigarette smoke. Immunoproteasome expression and activity were determined in bronchoalveolar lavage (BAL) and lungs of human donors and patients with COPD or idiopathic pulmonary fibrosis (IPF), as well as in cigarette smoke-exposed mice. Smoke-mediated alterations of immunoproteasome activity and MHC I surface expression were analyzed in human blood-derived macrophages. Immunoproteasome-specific MHC I antigen presentation was evaluated in spleen and lung immune cells that had been smoke-exposed in vitro or in vivo. Immunoproteasome and MHC I mRNA expression was reduced in BAL cells of patients with COPD and in isolated alveolar macrophages of patients with COPD or IPF. Exposure of immune cells to cigarette smoke extract in vitro reduced immunoproteasome activity and impaired immunoproteasome-specific MHC I antigen presentation. In vivo, acute cigarette smoke exposure dynamically regulated immunoproteasome function and MHC I antigen presentation in mouse BAL cells. End-stage COPD lungs showed markedly impaired immunoproteasome activities. We here show that the activity of the immunoproteasome is impaired by cigarette smoke resulting in reduced MHC I antigen presentation. Regulation of immunoproteasome function by cigarette smoke may thus alter adaptive immune responses and add to prolonged infections and exacerbations in COPD and IPF.

MAFB Determines Human Macrophage Anti-Inflammatory Polarization: Relevance for the Pathogenic Mechanisms Operating in Multicentric Carpotarsal Osteolysis.

PubMed

Cuevas, Víctor D; Anta, Laura; Samaniego, Rafael; Orta-Zavalza, Emmanuel; Vladimir de la Rosa, Juan; Baujat, Geneviève; Domínguez-Soto, Ángeles; Sánchez-Mateos, Paloma; Escribese, María M; Castrillo, Antonio; Cormier-Daire, Valérie; Vega, Miguel A; Corbí, Ángel L

2017-03-01

Macrophage phenotypic and functional heterogeneity derives from tissue-specific transcriptional signatures shaped by the local microenvironment. Most studies addressing the molecular basis for macrophage heterogeneity have focused on murine cells, whereas the factors controlling the functional specialization of human macrophages are less known. M-CSF drives the generation of human monocyte-derived macrophages with a potent anti-inflammatory activity upon stimulation. We now report that knockdown of MAFB impairs the acquisition of the anti-inflammatory profile of human macrophages, identify the MAFB-dependent gene signature in human macrophages and illustrate the coexpression of MAFB and MAFB-target genes in CD163 + tissue-resident and tumor-associated macrophages. The contribution of MAFB to the homeostatic/anti-inflammatory macrophage profile is further supported by the skewed polarization of monocyte-derived macrophages from multicentric carpotarsal osteolysis (Online Mendelian Inheritance in Man #166300), a pathology caused by mutations in the MAFB gene. Our results demonstrate that MAFB critically determines the acquisition of the anti-inflammatory transcriptional and functional profiles of human macrophages. Copyright © 2017 by The American Association of Immunologists, Inc.

Effect of age on marrow macrophage number and function.

PubMed

Wang, C Q; Udupa, K B; Xiao, H; Lipschitz, D A

1995-10-01

Employing flow cytometry and a monoclonal antibody against the murine macrophage antigen, Mac-1, we found a significant increase in the number of marrow macrophages in aged mice. This was reflected as significant increase with age in the number of alpha-naphthyl acetate esterase positive cells, as well as in colony forming unit-macrophage (CFU-M) progenitor cells. Macrophages from the marrow of old mice generated significantly less tumor necrosis factor alpha (TNF alpha) than did macrophages from young mice, either spontaneously or when activated by granulocyte-macrophage colony stimulating factor (GM-CSF). Furthermore, conditioned medium (CM) derived from either marrow or peritoneal macrophages of old mice caused less suppression of burst forming unit-erythroid (BFU-E) colony growth than did CM obtained from young mice. Aging, therefore, is associated with an increase in the number of marrow macrophages that have an impaired ability to generate or release cytokines. The increase in macrophage number may reflect a compensation for their reduced function. Altered macrophage number and function may contribute to the age-related decline in hematopoietic reserve capacity.

Role of Macrophages in the Repair Process during the Tissue Migrating and Resident Helminth Infections

PubMed Central

Faz-López, Berenice

2016-01-01

The Th1/Th2/Th17 balance is a fundamental feature in the regulation of the inflammatory microenvironment during helminth infections, and an imbalance in this paradigm greatly contributes to inflammatory disorders. In some cases of helminthiasis, an initial Th1 response could occur during the early phases of infection (acute), followed by a Th2 response that prevails in chronic infections. During the late phase of infection, alternatively activated macrophages (AAMs) are important to counteract the inflammation caused by the Th1/Th17 response and larval migration, limiting damage and repairing the tissue affected. Macrophages are the archetype of phagocytic cells, with the primary role of pathogen destruction and antigen presentation. Nevertheless, other subtypes of macrophages have been described with important roles in tissue repair and immune regulation. These types of macrophages challenge the classical view of macrophages activated by an inflammatory response. The role of these subtypes of macrophages during helminthiasis is a controversial topic in immunoparasitology. Here, we analyze some of the studies regarding the role of AAMs in tissue repair during the tissue migration of helminths. PMID:27648452

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

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

Stavri, Simona; Trusca, Violeta G.; Simionescu, Maya

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 thatmore » 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.« less

Down-regulation of IL-8 by high-dose vitamin D is specific to hyperinflammatory macrophages and involves mechanisms beyond up-regulation of DUSP1

PubMed Central

Dauletbaev, N; Herscovitch, K; Das, M; Chen, H; Bernier, J; Matouk, E; Bérubé, J; Rousseau, S; Lands, L C

2015-01-01

Background and Purpose There is current interest in vitamin D as a potential anti-inflammatory treatment for chronic inflammatory lung disease, including cystic fibrosis (CF). Vitamin D transcriptionally up-regulates the anti-inflammatory gene DUSP1, which partly controls production of the inflammatory chemokine IL-8. IL-8 is overabundant in CF airways, potentially due to hyperinflammatory responses of CF macrophages. We tested the ability of vitamin D metabolites to down-regulate IL-8 production in CF macrophages. Experimental Approach CF and healthy monocyte-derived macrophages (MDM) were treated with two vitamin D metabolites, 25-hydroxyvitamin D3 (25OHD3) and 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), or paricalcitol, synthetic analogue of 1,25(OH)2D3. 25OHD3 was tested at doses of 25–150 nM, whereas 1,25(OH)2D3 and paricalcitol at doses of up to 100 nM. IL-8 was stimulated by bacterial virulence factors. As potential anti-inflammatory mechanism of vitamin D metabolites, we assessed up-regulation of DUSP1. Key Results MDM from patients with CF and some healthy donors showed excessive production of stimulated IL-8, highlighting their hyperinflammatory phenotype. Vitamin D metabolites down-regulated stimulated IL-8 only in those hyperinflammatory MDM, and only when used at high doses (>100 nM for 25OHD3, or >1 nM for 1,25(OH)2D3 and paricalcitol). The magnitude of IL-8 down-regulation by vitamin D metabolites or paricalcitol was moderate (∼30% vs. >70% by low-dose dexamethasone). Transcriptional up-regulation of DUSP1 by vitamin D metabolites was seen in all tested MDM, regardless of IL-8 down-regulation. Conclusions and Implications Vitamin D metabolites and their analogues moderately down-regulate IL-8 in hyperinflammatory macrophages, including those from CF. This down-regulation appears to go through DUSP1-independent mechanisms. PMID:26178144

Down-regulation of IL-8 by high-dose vitamin D is specific to hyperinflammatory macrophages and involves mechanisms beyond up-regulation of DUSP1.

PubMed

Dauletbaev, N; Herscovitch, K; Das, M; Chen, H; Bernier, J; Matouk, E; Bérubé, J; Rousseau, S; Lands, L C

2015-10-01

There is current interest in vitamin D as a potential anti-inflammatory treatment for chronic inflammatory lung disease, including cystic fibrosis (CF). Vitamin D transcriptionally up-regulates the anti-inflammatory gene DUSP1, which partly controls production of the inflammatory chemokine IL-8. IL-8 is overabundant in CF airways, potentially due to hyperinflammatory responses of CF macrophages. We tested the ability of vitamin D metabolites to down-regulate IL-8 production in CF macrophages. CF and healthy monocyte-derived macrophages (MDM) were treated with two vitamin D metabolites, 25-hydroxyvitamin D3 (25OHD3 ) and 1,25-dihydroxyvitamin D3 (1,25(OH)2 D3 ), or paricalcitol, synthetic analogue of 1,25(OH)2 D3 . 25OHD3 was tested at doses of 25-150 nM, whereas 1,25(OH)2 D3 and paricalcitol at doses of up to 100 nM. IL-8 was stimulated by bacterial virulence factors. As potential anti-inflammatory mechanism of vitamin D metabolites, we assessed up-regulation of DUSP1. MDM from patients with CF and some healthy donors showed excessive production of stimulated IL-8, highlighting their hyperinflammatory phenotype. Vitamin D metabolites down-regulated stimulated IL-8 only in those hyperinflammatory MDM, and only when used at high doses (>100 nM for 25OHD3 , or >1 nM for 1,25(OH)2 D3 and paricalcitol). The magnitude of IL-8 down-regulation by vitamin D metabolites or paricalcitol was moderate (∼30% vs. >70% by low-dose dexamethasone). Transcriptional up-regulation of DUSP1 by vitamin D metabolites was seen in all tested MDM, regardless of IL-8 down-regulation. Vitamin D metabolites and their analogues moderately down-regulate IL-8 in hyperinflammatory macrophages, including those from CF. This down-regulation appears to go through DUSP1-independent mechanisms. © 2015 The British Pharmacological Society.

Viral hemorrhagic septicaemia virus (VHSV) up-regulates the cytotoxic activity and the perforin/granzyme pathway in the rainbow trout RTS11 cell line.

PubMed

Ordás, M C; Cuesta, A; Mercado, L; Bols, N C; Tafalla, C

2011-08-01

A survey of immune-relevant genes that might be up-regulated in response to viral hemorrhagic septicaemia virus (VHSV) in the rainbow trout monocyte-macrophage cell line, RTS11, unexpectedly revealed an increased expression of perforin (PRF) and granzyme (GRZ) genes, which represent components of the major cytotoxic pathway. The natural killer-enhancing factor (NKEF), also known to modulate cytotoxic activity, was up-regulated at the gene but strikingly down-regulated at protein level. The expression of these genes was not affected in head kidney leukocytes (HKLs) infected with VHSV, leading us to evaluate the potential cytotoxic activity of RTS11 and HKLs. For the first time, the cytotoxic activity of RTS11 against xenogeneic targets has been demonstrated, although this was modest relative to HKLs. Yet the activity in RTS11 was significantly increased by VHSV, as in HKLs. This cytotoxic activity elicited by viral infection appeared to require viral gene expression because inactivated VHSV failed to increase RTS11 cytotoxic activity. As for other immune functions, RTS11 cells provide a model for further studying cytotoxic activities of fish monocyte-macrophages. Copyright © 2011 Elsevier Ltd. All rights reserved.

Temporal Phenotypic Features Distinguish Polarized Macrophages In Vitro

PubMed Central

Melton, David W.; McManus, Linda M.; Gelfond, Jonathan A.L.; Shireman, Paula K.

2015-01-01

Macrophages are important in vascular inflammation and environmental factors influence macrophage plasticity. Macrophage transitions into pro-inflammatory (M1) or anti-inflammatory (M2) states have been defined predominately by measuring cytokines in culture media (CM). However, temporal relationships between cellular and secreted cytokines have not been established. We measured phenotypic markers and cytokines in cellular and CM of murine bone marrow-derived macrophages at multiple time points following stimulation with IFN-γ+LPS (M1), IL-4 (M2a), or IL-10 (M2c). Cytokines/proteins in M1-polarized macrophages exhibited two distinct temporal patterns; an early (0.5–3 hr), transient increase in cellular cytokines (GM-CSF, KC-GRO, MIP-2, IP-10 and MIP-1β) and a delayed (3–6 hrs) response that was more sustained [IL-3, regulated on activation normal T cell expressed and secreted (RANTES), and tissue inhibitor of metalloproteinases 1 (TIMP-1)]. M2a-related cytokine/cell markers (IGF-1, Fizz1, and Ym1) were progressively (3–24 hrs) increased post-stimulation. Additionally, novel patterns were observed. First, and unexpectedly, cellular pro-inflammatory chemokines, MCP-1 and MCP-3 but not MCP-5, were comparably increased in M1 and M2a macrophages. Second, Vegfr1 mRNA was decreased in M1 and increased in M2a macrophages. Finally, VEGF-A was increased in the CM of M1 cultures and strikingly reduced in M2a coinciding with increased Vegfr1 expression, suggesting decreased VEGF-A in M2a CM was secondary to increased soluble VEGFR1. In conclusion, macrophage cytokine production and marker expression were temporally regulated and relative levels compared across polarizing conditions were highly dependent upon the timing and location (cellular vs. CM) of the sample collection. For most cytokines, cellular production preceded increases in the CM suggesting that cellular regulatory pathways should be studied within 6 hours of stimulation. The divergent polarization-dependent expression of Vegfr1 may be essential to controlling VEGF potentially regulating angiogenesis and inflammatory cell infiltration in the vascular niche. The current study expands the repertoire of cytokines produced by polarized macrophages and provides insights into the dynamic regulation of macrophage polarization and resulting cytokines, proteins, and gene expression that influence vascular inflammation. PMID:25826285

Monocyte and macrophage-targeted NADPH oxidase mediates antifungal host defense and regulation of acute inflammation in mice

PubMed Central

Grimm, Melissa J.; Vethanayagam, R. Robert; Almyroudis, Nikolaos G.; Dennis, Carly G.; Khan, A. Nazmul H.; D’Auria, Anthony; Singel, Kelly L.; Davidson, Bruce A.; Knight, Paul R.; Blackwell, Timothy S.; Hohl, Tobias M.; Mansour, Michael K.; Vyas, Jatin M.; Röhm, Marc; Urban, Constantin F.; Kelkka, Tiina; Holmdahl, Rikard; Segal, Brahm H.

2013-01-01

Chronic granulomatous disease, an inherited disorder of the NADPH oxidase in which phagocytes are defective in the generation of superoxide anion and downstream reactive oxidant species, is characterized by severe bacterial and fungal infections and excessive inflammation. Although NADPH oxidase isoforms exist in several lineages, reactive oxidant generation is greatest in neutrophils, where NADPH oxidase has been deemed vital for pathogen killing. In contrast, the function and importance of NADPH oxidase in macrophages are less clear. Therefore, we evaluated susceptibility to pulmonary aspergillosis in globally NADPH oxidase-deficient mice versus transgenic mice with monocyte/macrophage-targeted NADPH oxidase activity. We found that the lethal inoculum was more than 100-fold greater in transgenic versus globally NADPH oxidase-deficient mice. Consistent with these in vivo results, NADPH oxidase in mouse alveolar macrophages limited germination of phagocytosed Aspergillus fumigatus spores. Finally, globally NADPH oxidase-deficient mice developed exuberant neutrophilic lung inflammation and pro-inflammatory cytokine responses to zymosan, a fungal cell wall-derived product composed principally of particulate beta-glucans, whereas inflammation in transgenic and wildtype mice was mild and transient. Together, our studies identify a central role for monocyte/macrophage NADPH oxidase in controlling fungal infection and in limiting acute lung inflammation. PMID:23509361

SARS-CoV Regulates Immune Function-Related Gene Expression in Human Monocytic Cells

PubMed Central

Hu, Wanchung; Yen, Yu-Ting; Singh, Sher; Kao, Chuan-Liang

2012-01-01

Abstract Severe acute respiratory syndrome (SARS) is characterized by acute respiratory distress syndrome (ARDS) and pulmonary fibrosis, and monocytes/macrophages are the key players in the pathogenesis of SARS. In this study, we compared the transcriptional profiles of SARS coronavirus (SARS-CoV)-infected monocytic cells against that infected by coronavirus 229E (CoV-229E). Total RNA was extracted from infected DC-SIGN-transfected monocytes (THP-1-DC-SIGN) at 6 and 24 h after infection, and the gene expression was profiled in oligonucleotide-based microarrays. Analysis of immune-related gene expression profiles showed that at 24 h after SARS-CoV infection: (1) IFN-α/β-inducible and cathepsin/proteasome genes were downregulated; (2) hypoxia/hyperoxia-related genes were upregulated; and (3) TLR/TLR-signaling, cytokine/cytokine receptor-related, chemokine/chemokine receptor-related, lysosome-related, MHC/chaperon-related, and fibrosis-related genes were differentially regulated. These results elucidate that SARS-CoV infection regulates immune-related genes in monocytes/macrophages, which may be important to the pathogenesis of SARS. PMID:22876772

SARS-CoV regulates immune function-related gene expression in human monocytic cells.

PubMed

Hu, Wanchung; Yen, Yu-Ting; Singh, Sher; Kao, Chuan-Liang; Wu-Hsieh, Betty A

2012-08-01

Severe acute respiratory syndrome (SARS) is characterized by acute respiratory distress syndrome (ARDS) and pulmonary fibrosis, and monocytes/macrophages are the key players in the pathogenesis of SARS. In this study, we compared the transcriptional profiles of SARS coronavirus (SARS-CoV)-infected monocytic cells against that infected by coronavirus 229E (CoV-229E). Total RNA was extracted from infected DC-SIGN-transfected monocytes (THP-1-DC-SIGN) at 6 and 24 h after infection, and the gene expression was profiled in oligonucleotide-based microarrays. Analysis of immune-related gene expression profiles showed that at 24 h after SARS-CoV infection: (1) IFN-α/β-inducible and cathepsin/proteasome genes were downregulated; (2) hypoxia/hyperoxia-related genes were upregulated; and (3) TLR/TLR-signaling, cytokine/cytokine receptor-related, chemokine/chemokine receptor-related, lysosome-related, MHC/chaperon-related, and fibrosis-related genes were differentially regulated. These results elucidate that SARS-CoV infection regulates immune-related genes in monocytes/macrophages, which may be important to the pathogenesis of SARS.

Dysregulated expression of miR-101b and miR-26b lead to age-associated increase in LPS-induced COX-2 expression in murine macrophage.

PubMed

Liu, Dan; Wang, Dongsheng; Xu, Zhenbiao; Gao, Jing; Liu, Min; Liu, Yanxin; Jiang, Minghong; Zheng, Dexian

2015-10-01

Aging is the natural process of decline in physiological structure and function of various molecules, cells, tissues, and organs. Growing evidence indicates that increased immune genetic diversity and dysfunction of immune system cause aging-related pathophysiological process with the growth of age. In the present study, we observed that LPS-induced higher activation of cyclooxygenase (COX)-2 promoter is associated with the upregulated binding activity of nuclear factor kappa B (NF-κB) in peritoneal macrophages of aged mice than young ones. Additionally, COX-2 is a direct target of miR-101b and miR-26b in the macrophages. Significant upregulation of miR-101b and miR-26b effectively prevented LPS-induced excessive expression of COX-2 in the young mice. Because these negative regulatory factors were unresponsive to LPS stimulation, the levels of COX-2 were markedly higher in the macrophages of aged mice. Further study showed that NF-κB activation contributed to the increase in the expression of miR-101b and miR-26b in the LPS-stimulated macrophages of young mice, but not aged ones. Moreover, histone deacetylase (HDAC) inhibitor trichostatin A (TSA) upregulated expression of miR-101b and miR-26b in the aged mouse macrophages only, but not the young cells. This demonstrated that HDAC suppressed the expression of miR-101b and miR-26b in the LPS-treated macrophages of aged mice and contributed to the aging process. TSA-induced increased expression of miR-101b and miR-26b could further suppress COX-2 expression. These findings provide novel evidence on the regulation of immune senescence and miR-101b and miR-26b, which might be promising targets in treating aged-related inflammatory diseases. Epigenetic regulation of the microRNAs (miRNAs) provides an important evidence for the treatment of innate inflammatory disease with HDAC inhibitors in elderly.

Nanoparticle delivery of miR-223 to attenuate macrophage fusion

PubMed Central

Moore, Laura Beth; Sawyer, Andrew J.; Saucier-Sawyer, Jennifer; Saltzman, W. Mark; Kyriakides, Themis R.

2016-01-01

The foreign body response (FBR) begins with injury acquired during implantation of a biomaterial (BM) and is detrimental due to the eventual encapsulation of the implant. Fusion of macrophages to form foreign body giant cells (FBGC), a hallmark of the FBR, is the consequence of a multistep mechanism induced by interleukin (IL)-4 that includes the acquisition of a fusion competent state and subsequent cytoskeletal rearrangements. However, the precise mechanism, regulation, and interplay among molecular mediators to generate FBGCs are insufficiently understood. Seeking novel mediators of fusion that might be regulated at the post-transcriptional level, we examined the role of microRNAs (miRs) in this process. A miR microarray was screened and identified miR-223 as a negative regulator of macrophage fusion. In addition, transfection of primary macrophages with a mir-223 mimic attenuated IL-4-induced fusion. Furthermore, miR-223 KO mice and mir-223 deficient cells displayed increased fusion in vivo and in vitro, respectively. Finally, we developed a method for in vivo delivery of miR-223 mimic utilizing PLGA nanoparticles, which inhibited FBGC formation in a biomaterial implant model. Our results identify miR-223 as a negative regulator of fusion and demonstrate miR-223 mimic-loaded nanoparticles as a therapeutic inhibitor of macrophage fusion. PMID:26967647

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

Baker, Anna D.; Malur, Anagha; Barna, Barbara P.

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} andmore » 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.« less

Acetylation-Dependent Regulation of Notch Signaling in Macrophages by SIRT1 Affects Sepsis Development

PubMed Central

Bai, Xiaozhi; He, Ting; Liu, Yang; Zhang, Julei; Li, Xiaoqiang; Shi, Jihong; Wang, Kejia; Han, Fu; Zhang, Wei; Zhang, Yijie; Cai, Weixia; Hu, Dahai

2018-01-01

SIRT1 is reported to participate in macrophage differentiation and affect sepsis, and Notch signaling is widely reported to influence inflammation and macrophage activation. However, the specific mechanisms through which SIRT1 regulates sepsis and the relationship between SIRT1 and Notch signaling remain poorly elucidated. In this study, we found that SIRT1 levels were decreased in sepsis both in vitro and in vivo and that SIRT1 regulation of Notch signaling affected inflammation. In lipopolysaccharide (LPS)-induced sepsis, the levels of Notch signaling molecules, including Notch1, Notch2, Hes1, and intracellular domain of Notch (NICD), were increased. However, NICD could be deacetylated by SIRT1, and this led to the suppression of Notch signaling. Notably, in macrophages from myeloid-specific RBP-J−/− mice, in which Notch signaling is inhibited, pro-inflammatory cytokines were expressed at lower levels than in macrophages from wild-type littermates and in RBP-J−/− macrophages, and the NF-κB pathway was also inhibited. Accordingly, in the case of RBP-J−/− mice, LPS-induced inflammation and mortality were lower than in wild-type mice. Our results indicate that SIRT1 inhibits Notch signaling through NICD deacetylation and thus ultimately alleviates sepsis. PMID:29867921

Regulation of CD4 Receptor and HIV-1 Entry by MicroRNAs-221 and -222 during Differentiation of THP-1 Cells.

PubMed

Lodge, Robert; Gilmore, Julian C; Ferreira Barbosa, Jérémy A; Lombard-Vadnais, Félix; Cohen, Éric A

2017-12-30

Human immunodeficiency virus type-1 (HIV-1) infection of monocyte/macrophages is modulated by the levels of entry receptors cluster of differentiation 4 (CD4) and C-C chemokine receptor type 5 (CCR5), as well as by host antiviral restriction factors, which mediate several post-entry blocks. We recently identified two microRNAs, miR-221 and miR-222, which limit HIV-1 entry during infection of monocyte-derived macrophages (MDMs) by down-regulating CD4 expression. Interestingly, CD4 is also down-regulated during the differentiation of monocytes into macrophages. In this study, we compared microRNA expression profiles in primary monocytes and macrophages by RNAseq and found that miR-221/miR-222 are enhanced in macrophages. We took advantage of the monocytic THP-1 cell line that, once differentiated, is poorly susceptible to HIV-1. Accordingly, we found that CD4 levels are very low in THP-1 differentiated cells and that this down-regulation of the virus receptor is the result of miR-221/miR-222 up-regulation during differentiation. We thus established a THP-1 cell line stably expressing a modified CD4 (THP-1-CD4 R ) that is not modulated by miR-221/miR-222. We show that in contrast to parental THP-1, this line is productively infected by HIV-1 following differentiation, sustaining efficient HIV-1 CD4-dependent replication and spread. This new THP-1-CD4 R cell line represents a useful tool for the study of HIV-1-macrophage interactions particularly in contexts where spreading of viral infection is necessary.

Regulation of CD4 Receptor and HIV-1 Entry by MicroRNAs-221 and -222 during Differentiation of THP-1 Cells

PubMed Central

Gilmore, Julian C.; Ferreira Barbosa, Jérémy A.; Lombard-Vadnais, Félix

2017-01-01

Human immunodeficiency virus type-1 (HIV-1) infection of monocyte/macrophages is modulated by the levels of entry receptors cluster of differentiation 4 (CD4) and C-C chemokine receptor type 5 (CCR5), as well as by host antiviral restriction factors, which mediate several post-entry blocks. We recently identified two microRNAs, miR-221 and miR-222, which limit HIV-1 entry during infection of monocyte-derived macrophages (MDMs) by down-regulating CD4 expression. Interestingly, CD4 is also down-regulated during the differentiation of monocytes into macrophages. In this study, we compared microRNA expression profiles in primary monocytes and macrophages by RNAseq and found that miR-221/miR-222 are enhanced in macrophages. We took advantage of the monocytic THP-1 cell line that, once differentiated, is poorly susceptible to HIV-1. Accordingly, we found that CD4 levels are very low in THP-1 differentiated cells and that this down-regulation of the virus receptor is the result of miR-221/miR-222 up-regulation during differentiation. We thus established a THP-1 cell line stably expressing a modified CD4 (THP-1-CD4R) that is not modulated by miR-221/miR-222. We show that in contrast to parental THP-1, this line is productively infected by HIV-1 following differentiation, sustaining efficient HIV-1 CD4-dependent replication and spread. This new THP-1-CD4R cell line represents a useful tool for the study of HIV-1-macrophage interactions particularly in contexts where spreading of viral infection is necessary. PMID:29301198

Methylglyoxal-bis-guanylhydrazone inhibits osteopontin expression and differentiation in cultured human monocytes.

PubMed

Jin, Xia; Xu, Hua; McGrath, Michael S

2018-01-01

Monocyte activation and polarization play essential roles in many chronic inflammatory diseases. An imbalance of M1 and M2 macrophage activation (pro-inflammatory and alternatively activated, respectively) is believed to be a key aspect in the etiology of these diseases, thus a therapeutic approach that regulates macrophage activation could be of broad clinical relevance. Methylglyoxal-bis-guanylhydrazone (MGBG), a regulator of polyamine metabolism, has recently been shown to be concentrated in monocytes and macrophages, and interfere with HIV integration into the DNA of these cells in vitro. RNA expression analysis of monocytes from HIV+ and control donors with or without MGBG treatment revealed the only gene to be consistently down regulated by MGBG to be osteopontin (OPN). The elevated expression of this pro-inflammatory cytokine and monocyte chemoattractant is associated with various chronic inflammatory diseases. We demonstrate that MGBG is a potent inhibitor of secreted OPN (sOPN) in cultured monocytes with 50% inhibition achieved at 0.1 μM of the drug. Furthermore, inhibition of OPN RNA transcription in monocyte cultures occurs at similar concentrations of the drug. During differentiation of monocytes into macrophages in vitro, monocytes express cell surface CD16 and the cells undergo limited DNA synthesis as measured by uptake of BrdU. MGBG inhibited both activities at similar doses to those regulating OPN expression. In addition, monocyte treatment with MGBG inhibited differentiation into both M1 and M2 classes of macrophages at non-toxic doses. The inhibition of differentiation and anti-OPN effects of MGBG were specific for monocytes in that differentiated macrophages were nearly resistant to MGBG activities. Thus MGBG may have potential therapeutic utility in reducing or normalizing OPN levels and regulating monocyte activation in diseases that involve chronic inflammation.

Dendritic cells and macrophages in the kidney: a spectrum of good and evil

PubMed Central

Rogers, NM; Ferenbach, DA; Isenberg, JS; Thomson, AW; Hughes, J

2015-01-01

Renal dendritic cells (DC) and macrophages (Mac) represent a constitutive, extensive and contiguous network of innate immune cells that provide sentinel and immune intelligence function. They induce and regulate inflammatory responses to freely-filtered antigenic material and protect the kidney from infection. Tissue–resident or infiltrating DC and Mac are key to the initiation and propagation of renal disease, as well as essential contributors to subsequent tissue regeneration regardless of its etiology and pathogenesis. Their identification, functional and phenotypic distinction, interplay and relationship with effector and regulatory adaptive immune cells is complex and incompletely understood. This review discusses both the common and distinct characteristics of these cells, as well as recent key advances in the field that have identified renal-specific functions of DC and Mac that enable these important, phagocytic, antigen-presenting, cells to mediate or mitigate intrinsic kidney disease. We also identify priority areas for further investigation and prospects for translational and therapeutic application of acquired knowledge. PMID:25266210

3′UTR AU-rich elements (AREs) and the RNA-binding protein Tristetraprolin (TTP) are not required for the LPS-mediated destabilization of phospholipase-Cβ-2 mRNA in murine macrophages

PubMed Central

Shukla, Smita; Elson, Genie; Blackshear, Perry J.; Lutz, Carol S.; Leibovich, S. Joseph

2017-01-01

We have shown previously that bacterial lipopolysaccharide (LPS)-mediated suppression of Phospholipase-Cβ-2 (PLCβ-2) expression is involved in M1 (inflammatory) to M2-like (wound healing) phenotypic switching of macrophages triggered by adenosine. This suppression is mediated post-transcriptionally by destabilization of PLCβ-2 mRNA. To investigate the mechanism of this LPS-mediated destabilization, we examined the roles of RNA-binding agents including microRNAs and RNA-binding proteins that are involved in regulating stability of mRNAs encoding growth factors, inflammatory mediators and proto-oncogenes. Adenylate and Uridylate (AU)-rich elements (AREs) in 3′UTRs are specific recognition sites for RNA-binding proteins including Tristetraprolin (TTP), HuR and AUF1, and for microRNAs that are involved in regulating mRNA stability. In this study, we investigated the role of TTP and AREs in regulating PLCβ-2 mRNA stability. The 3′UTR of the PLCβ-2 gene was inserted into the pLightswitch luciferase reporter plasmid and transfected into RAW264.7 cells. LPS suppressed Luciferase expression from this reporter. Luciferase expression from mutant 3′UTR constructs lacking AREs was similarly down-regulated, suggesting that these regions are not required for LPS-mediated suppression of PLCβ-2. TTP was rapidly upregulated in both primary murine macrophages and RAW264.7 cells in response to LPS. Suppression of PLCβ-2 by LPS was examined using macrophages from mice lacking TTP. LPS suppressed PLCβ-2 expression to the same extent in wild type and TTP−/− macrophages. Also, the rate of decay of PLCβ-2 mRNA in LPS-treated macrophages following transcriptional blockade was similar in wild type and TTP−/− macrophages, clearly indicating that TTP is not involved in LPS-mediated destabilization of PLCβ-2 mRNA in macrophages. PMID:28124257

Piperine metabolically regulates peritoneal resident macrophages to potentiate their functions against bacterial infection

PubMed Central

Huang, Mei-Yun; Zha, Qing-Bing; Zhao, Gao-Xiang; Hou, Xiao-Feng; Shi, Zi-Jian; Lin, Qiu-Ru; Ouyang, Dong-Yun; He, Xian-Hui

2015-01-01

Pepper, a daily-used seasoning for promoting appetite, is widely used in folk medicine for treating gastrointestinal diseases. Piperine is the major alkaloid in pepper and possesses a wide range of pharmacological activities. However, the mechanism for linking metabolic and medicinal activities of piperine remains unknown. Here we report that piperine robustly boosts mTORC1 activity by recruiting more system L1 amino acid transporter (SLC7A5/SLC3A2) to the cell membrane, thus promoting amino acid metabolism. Piperine-induced increase of mTORC1 activity in resident peritoneal macrophages (pMΦs) is correlated with enhanced production of IL-6 and TNF-α upon LPS stimulation. Such an enhancement of cytokine production could be abrogated by inhibitors of the mTOR signaling pathway, indicating mTOR's action in this process. Moreover, piperine treatment protected resident pMΦs from bacterium-induced apoptosis and disappearance, and increased their bacterial phagocytic ability. Consequently, piperine administration conferred mice resistance against bacterial infection and even sepsis. Our data highlight that piperine has the capacity to metabolically reprogram peritoneal resident macrophages to fortify their innate functions against bacterial infection. PMID:26439699

Immunomodulatory effects of Hericium erinaceus derived polysaccharides are mediated by intestinal immunology.

PubMed

Sheng, Xiaotong; Yan, Jingmin; Meng, Yue; Kang, Yuying; Han, Zhen; Tai, Guihua; Zhou, Yifa; Cheng, Hairong

2017-03-22

This study was aimed at investigating the immunomodulating activity of Hericium erinaceus polysaccharide (HEP) in mice, by assessing splenic lymphocyte proliferation (cell-mediated immunity), serum hemolysin levels (humoral immunity), phagocytic capacity of peritoneal cavity phagocytes (macrophage phagocytosis), and NK cell activity. ELISA of immunoglobulin A (SIgA) in the lamina propria, and western blotting of small intestinal proteins were also performed to gain insight into the mechanism by which HEP affects the intestinal immune system. Here, we report that HEP improves immune function by functionally enhancing cell-mediated and humoral immunity, macrophage phagocytosis, and NK cell activity. In addition, HEP was found to upregulate the secretion of SIgA and activate the MAPK and AKT cellular signaling pathways in the intestine. In conclusion, all these results allow us to postulate that the immunomodulatory effects of HEP are most likely attributed to the effective regulation of intestinal mucosal immune activity.

Comparative and network-based proteomic analysis of low dose ethanol- and lipopolysaccharide-induced macrophages.

PubMed

Kamal, Abu Hena M; Fessler, Michael B; Chowdhury, Saiful M

2018-01-01

Macrophages are specialized phagocytes that play an essential role in inflammation, immunity, and tissue repair. Profiling the global proteomic response of macrophages to microbial molecules such as bacterial lipopolysaccharide is key to understanding fundamental mechanisms of inflammatory disease. Ethanol is a widely abused substance that has complex effects on inflammation. Reports have indicated that ethanol can activate or inhibit the lipopolysaccharide receptor, Toll-like Receptor 4, in different settings, with important consequences for liver and neurologic inflammation, but the underlying mechanisms are poorly understood. To profile the sequential effect of low dose ethanol and lipopolysaccharide on macrophages, a gel-free proteomic technique was applied to RAW 264.7 macrophages. Five hundred four differentially expressed proteins were identified and quantified with high confidence using ≥ 5 peptide spectral matches. Among these, 319 proteins were shared across all treatment conditions, and 69 proteins were exclusively identified in ethanol-treated or lipopolysaccharide-stimulated cells. The interactive impact of ethanol and lipopolysaccharide on the macrophage proteome was evaluated using bioinformatics tools, enabling identification of differentially responsive proteins, protein interaction networks, disease- and function-based networks, canonical pathways, and upstream regulators. Five candidate protein coding genes (PGM2, ISYNA1, PARP1, and PSAP) were further validated by qRT-PCR that mostly related to glucose metabolism and fatty acid synthesis pathways. Taken together, this study describes for the first time at a systems level the interaction between ethanol and lipopolysaccharide in the proteomic programming of macrophages, and offers new mechanistic insights into the biology that may underlie the impact of ethanol on infectious and inflammatory disease in humans.

l-Arginine Uptake by Cationic Amino Acid Transporter Promotes Intra-Macrophage Survival of Leishmania donovani by Enhancing Arginase-Mediated Polyamine Synthesis

PubMed Central

Mandal, Abhishek; Das, Sushmita; Kumar, Ajay; Roy, Saptarshi; Verma, Sudha; Ghosh, Ayan Kumar; Singh, Ruby; Abhishek, Kumar; Saini, Savita; Sardar, Abul Hasan; Purkait, Bidyut; Kumar, Ashish; Mandal, Chitra; Das, Pradeep

2017-01-01

The survival of intracellular protozoan parasite, Leishmania donovani, the causative agent of Indian visceral leishmaniasis (VL), depends on the activation status of macrophages. l-Arginine, a semi-essential amino acid plays a crucial regulatory role for activation of macrophages. However, the role of l-arginine transport in VL still remains elusive. In this study, we demonstrated that intra-macrophage survival of L. donovani depends on the availability of extracellular l-arginine. Infection of THP-1-derived macrophage/human monocyte-derived macrophage (hMDM) with Leishmania, resulted in upregulation of l-arginine transport. While investigating the involvement of the transporters, we observed that Leishmania survival was greatly impaired when the transporters were blocked either using inhibitor or siRNA-mediated downregulation. CAT-2 was found to be the main isoform associated with l-arginine transport in L. donovani-infected macrophages. l-arginine availability and its transport regulated the host arginase in Leishmania infection. Arginase and inducible nitric oxide synthase (iNOS) expression were reciprocally regulated when assayed using specific inhibitors and siRNA-mediated downregulation. Interestingly, induction of iNOS expression and nitric oxide production were observed in case of inhibition of arginase in infected macrophages. Furthermore, inhibition of l-arginine transport as well as arginase resulted in decreased polyamine production, limiting parasite survival inside macrophages. l-arginine availability and transport regulated Th1/Th2 cytokine levels in case of Leishmania infection. Upregulation of l-arginine transport, induction of host arginase, and enhanced polyamine production were correlated with increased level of IL-10 and decreased level of IL-12 and TNF-α in L. donovani-infected macrophages. Our findings provide clear evidence for targeting the metabolism of l-arginine and l-arginine-metabolizing enzymes as an important therapeutic and prophylactic strategy to treat VL. PMID:28798743

Multipotent mesenchymal stromal cells decrease transforming growth factor β1 expression in microglia/macrophages and down-regulate plasminogen activator inhibitor 1 expression in astrocytes after stroke.

PubMed

Xin, Hongqi; Chopp, Michael; Shen, Li Hong; Zhang, Rui Lan; Zhang, Li; Zhang, Zheng Gang; Li, Yi

2013-05-10

Multipotent mesenchymal stromal cells (MSCs) decrease the expression of transforming growth factor β1 (TGFβ1) in astrocytes and subsequently decrease astrocytic plasminogen activator inhibitor 1 (PAI-1) level in an autocrine manner. Since activated microglia/macrophages are also a source of TGFβ1 after stroke, we therefore tested whether MSCs regulate TGFβ1 expression in microglia/macrophages and subsequently alters PAI-1 expression after ischemia. TGFβ1 and its downstream effector phosphorylated SMAD 2/3 (p-SMAD 2/3) were measured in mice subjected to middle cerebral artery occlusion (MCAo). MSC treatment significantly decreased TGFβ1 protein expression in both astrocytes and microglia/macrophages in the ischemic boundary zone (IBZ) at day 14 after stroke. However, the p-SMAD 2/3 was only detected in astrocytes and decreased after MSC treatment. In vitro, RT-PCR results showed that the TGFβ1 mRNA level was increased in both astrocytes and microglia/macrophages in an astrocyte-microglia/macrophage co-culture system after oxygen-glucose deprived (OGD) treatment. MSCs treatment significantly decreased the above TGFβ1 mRNA level under OGD conditions, respectively. OGD increased the PAI-1 mRNA in astrocytes in the astrocyte-microglia/macrophage co-culture system, and MSC administration significantly decreased this level. PAI-1 mRNA was very low in microglia/macrophages compared with that in astrocytes under different conditions. Western blot results also verified that MSC administration significantly decreased p-SMAD 2/3 and PAI-1 level in astrocytes in astrocyte-microglia/macrophage co-culture system under OGD conditions. Our in vivo and in vitro data, in concert, suggest that MSCs decrease TGFβ1 expression in microglia/macrophages in the IBZ which contribute to the down-regulation of PAI-1 level in astrocytes. Published by Elsevier Ireland Ltd.

FcγR-induced production of superoxide and inflammatory cytokines is differentially regulated by SHIP through its influence on PI3K and/or Ras/Erk pathways

PubMed Central

Ganesan, Latha P.; Joshi, Trupti; Fang, Huiqing; Kutala, Vijay Kumar; Roda, Julie; Trotta, Rossana; Lehman, Amy; Kuppusamy, Periannan; Byrd, John C.; Carson, William E.; Caligiuri, Michael A.; Tridandapani, Susheela

2006-01-01

Phagocytosis of IgG-coated particles via FcγR is accompanied by the generation of superoxide and inflammatory cytokines, which can cause collateral tissue damage in the absence of regulation. Molecular mechanisms regulating these phagocytosis-associated events are not known. SHIP is an inositol phosphatase that downregulates PI3K-mediated activation events. Here, we have examined the role of SHIP in FcγR-induced production of superoxide and inflammatory cytokines. We report that primary SHIP-deficient bone marrow macrophages produce elevated levels of superoxide upon FcγR clustering. Analysis of the molecular mechanism revealed that SHIP regulates upstream Rac-GTP binding, an obligatory event for superoxide production. Likewise, SHIP-deficient macrophages displayed enhanced IL-1β and IL-6 production in response to FcγR clustering. Interestingly, whereas IL-6 production required activation of both PI3K and Ras/Erk pathways, IL-1β production was dependent only on Ras/Erk activation, suggesting that SHIP may also regulate the Ras/Erk pathway in macrophages. Consistently, SHIP-deficient macrophages displayed enhanced activation of Erk upon FcγR clustering. Inhibition of Ras/Erk or PI3K suppressed the enhanced production of IL-6 in SHIP-deficient macrophages. In contrast, inhibition of Ras/Erk, but not PI3K, suppressed IL-1β production in these cells. Together, these data demonstrate that SHIP regulates phagocytosis-associated events through the inhibition of PI3K and Ras/Erk pathways. PMID:16543474

Single-Cell RNA-Seq Reveals the Transcriptional Landscape and Heterogeneity of Aortic Macrophages in Murine Atherosclerosis.

PubMed

Cochain, Clément; Vafadarnejad, Ehsan; Arampatzi, Panagiota; Jaroslav, Pelisek; Winkels, Holger; Ley, Klaus; Wolf, Dennis; Saliba, Antoine-Emmanuel; Zernecke, Alma

2018-03-15

Rationale: It is assumed that atherosclerotic arteries contain several macrophage subsets endowed with specific functions. The precise identity of these subsets is poorly characterized as they ha ve been defined by the expression of a restricted number of markers. Objective: We have applied single-cell RNA-seq as an unbiased profiling strategy to interrogate and classify aortic macrophage heterogeneity at the single-cell level in atherosclerosis. Methods and Results: We performed single-cell RNA sequencing of total aortic CD45 + cells extracted from the non-diseased (chow fed) and atherosclerotic (11 weeks of high fat diet) aorta of Ldlr -/- mice. Unsupervised clustering singled out 13 distinct aortic cell clusters. Among the myeloid cell populations, Resident-like macrophages with a gene expression profile similar to aortic resident macrophages were found in healthy and diseased aortae, whereas monocytes, monocyte-derived dendritic cells (MoDC), and two populations of macrophages were almost exclusively detectable in atherosclerotic aortae, comprising Inflammatory macrophages showing enrichment in I l1b , and previously undescribed TREM2 hi macrophages. Differential gene expression and gene ontology enrichment analyses revealed specific gene expression patterns distinguishing these three macrophage subsets and MoDC, and uncovered putative functions of each cell type. Notably, TREM2 hi macrophages appeared to be endowed with specialized functions in lipid metabolism and catabolism, and presented a gene expression signature reminiscent of osteoclasts, suggesting a role in lesion calcification. TREM2 expression was moreover detected in human lesional macrophages. Importantly, these macrophage populations were present also in advanced atherosclerosis and in Apoe -/- aortae, indicating relevance of our findings in different stages of atherosclerosis and mouse models. Conclusions: These data unprecedentedly uncovered the transcriptional landscape and phenotypic heterogeneity of aortic macrophages and MoDCs in atherosclerotic and identified previously unrecognized macrophage populations and their gene expression signature, suggesting specialized functions. Our findings will open up novel opportunities to explore distinct myeloid cell populations and their functions in atherosclerosis.

Malondialdehyde-Acetaldehyde (MAA) Adducted Surfactant Protein Alters Macrophage Functions through Scavenger Receptor A

PubMed Central

Sapkota, Muna; Kharbanda, Kusum K.; Wyatt, Todd A.

2016-01-01

Background Reactive aldehydes like acetaldehyde and malondialdehyde generated as a result of alcohol metabolism and cigarette smoke exposure lead to the formation of malondialdehyde-acetaldehyde-adducted proteins (MAA adducts). These aldehydes can adduct to different proteins such as bovine serum album (BSA) and surfactant proteins A or D (SPA, SPD). Macrophages play an important role in innate immunity, but the effect of MAA adducts on macrophage function has not yet been examined. Because macrophage scavenger receptor A (SRA; CD204) mediates the uptake of modified proteins, we hypothesized that the effects of MAA modified proteins on macrophage function are primarily mediated through SRA. Methods and Results We tested this hypothesis by exposing SPD-MAA to macrophages and measuring functions. SPD-MAA treatment significantly stimulated pro-inflammatory cytokine TNF-α release in the macrophage cell line, RAW 264.7. A significant reduction in phagocytosis of zymosan particles was also observed. SPD-MAA stimulated a significant dose-dependent increase in TNF-α and IL-6 release from peritoneal macrophages of WT mice. But a significantly less TNF-α and IL-6 were released from peritoneal macrophages of SRA−/− mice. We observed a significant reduction in phagocytosis of zymosan particles in peritoneal macrophages from WT mice treated with SPD-MAA. No further SPD-MAA-induced reduction was seen in peritoneal macrophages form SRA−/− mice. SPD-MAA treatment significantly increased SRA mRNA expression, but had no effect on surface receptor protein expression. Protein kinase C alpha inhibitor and NF-κB inhibitor significantly reduced pro-inflammatory cytokine release in response to SPD-MAA. Conclusion In conclusion, our data demonstrate that SRA is important for MAA-adducted protein-mediated effect on macrophage functions. PMID:27783409

Maternal low protein diet leads to placental angiogenic compensation via dysregulated M1/M2 macrophages and TNFa expression in Sprague-Dawley rats

USDA-ARS?s Scientific Manuscript database

A maternal low-protein (LP) diet results in low birth weight, increased offspring rapid adipose tissue catch-up growth, adult obesity, and insulin resistance in Sprague-Dawley rats. The placenta plays key roles in nutrient transport and fetal growth. Placental function is dependent on regulation of ...

Induction of Mincle by Helicobacter pylori and consequent anti-inflammatory signaling denote a bacterial survival strategy

PubMed Central

Devi, Savita; Rajakumara, Eerappa; Ahmed, Niyaz

2015-01-01

Evasion of innate immune recognition is one of the key strategies for persistence of Helicobacter pylori, by virtue of its ability to modulate or escape the host innate immune receptors and signaling pathways. C-type lectin receptors (CLRs) predominantly expressed by macrophages are pivotal in tailoring immune response against pathogens. The recognition of glyco or carbohydrate moieties by Mincle (Macrophage inducible C-type lectin) is emerging as a crucial element in anti-fungal and anti-mycobacterial immunity. Herein, we demonstrate the role of Mincle in modulation of innate immune response against H. pylori infection. Our results revealed an upregulated expression of Mincle which was independent of direct host cell contact. Upon computational modelling, Mincle was observed to interact with the Lewis antigens of H. pylori LPS and possibly activating an anti-inflammatory cytokine production, thereby maintaining a balance between pro- and anti-inflammatory cytokine production. Furthermore, siRNA mediated knockdown of Mincle in human macrophages resulted in up regulation of pro-inflammatory cytokines and consequent down regulation of anti-inflammatory cytokines. Collectively, our study demonstrates a novel mechanism employed by H. pylori to escape clearance by exploiting functional plasticity of Mincle to strike a balance between pro-and anti-inflammatory responses ensuring its persistence in the host. PMID:26456705

A Novel Soluble Immune-Type Receptor (SITR) in Teleost Fish: Carp SITR Is Involved in the Nitric Oxide-Mediated Response to a Protozoan Parasite

PubMed Central

Ribeiro, Carla M. S.; Bird, Steve; Raes, Geert; Ghassabeh, Gholamreza H.; Schijns, Virgil E. J. C.; Pontes, Maria J. S. L.; Savelkoul, Huub F. J.; Wiegertjes, Geert F.

2011-01-01

Background The innate immune system relies upon a wide range of germ-line encoded receptors including a large number of immunoglobulin superfamily (IgSF) receptors. Different Ig-like immune receptor families have been reported in mammals, birds, amphibians and fish. Most innate immune receptors of the IgSF are type I transmembrane proteins containing one or more extracellular Ig-like domains and their regulation of effector functions is mediated intracellularly by distinct stimulatory or inhibitory pathways. Methodology/Principal Findings Carp SITR was found in a substracted cDNA repertoire from carp macrophages, enriched for genes up-regulated in response to the protozoan parasite Trypanoplasma borreli. Carp SITR is a type I protein with two extracellular Ig domains in a unique organisation of a N-proximal V/C2 (or I-) type and a C-proximal V-type Ig domain, devoid of a transmembrane domain or any intracytoplasmic signalling motif. The carp SITR C-proximal V-type Ig domain, in particular, has a close sequence similarity and conserved structural characteristics to the mammalian CD300 molecules. By generating an anti-SITR antibody we could show that SITR protein expression was restricted to cells of the myeloid lineage. Carp SITR is abundantly expressed in macrophages and is secreted upon in vitro stimulation with the protozoan parasite T. borreli. Secretion of SITR protein during in vivo T. borreli infection suggests a role for this IgSF receptor in the host response to this protozoan parasite. Overexpression of carp SITR in mouse macrophages and knock-down of SITR protein expression in carp macrophages, using morpholino antisense technology, provided evidence for the involvement of carp SITR in the parasite-induced NO production. Conclusion/Significance We report the structural and functional characterization of a novel soluble immune-type receptor (SITR) in a teleost fish and propose a role for carp SITR in the NO-mediated response to a protozoan parasite. PMID:21305002

Macrophage Polarization in Chronic Inflammatory Diseases: Killers or Builders?

PubMed Central

Baci, Denisa; Tremolati, Marco; Fanuli, Matteo; Farronato, Giampietro; Mortara, Lorenzo

2018-01-01

Macrophages are key cellular components of the innate immunity, acting as the main player in the first-line defence against the pathogens and modulating homeostatic and inflammatory responses. Plasticity is a major feature of macrophages resulting in extreme heterogeneity both in normal and in pathological conditions. Macrophages are not homogenous, and they are generally categorized into two broad but distinct subsets as either classically activated (M1) or alternatively activated (M2). However, macrophages represent a continuum of highly plastic effector cells, resembling a spectrum of diverse phenotype states. Induction of specific macrophage functions is closely related to the surrounding environment that acts as a relevant orchestrator of macrophage functions. This phenomenon, termed polarization, results from cell/cell, cell/molecule interaction, governing macrophage functionality within the hosting tissues. Here, we summarized relevant cellular and molecular mechanisms driving macrophage polarization in “distant” pathological conditions, such as cancer, type 2 diabetes, atherosclerosis, and periodontitis that share macrophage-driven inflammation as a key feature, playing their dual role as killers (M1-like) and/or builders (M2-like). We also dissect the physio/pathological consequences related to macrophage polarization within selected chronic inflammatory diseases, placing polarized macrophages as a relevant hallmark, putative biomarkers, and possible target for prevention/therapy. PMID:29507865

Disease-causing Mutations in the Cystic Fibrosis Transmembrane Conductance Regulator Determine the Functional Responses of Alveolar Macrophages*

PubMed Central

Deriy, Ludmila V.; Gomez, Erwin A.; Zhang, Guangping; Beacham, Daniel W.; Hopson, Jessika A.; Gallan, Alexander J.; Shevchenko, Pavel D.; Bindokas, Vytautas P.; Nelson, Deborah J.

2009-01-01

Alveolar macrophages (AMs) play a major role in host defense against microbial infections in the lung. To perform this function, these cells must ingest and destroy pathogens, generally in phagosomes, as well as secrete a number of products that signal other immune cells to respond. Recently, we demonstrated that murine alveolar macrophages employ the cystic fibrosis transmembrane conductance regulator (CFTR) Cl− channel as a determinant in lysosomal acidification (Di, A., Brown, M. E., Deriy, L. V., Li, C., Szeto, F. L., Chen, Y., Huang, P., Tong, J., Naren, A. P., Bindokas, V., Palfrey, H. C., and Nelson, D. J. (2006) Nat. Cell Biol. 8, 933–944). Lysosomes and phagosomes in murine cftr−/− AMs failed to acidify, and the cells were deficient in bacterial killing compared with wild type controls. Cystic fibrosis is caused by mutations in CFTR and is characterized by chronic lung infections. The information about relationships between the CFTR genotype and the disease phenotype is scarce both on the organismal and cellular level. The most common disease-causing mutation, ΔF508, is found in 70% of patients with cystic fibrosis. The mutant protein fails to fold properly and is targeted for proteosomal degradation. G551D, the second most common mutation, causes loss of function of the protein at the plasma membrane. In this study, we have investigated the impact of CFTR ΔF508 and G551D on a set of core intracellular functions, including organellar acidification, granule secretion, and microbicidal activity in the AM. Utilizing primary AMs from wild type, cftr−/−, as well as mutant mice, we show a tight correlation between CFTR genotype and levels of lysosomal acidification, bacterial killing, and agonist-induced secretory responses, all of which would be expected to contribute to a significant impact on microbial clearance in the lung. PMID:19837664

Signal regulatory protein α associated with the progression of oral leukoplakia and oral squamous cell carcinoma regulates phenotype switch of macrophages.

PubMed

Ye, Xiaojing; Zhang, Jing; Lu, Rui; Zhou, Gang

2016-12-06

Signal regulatory protein α (SIRPα) is a cell-surface protein expressed on macrophages that are regarded as an important component of the tumor microenvironment. The expression of SIRPα in oral leukoplakia (OLK) and oral squamous cell carcinoma (OSCC), and further explored the role of SIRPα on the phenotype, phagocytosis ability, migration, and invasion of macrophages in OSCC were investigated. The expression of SIRPα in OLK was higher than in OSCC, correlating with the expression of CD68 and CD163 on macrophages. After cultured with the conditioned media of oral cancer cells, the expression of SIRPα on THP-1 cells was decreased gradually. In co-culture system, macrophages were induced into M2 phenotype by oral cancer cells. Blockade of SIRPα inhibited phagocytosis ability and IL-6, TNF-α productions of macrophages. In addition, the proliferation, migration, and IL-10, TGF-β productions of macrophages were upregulated after blockade of SIRPα. Macrophages upregulated the expression of SIRPα and phagocytosis ability, and inhibited the migration and invasion when the activation of NF-κB was inhibited by pyrrolidine dithiocarbamate ammonium (PDTC). Hence, SIRPα might play an important role in the progression of OLK and oral cancer, and could be a pivotal therapeutic target in OSCC by regulating the phenotype of macrophages via targeting NF-κB.

Carcinoembryonic antigen-stimulated THP-1 macrophages activate endothelial cells and increase cell-cell adhesion of colorectal cancer cells.

PubMed

Aarons, Cary B; Bajenova, Olga; Andrews, Charles; Heydrick, Stanley; Bushell, Kristen N; Reed, Karen L; Thomas, Peter; Becker, James M; Stucchi, Arthur F

2007-01-01

The liver is the most common site for metastasis by colorectal cancer, and numerous studies have shown a relationship between serum carcinoembryonic antigen (CEA) levels and metastasis to this site. CEA activates hepatic macrophages or Kupffer cells via binding to the CEA receptor (CEA-R), which results in the production of cytokines and the up-regulation of endothelial adhesion molecules, both of which are implicated in hepatic metastasis. Since tissue macrophages implicated in the metastatic process can often be difficult to isolate, the aim of this study was to develop an in vitro model system to study the complex mechanisms of CEA-induced macrophage activation and metastasis. Undifferentiated, human monocytic THP-1 (U-THP) cells were differentiated (D-THP) to macrophages by exposure to 200 ng/ml phorbol myristate acetate (PMA) for 18 h. Immunohistochemistry showed two CEA-R isoforms present in both U- and D-THP cells. The receptors were localized primarily to the nucleus in U-THP cells, while a significant cell-surface presence was observed following PMA-differentiation. Incubation of D-THP-1 cells with CEA resulted in a significant increase in tumor necrosis factor-alpha (TNF-alpha) release over 24 h compared to untreated D-THP-1 or U-THP controls confirming the functionality of these cell surface receptors. U-THP cells were unresponsive to CEA. Attachment of HT-29 cells to human umbilical vein endothelial cells significantly increased at 1 h after incubation with both recombinant TNF-alpha and conditioned media from CEA stimulated D-THP cells by six and eightfold, respectively. This study establishes an in vitro system utilizing a human macrophage cell line expressing functional CEA-Rs to study activation and signaling mechanisms of CEA that facilitate tumor cell attachment to activated endothelial cells. Utilization of this in vitro system may lead to a more complete understanding of the expression and function of CEA-R and facilitate the design of anti-CEA-R therapeutic modalities that may significantly diminish the metastatic potential of CEA overexpressing colorectal tumors.

Structural basis for decreased induction of class IB PI3-kinases expression by MIF inhibitors

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

Singh, Abhay Kumar; Pantouris, Georgios; Borosch, Sebastian

Macrophage migration inhibitory factor (MIF) is a master regulator of proinflammatory cytokines and plays pathological roles when not properly regulated in rheumatoid arthritis, lupus, atherosclerosis, asthma and cancer. Unlike canonical cytokines, MIF has vestigial keto-enol tautomerase activity. Most of the current MIF inhibitors were screened for the inhibition of this enzymatic activity. However, only some of the enzymatic inhibitors inhibit receptor-mediated biological functions of MIF, such as cell recruitment, through an unknown molecular mechanism. The goal of this study was to understand the molecular basis underlying the pharmacological inhibition of biological functions of MIF. Here, we demonstrate how the structuralmore » changes caused upon inhibitor binding translate into the alteration of MIF-induced downstream signalling. Macrophage migration inhibitory factor activates phosphoinositide 3-kinases (PI3Ks) that play a pivotal role in immune cell recruitment in health and disease. There are several different PI3K isoforms, but little is known about how they respond to MIF. We demonstrate that MIF up-regulates the expression of Class IB PI3Ks in leucocytes. We also demonstrate that MIF tautomerase active site inhibitors down-regulate the expression of Class IB PI3Ks as well as leucocyte recruitment in vitro and in vivo. Finally, based on our MIF:inhibitor complex crystal structures, we hypothesize that the reduction in Class IB PI3K expression occurs because of the displacement of Pro1 towards the second loop of MIF upon inhibitor binding, which results in increased flexibility of the loop 2 and sub-optimal MIF binding to its receptors. These results will provide molecular insights for fine-tuning the biological functions of MIF.« less

Regulation of immunophenotype modulation of monocytes-macrophages from M1 into M2 by prostate cancer cell-culture supernatant via transcription factor STAT3.

PubMed

Solís-Martínez, R; Cancino-Marentes, M; Hernández-Flores, G; Ortiz-Lazareno, P; Mandujano-Álvarez, G; Cruz-Gálvez, C; Sierra-Díaz, E; Rodríguez-Padilla, C; Jave-Suárez, L F; Aguilar-Lemarroy, A; Bravo-Cuellar, A

2018-04-01

Transcription factor STAT3 has a prominent innate immunity effect on cancer progression. We determined the regulation of STAT3 in the immunophenotype modulation of macrophages from M1 into M2 induced by the cell-culture supernatant of the Prostate-Cancer line PC3. Monocytes-macrophages from healthy donors were cultured in the supernatant of PC3 cells, membrane proteins, and intracytoplasmic and phosphorylated STAT3 were measured using flow cytometry, while cytokines and growth factors were studied using luminescence. Cytotoxicity and nitric oxide were evaluated via colorimetric assays. The supernatant of PC3 prostate-tumor cells effectively induced macrophages toward an M2 profile, and the expression of phosphorylated STAT3 in the monocytes-macrophages notably increased, and mainly related to IL-10. In the group of monocytes-macrophages treated with a STAT3 inhibitor, the macrophages were induced toward an M1 phenotype. In this study, we showed that the secretion profile of PC3 prostate-cancer cells induces a change in macrophage phenotype from M1 into M2, and that the phenomenon is related to phosphorylation of transcription factor STAT3 and IL-10. Copyright © 2018. Published by Elsevier B.V.

NAMPT-Mediated Salvage Synthesis of NAD+ Controls Morphofunctional Changes of Macrophages

PubMed Central

Venter, Gerda; Oerlemans, Frank T. J. J.; Willemse, Marieke; Wijers, Mietske; Fransen, Jack A. M.; Wieringa, Bé

2014-01-01

Functional morphodynamic behavior of differentiated macrophages is strongly controlled by actin cytoskeleton rearrangements, a process in which also metabolic cofactors ATP and NAD(H) (i.e. NAD+ and NADH) and NADP(H) (i.e. NADP+ and NADPH) play an essential role. Whereas the link to intracellular ATP availability has been studied extensively, much less is known about the relationship between actin cytoskeleton dynamics and intracellular redox state and NAD+-supply. Here, we focus on the role of nicotinamide phosphoribosyltransferase (NAMPT), found in extracellular form as a cytokine and growth factor, and in intracellular form as one of the key enzymes for the production of NAD+ in macrophages. Inhibition of NAD+ salvage synthesis by the NAMPT-specific drug FK866 caused a decrease in cytosolic NAD+ levels in RAW 264.7 and Maf-DKO macrophages and led to significant downregulation of the glycolytic flux without directly affecting cell viability, proliferation, ATP production capacity or mitochondrial respiratory activity. Concomitant with these differential metabolic changes, the capacity for phagocytic ingestion of particles and also substrate adhesion of macrophages were altered. Depletion of cytoplasmic NAD+ induced cell-morphological changes and impaired early adhesion in phagocytosis of zymosan particles as well as spreading performance. Restoration of NAD+ levels by NAD+, NMN, or NADP+ supplementation reversed the inhibitory effects of FK866. We conclude that direct coupling to local, actin-based, cytoskeletal dynamics is an important aspect of NAD+’s cytosolic role in the regulation of morphofunctional characteristics of macrophages. PMID:24824795

Macrophage P2X4 receptors augment bacterial killing and protect against sepsis

PubMed Central

Csóka, Balázs; Németh, Zoltán H.; Szabó, Ildikó; Davies, Daryl L.; Varga, Zoltán V.; Pálóczi, János; Falzoni, Simonetta; Di Virgilio, Francesco; Muramatsu, Rieko; Pacher, Pál

2018-01-01

The macrophage is a major phagocytic cell type, and its impaired function is a primary cause of immune paralysis, organ injury, and death in sepsis. An incomplete understanding of the endogenous molecules that regulate macrophage bactericidal activity is a major barrier for developing effective therapies for sepsis. Using an in vitro killing assay, we report here that the endogenous purine ATP augments the killing of sepsis-causing bacteria by macrophages through P2X4 receptors (P2X4Rs). Using newly developed transgenic mice expressing a bioluminescent ATP probe on the cell surface, we found that extracellular ATP levels increase during sepsis, indicating that ATP may contribute to bacterial killing in vivo. Studies with P2X4R-deficient mice subjected to sepsis confirm the role of extracellular ATP acting on P2X4Rs in killing bacteria and protecting against organ injury and death. Results with adoptive transfer of macrophages, myeloid-specific P2X4R-deficient mice, and P2rx4 tdTomato reporter mice indicate that macrophages are essential for the antibacterial, antiinflammatory, and organ protective effects of P2X4Rs in sepsis. Pharmacological targeting of P2X4Rs with the allosteric activator ivermectin protects against bacterial dissemination and mortality in sepsis. We propose that P2X4Rs represent a promising target for drug development to control bacterial growth in sepsis and other infections. PMID:29875325

Evidence for nucleotide receptor modulation of cross talk between MAP kinase and NF-kappa B signaling pathways in murine RAW 264.7 macrophages.

PubMed

Aga, Mini; Watters, Jyoti J; Pfeiffer, Zachary A; Wiepz, Gregory J; Sommer, Julie A; Bertics, Paul J

2004-04-01

Extracellular nucleotides such as ATP are present in abundance at sites of inflammation and tissue damage, and these agents exert a potent modulatory effect on macrophage/monocyte function via the nucleotide receptor P2X(7). In this regard, after exposure to bacterial LPS, P2X(7) activation augments expression of the inducible nitric oxide (NO) synthase and production of NO in macrophages. Because P2X(7) has been reported to stimulate certain members of the MAP kinase family (ERK1/2) and can enhance the DNA-binding activity of NF-kappa B, we tested the hypothesis that LPS and nucleotides regulate NF-kappa B-dependent inflammatory events via cross talk with MAPK-associated pathways. In this regard, the present studies revealed that cotreatment of macrophages with LPS and the P2X(7)-selective ligand 2'-3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP) results in the cooperative activation of NF-kappa B DNA-binding activity and a sustained attenuation of levels of the NF-kappa B inhibitory protein I kappa B alpha. Interestingly, a persistent reduction in I kappa B alpha levels is also observed when the MEK1/2 inhibitor U0126 is coadministered with LPS, suggesting that components of the MEK/ERK pathway are involved in regulating I kappa B alpha protein expression and/or turnover. The observation that U0126 and BzATP exhibit overlapping actions with respect to LPS-induced changes in I kappa B alpha levels is supported by the finding that Ras activation, which is upstream of MEK/ERK activation, is reduced upon macrophage cotreatment with BzATP and LPS compared with the effects of BzATP treatment alone. These data are consistent with the concept that the Ras/MEK/ERK pathways are involved in regulating NF-kappa B/I kappa B-dependent inflammatory mediator production and suggest a previously unidentified mechanism by which nucleotides can modulate LPS-induced action via cross talk between NF-kappa B and Ras/MEK/MAPK-associated pathways.

Involvement of endoplasmic reticulum stress in the necroptosis of microglia/macrophages after spinal cord injury.

PubMed

Fan, H; Tang, H-B; Kang, J; Shan, L; Song, H; Zhu, K; Wang, J; Ju, G; Wang, Y-Z

2015-12-17

Microglia/macrophages play a crucial role in inflammation after spinal cord injury (SCI). Although extensive studies have been performed on the mechanisms of microglia/macrophage activation and recruitment, how microglia/macrophages are eliminated remains unclear. In the present study, we observed a high-level expression of mixed lineage kinase domain-like protein (MLKL), a key molecule in the execution of necroptosis, in microglia/macrophages after SCI in mice. In vivo PI-labeling and Necrostatin-1 treatment confirmed the necroptosis of microglia/macrophages. Interestingly, our electronic microscopic (EM) study revealed that MLKL localized not only at the membrane but also on the endoplasmic reticulum (ER) of necroptotic microglia/macrophages. Furthermore, receptor-interacting protein 3 (RIP3), another necrosome component, was also found on the ER of necroptotic microglia/macrophages. And Glucose-regulated protein 78 (GRP78), an ER stress sensor, was up-regulated in MLKL-positive microglia/macrophages after SCI, suggesting a possible link between necroptosis and ER stress. In vitro, oxygen-glucose deprivation (OGD) stress induced ER stress and necroptosis in microglia. Inhibiting ER stress by 4-phenylbutyrate (4-PBA) significantly blocked the OGD-induced necroptosis of microglia. In the end, our data showed that, GRP78 and phosphorylated MLKL were co-expressed by the microglia/macrophages in the injured human spinal cord. Taken together, these results suggested that microglia/macrophages undergo an ER-stress involved necroptosis after SCI, implying that ER stress and necroptosis could be manipulated for modulating inflammation post-SCI. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

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

PubMed

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.

TAM receptor-dependent regulation of SOCS3 and MAPKs contributes to pro-inflammatory cytokine downregulation following chronic NOD2 stimulation of human macrophages1

PubMed Central

Zheng, Shasha; Hedl, Matija; Abraham, Clara

2014-01-01

Microbial-induced cytokine regulation is critical to intestinal immune homeostasis. Acute stimulation of NOD2, the Crohn’s disease-associated sensor of bacterial peptidoglycan, induces cytokines. However, cytokines are attenuated after chronic NOD2 and pattern recognition receptor (PRR) stimulation of macrophages; similar attenuation is observed in intestinal macrophages. The role of Tyro3, Axl and Mer (TAM) receptors in regulating chronic PRR 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 pro-inflammatory 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 TGFβ-dependent TAM upregulation in human macrophages, which in turn, upregulated SOCS3 expression. Restoring SOCS3 expression under TAM knockdown conditions restored chronic NOD2-mediated pro-inflammatory cytokine downregulation. In contrast to the upregulated pro-inflammatory 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, MAFK and PU.1 binding to the IL-10 promoter and IL-10 secretion. Therefore, TAM receptors are critical for downregulating pro-inflammatory cytokines under the chronic NOD2 stimulation conditions observed in the intestinal environment. PMID:25567680

[Autophagy modulates the levels of inflammatory cytokines in macrophages induced by lipopolysaccharide].

PubMed

Ren, Caiyuan; Zhang, Xiangying; Shi, Hongbo; Chen, Dexi; Duan, Zhongping; Zhang, Huanhu; Ren, Feng

2017-05-01

Objective To analyze the effect of autophagy on inflammatory response in macrophages induced by lipopolysaccharide (LPS) and investigate its molecular mechanism. Methods Bone marrow mesenchymal stem cells, which were separated from the femora of mice, were cultured and induced to differentiate into primary macrophages in vitro. The inflammatory cell model was established by stimulating the primary macrophages with LPS. Autophagy was inhibited by 3-methyladenine (3-MA) or promoted by rapamycin. Green fluorescent protein-microtubule associated protein 1 light chain 3 (GFP-LC3) plasmid was used to transfect primary macrophages and the percentage of cells with GFP-LC3 puncta were counted in the different groups. The mRNA levels of LC3B, tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), IL-6 and IL-12p40 were detected by real-time quantitative PCR, and the protein levels of LC3B, nuclear factor κB (NF-κB) and IκBα were determined by Western blotting. Results LC3B mRNA and protein expression levels were gradually up-regulated and the autophagosomes increased in the macrophages 2, 4 and 6 hours after treated by LPS. Compared with only LPS treatment group, autophagy inhibition by 3-MA pretreatment promoted the mRNA expressions of inflammatory cytokines including TNF-α, IL-1β, IL-6 and IL-12p40, and the autophagy induction by rapamycin pretreatment suppressed TNF-α, IL-1β, IL-6 and IL-12p40. Meanwhile, 3-MA or rapamycin pretreatment further regulated the protein expressions of IκBα and p-NF-kBp65 induced by LPS in macrophages. Conclusion Autophagy can suppress the LPS-induced inflammatory response in macrophages by regulating NF-κB signaling pathway.

High and low molecular weight hyaluronic acid differentially influence macrophage activation

PubMed Central

Rayahin, Jamie E.; Buhrman, Jason S.; Zhang, Yu; Koh, Timothy J.; Gemeinhart, Richard A.

2015-01-01

Macrophages exhibit phenotypic diversity permitting wide-ranging roles in maintaining physiologic homeostasis. Hyaluronic acid, a major glycosaminoglycan of the extracellular matrix, has been shown to have differential signaling based on its molecular weight. With this in mind, the main objective of this study was to elucidate the role of hyaluronic acid molecular weight on macrophage activation and reprogramming. Changes in macrophage activation were assessed by activation state selective marker measurement, specifically quantitative real time polymerase chain reaction, and cytokine enzyme-linked immunoassays, after macrophage treatment with differing molecular weights of hyaluronic acid under four conditions: the resting state, concurrent with classical activation, and following inflammation involving either classically or alternatively activated macrophages. Regardless of initial polarization state, low molecular weight hyaluronic acid induced a classically activated-like state, confirmed by up-regulation of pro-inflammatory genes, including nos2, tnf, il12b, and cd80, and enhanced secretion of nitric oxide and TNF-α. High molecular weight hyaluronic acid promoted an alternatively activated-like state, confirmed by up regulation of pro-resolving gene transcription, including arg1, il10, and mrc1, and enhanced arginase activity. Overall, our observations suggest that macrophages undergo phenotypic changes dependent on molecular weight of hyaluronan that correspond to either (1) pro-inflammatory response for low molecular weight HA or (2) pro-resolving response for high molecular weight HA. These observations bring significant further understanding of the influence of extracellular matrix polymers, hyaluronic acid in particular, on regulating the inflammatory response of macrophages. This knowledge can be used to guide the design of HA-containing biomaterials to better utilize the natural response to HAs. PMID:26280020

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

PubMed

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.

IL-6 Mediates Macrophage Infiltration after Irradiation via Up-regulation of CCL2/CCL5 in Non-small Cell Lung Cancer.

PubMed

Wang, Xin; Yang, Xiaodong; Tsai, Ying; Yang, Li; Chuang, Kuang-Hsiang; Keng, Peter C; Lee, Soo Ok; Chen, Yuhchyau

2017-01-01

Radiotherapy is effective in reducing primary tumors, however, it may enhance macrophage infiltration to tumor sites, accelerating tumor progression in several ways. We investigated whether radiation can increase macrophage infiltration into non-small cell lung carcinoma (NSCLC) cells. Analysis of in vitro macrophage (differentiated THP-1 cells) migration to either nonirradiated or irradiated tumor cells showed increased migration to the irradiated tumor cells. Because the IL-6 levels in A549 and H157 cells were significantly increased after irradiation, we then investigated whether this increased IL-6 level contributes to radiation-induced macrophage migration. Radiation-induced macrophage infiltration was reduced when IL-6 was knocked down in tumor cells, indicating a positive IL-6 role in this process. To validate this in vitro result, an orthotopic mouse model was developed using a luciferase-tagged H157siIL-6/scramble control (sc) cell set. After tumors developed, the lungs were irradiated, and infiltration of endogenous macrophages and tail-vein injected fluorescent macrophages to tumor sites was investigated. In both groups, increased macrophage infiltration was observed in H157sc cell-derived xenografts compared to H157siIL-6 cell-derived xenografts, confirming the positive IL-6 role in the radiation-induced macrophage infiltration process. In mechanistic dissection studies, radiation-induced up-regulation of CCL2 and CCL5 by IL-6 was detected, and blocking the action of CCL2/CCL5 molecules significantly reduced the number of migrated macrophages to tumor cells after irradiation. These results demonstrate that targeting the IL-6 signaling or CCL2/CCL5 molecules in combination with conventional radiotherapy potentially blocks undesired radiation-induced macrophage infiltration.

CD74-Downregulation of Placental Macrophage-Trophoblastic Interactions in Preeclampsia.

PubMed

Przybyl, Lukasz; Haase, Nadine; Golic, Michaela; Rugor, Julianna; Solano, Maria Emilia; Arck, Petra Clara; Gauster, Martin; Huppertz, Berthold; Emontzpohl, Christoph; Stoppe, Christian; Bernhagen, Jürgen; Leng, Lin; Bucala, Richard; Schulz, Herbert; Heuser, Arnd; Weedon-Fekjær, M Susanne; Johnsen, Guro M; Peetz, Dirk; Luft, Friedrich C; Staff, Anne Cathrine; Müller, Dominik N; Dechend, Ralf; Herse, Florian

2016-06-24

We hypothesized that cluster of differentiation 74 (CD74) downregulation on placental macrophages, leading to altered macrophage-trophoblast interaction, is involved in preeclampsia. Preeclamptic pregnancies feature hypertension, proteinuria, and placental anomalies. Feto-placental macrophages regulate villous trophoblast differentiation during placental development. Disturbance of this well-balanced regulation can lead to pathological pregnancies. We performed whole-genome expression analysis of placental tissue. CD74 was one of the most downregulated genes in placentas from preeclamptic women. By reverse transcriptase-polymerase chain reaction, we confirmed this finding in early-onset (<34 gestational week, n=26) and late-onset (≥34 gestational week, n=24) samples from preeclamptic women, compared with healthy pregnant controls (n=28). CD74 protein levels were analyzed by Western blot and flow cytometry. We identified placental macrophages to express CD74 by immunofluorescence, flow cytometry, and RT-PCR. CD74-positive macrophages were significantly reduced in preeclamptic placentas compared with controls. CD74-silenced macrophages showed that the adhesion molecules ALCAM, ICAM4, and Syndecan-2, as well as macrophage adhesion to trophoblasts were diminished. Naive and activated macrophages lacking CD74 showed a shift toward a proinflammatory signature with an increased secretion of tumor necrosis factor-α, chemokine (C-C motif) ligand 5, and monocyte chemotactic protein-1, when cocultured with trophoblasts compared with control macrophages. Trophoblasts stimulated by these factors express more CYP2J2, sFlt1, TNFα, and IL-8. CD74-knockout mice showed disturbed placental morphology, reduced junctional zone, smaller placentas, and impaired spiral artery remodeling with fetal growth restriction. CD74 downregulation in placental macrophages is present in preeclampsia. CD74 downregulation leads to altered macrophage activation toward a proinflammatory signature and a disturbed crosstalk with trophoblasts. © 2016 American Heart Association, Inc.

SLC15A2 and SLC15A4 Mediate the Transport of Bacterially Derived Di/Tripeptides To Enhance the Nucleotide-Binding Oligomerization Domain-Dependent Immune Response in Mouse Bone Marrow-Derived Macrophages.

PubMed

Hu, Yongjun; Song, Feifeng; Jiang, Huidi; Nuñez, Gabriel; Smith, David E

2018-05-21

There is increasing evidence that proton-coupled oligopeptide transporters (POTs) can transport bacterially derived chemotactic peptides and therefore reside at the critical interface of innate immune responses and regulation. However, there is substantial contention regarding how these bacterial peptides access the cytosol to exert their effects and which POTs are involved in facilitating this process. Thus, the current study proposed to determine the (sub)cellular expression and functional activity of POTs in macrophages derived from mouse bone marrow and to evaluate the effect of specific POT deletion on the production of inflammatory cytokines in wild-type, Pept2 knockout and Pht1 knockout mice. We found that PEPT2 and PHT1 were highly expressed and functionally active in mouse macrophages, but PEPT1 was absent. The fluorescent imaging of muramyl dipeptide-rhodamine clearly demonstrated that PEPT2 was expressed on the plasma membrane of macrophages, whereas PHT1 was expressed on endosomal membranes. Moreover, both transporters could significantly influence the effect of bacterially derived peptide ligands on cytokine stimulation, as shown by the reduced responses in Pept2 knockout and Pht1 knockout mice as compared with wild-type animals. Taken as a whole, our results point to PEPT2 (at plasma membranes) and PHT1 (at endosomal membranes) working in concert to optimize the uptake of bacterial ligands into the cytosol of macrophages, thereby enhancing the production of proinflammatory cytokines. This new paradigm offers significant insight into potential drug development strategies along with transporter-targeted therapies for endocrine, inflammatory, and autoimmune diseases. Copyright © 2018 by The American Association of Immunologists, Inc.

Development of SH2 probes and pull-down assays to detect pathogen-induced, site-specific tyrosine phosphorylation of the TLR adaptor SCIMP.

PubMed

Luo, Lin; Tong, Samuel J; Wall, Adam A; Khromykh, Tatiana; Sweet, Matthew J; Stow, Jennifer L

2017-07-01

Protein tyrosine phosphorylation guides many molecular interactions for cellular functions. SCIMP is a transmembrane adaptor protein (TRAP) family member that mediates selective proinflammatory cytokine responses generated by pathogen-activated Toll-like receptor (TLR) pathways in macrophages. TLR activation triggers SCIMP phosphorylation and selective phosphorylation of distinct tyrosine residues on this adaptor offers the potential for regulating or biasing inflammatory responses. To analyze site-specific phosphorylation events, we developed three probes based on the SH2 domains of known SCIMP effectors, and used them for pull-downs from macrophage extracts. CRISPR-mediated SCIMP-deficient RAW264.7 macrophage-like cells were reconstituted with various phosphorylation-deficient (Y58F, Y96F, Y120F) SCIMPs, and used to demonstrate the specificity of LPS/TLR4-induced, site-specific phosphorylation of SCIMP for the temporal recruitment of the effectors Grb2, Csk and SLP65. Our findings reveal potential for differential SCIMP phosphorylation and specific effectors to influence TLR signaling and inflammatory programs. Furthermore, the use of Csk-SH2 pull-downs to identify additional known and new Csk targets in LPS-activated macrophages reveals the wider utility of our SH2 probes.

Candida albicans Chitin Increases Arginase-1 Activity in Human Macrophages, with an Impact on Macrophage Antimicrobial Functions.

PubMed

Wagener, Jeanette; MacCallum, Donna M; Brown, Gordon D; Gow, Neil A R

2017-01-24

The opportunistic human fungal pathogen Candida albicans can cause a variety of diseases, ranging from superficial mucosal infections to life-threatening systemic infections. Phagocytic cells of the innate immune response, such as neutrophils and macrophages, are important first-line responders to an infection and generate reactive oxygen and nitrogen species as part of their protective antimicrobial response. During an infection, host cells generate nitric oxide through the enzyme inducible nitric oxide synthase (iNOS) to kill the invading pathogen. Inside the phagocyte, iNOS competes with the enzyme arginase-1 for a common substrate, the amino acid l-arginine. Several pathogenic species, including bacteria and parasitic protozoans, actively modulate the production of nitric oxide by inducing their own arginases or the host's arginase activity to prevent the conversion of l-arginine to nitric oxide. We report here that C. albicans blocks nitric oxide production in human-monocyte-derived macrophages by induction of host arginase activity. We further determined that purified chitin (a fungal cell wall polysaccharide) and increased chitin exposure at the fungal cell wall surface induces this host arginase activity. Blocking the C. albicans-induced arginase activity with the arginase-specific substrate inhibitor Nω-hydroxy-nor-arginine (nor-NOHA) or the chitinase inhibitor bisdionin F restored nitric oxide production and increased the efficiency of fungal killing. Moreover, we determined that C. albicans influences macrophage polarization from a classically activated phenotype toward an alternatively activated phenotype, thereby reducing antimicrobial functions and mediating fungal survival. Therefore, C. albicans modulates l-arginine metabolism in macrophages during an infection, potentiating its own survival. The availability and metabolism of amino acids are increasingly recognized as crucial regulators of immune functions. In acute infections, the conversion of the "conditionally essential" amino acid l-arginine by the inducible nitric oxide synthase to nitric oxide is a resistance factor that is produced by the host to fight pathogens. Manipulation of these host defense mechanisms by the pathogen can be key to successful host invasion. We show here that the human opportunistic fungal pathogen Candida albicans influences l-arginine availability for nitric oxide production by induction of the substrate-competing host enzyme arginase-1. This led to a reduced production of nitric oxide and, moreover, reduced eradication of the fungus by human macrophages. We demonstrate that blocking of host arginase-1 activity restored nitric oxide production and increased the killing potential of macrophages. These results highlight the therapeutic potential of l-arginine metabolism in fungal diseases. Copyright © 2017 Wagener et al.

Monosodium Urate Crystals Induce Upregulation of NK1.1-Dependent Killing by Macrophages and Support Tumor-Resident NK1.1+ Monocyte/Macrophage Populations in Antitumor Therapy.

PubMed

Steiger, Stefanie; Kuhn, Sabine; Ronchese, Franca; Harper, Jacquie L

2015-12-01

Macrophages display phenotypic and functional heterogeneity dependent on the changing inflammatory microenvironment. Under some conditions, macrophages can acquire effector functions commonly associated with NK cells. In the current study, we investigated how the endogenous danger signal monosodium urate (MSU) crystals can alter macrophage functions. We report that naive, primary peritoneal macrophages rapidly upregulate the expression of the NK cell-surface marker NK1.1 in response to MSU crystals but not in response to LPS or other urate crystals. NK1.1 upregulation by macrophages was associated with mechanisms including phagocytosis of crystals, NLRP3 inflammasome activation, and autocrine proinflammatory cytokine signaling. Further analysis demonstrated that MSU crystal-activated macrophages exhibited NK cell-like cytotoxic activity against target cells in a perforin/granzyme B-dependent manner. Furthermore, analysis of tumor hemopoietic cell populations showed that effective, MSU-mediated antitumor activity required coadministration with Mycobacterium smegmatis to induce IL-1β production and significant accumulation of monocytes and macrophages (but not granulocytes or dendritic cells) expressing elevated levels of NK1.1. Our findings provide evidence that MSU crystal-activated macrophages have the potential to develop tumoricidal NK cell-like functions that may be exploited to boost antitumor activity in vivo. Copyright © 2015 by The American Association of Immunologists, Inc.

The Pore-Forming Protein Gasdermin D Regulates Interleukin-1 Secretion from Living Macrophages.

PubMed

Evavold, Charles L; Ruan, Jianbin; Tan, Yunhao; Xia, Shiyu; Wu, Hao; Kagan, Jonathan C

2018-01-16

The interleukin-1 (IL-1) family cytokines are cytosolic proteins that exhibit inflammatory activity upon release into the extracellular space. These factors are released following various cell death processes, with pyroptosis being a common mechanism. Recently, it was recognized that phagocytes can achieve a state of hyperactivation, which is defined by their ability to secrete IL-1 while retaining viability, yet it is unclear how IL-1 can be secreted from living cells. Herein, we report that the pyroptosis regulator gasdermin D (GSDMD) was necessary for IL-1β secretion from living macrophages that have been exposed to inflammasome activators, such as bacteria and their products or host-derived oxidized lipids. Cell- and liposome-based assays demonstrated that GSDMD pores were required for IL-1β transport across an intact lipid bilayer. These findings identify a non-pyroptotic function for GSDMD, and raise the possibility that GSDMD pores represent conduits for the secretion of cytosolic cytokines under conditions of cell hyperactivation. Copyright © 2017 Elsevier Inc. All rights reserved.

Leveraging the immune system to treat advanced thyroid cancers.

PubMed

French, Jena D; Bible, Keith; Spitzweg, Christine; Haugen, Bryan R; Ryder, Mabel

2017-06-01

Inflammation has long been associated with the thyroid and with thyroid cancers, raising seminal questions about the role of the immune system in the pathogenesis of advanced thyroid cancers. With a growing understanding of dynamic tumour-immune cell interactions and the mechanisms by which tumour cells evade antitumour immunity, the field of cancer immunotherapy has been revolutionised. In this Review, we provide evidence to support the presence of an antitumour immune response in advanced thyroid cancers linked to cytotoxic T cells and NK cells. This antitumour response, however, is likely blunted by the presence of immunosuppressive pathways within the microenvironment, facilitated by tumour-associated macrophages or increased expression of negative regulators of cytotoxic T-cell function. Current and future efforts to incorporate immune-based therapies into existing tumour cell or endothelial-derived therapies-eg, with kinase inhibitors targeting tumour-associated macrophages or antibodies blocking negative regulators on T cells-could provide improved and durable responses for patients with disease that is otherwise refractory to treatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

An Arginine Deprivation Response Pathway Is Induced in Leishmania during Macrophage Invasion

PubMed Central

Strasser, Rona; Zeituni-Molad, Michal; Bendelak, Keren; Rentsch, Doris; Ephros, Moshe; Wiese, Martin; Jardim, Armando; Myler, Peter J.; Zilberstein, Dan

2016-01-01

Amino acid sensing is an intracellular function that supports nutrient homeostasis, largely through controlled release of amino acids from lysosomal pools. The intracellular pathogen Leishmania resides and proliferates within human macrophage phagolysosomes. Here we describe a new pathway in Leishmania that specifically senses the extracellular levels of arginine, an amino acid that is essential for the parasite. During infection, the macrophage arginine pool is depleted due to its use to produce metabolites (NO and polyamines) that constitute part of the host defense response and its suppression, respectively. We found that parasites respond to this shortage of arginine by up-regulating expression and activity of the Leishmania arginine transporter (LdAAP3), as well as several other transporters. Our analysis indicates the parasite monitors arginine levels in the environment rather than the intracellular pools. Phosphoproteomics and genetic analysis indicates that the arginine-deprivation response is mediated through a mitogen-activated protein kinase-2-dependent signaling cascade. PMID:27043018

Brain perivascular macrophages: characterization and functional roles in health and disease.

PubMed

Faraco, Giuseppe; Park, Laibaik; Anrather, Josef; Iadecola, Costantino

2017-11-01

Perivascular macrophages (PVM) are a distinct population of resident brain macrophages characterized by a close association with the cerebral vasculature. PVM migrate from the yolk sac into the brain early in development and, like microglia, are likely to be a self-renewing cell population that, in the normal state, is not replenished by circulating monocytes. Increasing evidence implicates PVM in several disease processes, ranging from brain infections and immune activation to regulation of the hypothalamic-adrenal axis and neurovascular-neurocognitive dysfunction in the setting of hypertension, Alzheimer disease pathology, or obesity. These effects involve crosstalk between PVM and cerebral endothelial cells, interaction with circulating immune cells, and/or production of reactive oxygen species. Overall, the available evidence supports the idea that PVM are a key component of the brain-resident immune system with broad implications for the pathogenesis of major brain diseases. A better understanding of the biology and pathobiology of PVM may lead to new insights and therapeutic strategies for a wide variety of brain diseases.

Review: Impact of Helminth Infection on Antimycobacterial Immunity—A Focus on the Macrophage

PubMed Central

Lang, Roland; Schick, Judith

2017-01-01

Successful immune control of Mycobacterium tuberculosis (MTB) requires robust CD4+ T cell responses, with IFNγs as the key cytokine promoting killing of intracellular mycobacteria by macrophages. By contrast, helminth infections typically direct the immune system toward a type 2 response, characterized by high levels of the cytokines IL-4 and IL-10, which can antagonize IFNγ production and its biological effects. In many countries with high burden of tuberculosis, helminth infections are endemic and have been associated with increased risk to develop tuberculosis or to inhibit vaccination-induced immunity. Mechanistically, regulation of the antimycobacterial immune response by helminths has been mostly been attributed to the T cell compartment. Here, we review the current status of the literature on the impact of helminths on vaccine-induced and natural immunity to MTB with a focus on the alterations enforced on the capacity of macrophages to function as sensors of mycobacteria and effector cells to control their replication. PMID:29312343

Human recombinant RNASET2-induced inflammatory response and connective tissue remodeling in the medicinal leech.

PubMed

Baranzini, Nicolò; Pedrini, Edoardo; Girardello, Rossana; Tettamanti, Gianluca; de Eguileor, Magda; Taramelli, Roberto; Acquati, Francesco; Grimaldi, Annalisa

2017-05-01

In recent years, several studies have demonstrated that the RNASET2 gene is involved in the control of tumorigenicity in ovarian cancer cells. Furthermore, a role in establishing a functional cross-talk between cancer cells and the surrounding tumor microenvironment has been unveiled for this gene, based on its ability to act as an inducer of the innate immune response. Although several studies have reported on the molecular features of RNASET2, the details on the mechanisms by which this evolutionarily conserved ribonuclease regulates the immune system are still poorly defined. In the effort to clarify this aspect, we report here the effect of recombinant human RNASET2 injection and its role in regulating the innate immune response after bacterial challenge in an invertebrate model, the medicinal leech. We found that recombinant RNASET2 injection induces fibroplasias, connective tissue remodeling and the recruitment of numerous infiltrating cells expressing the specific macrophage markers CD68 and HmAIF1. The RNASET2-mediated chemotactic activity for macrophages has been further confirmed by using a consolidated experimental approach based on injection of the Matrigel biomatrice (MG) supplemented with recombinant RNASET2 in the leech body wall. One week after injection, a large number of CD68 + and HmAIF-1 + macrophages massively infiltrated MG sponges. Finally, in leeches challenged with lipopolysaccharides (LPS) or with the environmental bacteria pathogen Micrococcus nishinomiyaensis, numerous macrophages migrating to the site of inoculation expressed high levels of endogenous RNASET2. Taken together, these results suggest that RNASET2 is likely involved in the initial phase of the inflammatory response in leeches.

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

PubMed

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.

Norcantharidin Facilitates LPS-Mediated Immune Responses by Up-Regulation of AKT/NF-κB Signaling in Macrophages

PubMed Central

Li, Ruimei; Tan, Binghe; Han, Honghui; Liu, Mingyao; Qian, Min; Du, Bing

2012-01-01

Norcantharidin (NCTD), a demethylated analog of cantharidin, is a common used clinical drug to inhibit proliferation and metastasis of cancer cells. But the role of NCTD in modulating immune responses remains unknown. Here, we investigated the function and mechanism of NCTD in regulation of TLR4 associated immune response in macrophages. We evaluated the influence of NCTD on host defense against invaded pathogens by acute peritonitis mouse model, ELISA, Q-PCR, nitrite quantification, phagocytosis assay and gelatin zymography assay. Our data showed that the survival and the serum concentrations of IL-6 and TNF-α were all enhanced by NCTD significantly in peritonitis mouse model. Accordingly, LPS-induced cytokine, nitric oxide and MMP-9 production as well as the phagocytosis of bacteria were all up-regulated by NCTD in a dose dependent manner in both RAW264.7 cells and bone marrow-derived macrophages (BMMs). Then we further analyzed TLR4 associated signaling pathway by Western blot, Immunofluorescence and EMSA in the presence or absence of LPS. The phosphorylation of AKT and p65 at serine 536 but not serine 468 was enhanced obviously by NCTD in a dose dependent manner, whereas the degradation of IκBα was little effected. Consequently, the nuclear translocation and DNA binding ability of NF-κB was also increased by NCTD obviously in RAW264.7 cells. Our results demonstrated that NCTD could facilitate LPS-mediated immune response through promoting the phosphorylation of AKT/p65 and transcriptional activity of NF-κB, thus reprofiling the traditional anti-tumor drug NCTD as a novel immune regulator in promoting host defense against bacterial infection. PMID:22984593

Endoglin regulates renal ischaemia-reperfusion injury.

PubMed

Docherty, Neil G; López-Novoa, José M; Arevalo, Miguel; Düwel, Annette; Rodriguez-Peña, Ana; Pérez-Barriocanal, Fernando; Bernabeu, Carmelo; Eleno, Nélida

2006-08-01

Renal ischaemia-reperfusion (I-R) can cause acute tubular necrosis and chronic renal deterioration. Endoglin, an accessory receptor for Transforming Growth Factor-beta1 (TGF-beta1), is expressed on activated endothelium during macrophage maturation and implicated in the control of fibrosis, angiogenesis and inflammation. Endoglin expression was monitored over 14 days after renal I-R in rats. As endoglin-null mice are not viable, the role of endoglin in I-R was studied by comparing renal I-R injury in haploinsufficient mice (Eng(+/-)) and their wild-type littermates (Eng(+/+)). Renal function, morphology and molecular markers of acute renal injury and inflammation were compared. Endoglin mRNA up-regulation in the post-ischaemic kidneys of rats occurred at 12 h after I-R; endoglin protein levels were elevated throughout the study period. Expression was initially localized to the vascular endothelium, then extended to fibrotic and inflamed areas of the interstitium. Two days after I-R, plasma creatinine elevation and acute tubular necrosis were less marked in Eng(+/-) than in Eng(+/+) mice. Significant up-regulation of endoglin protein was found only in the post-ischaemic kidneys of Eng(+/+) mice and coincided with an increased mRNA expression of the TGF-beta1 and collagen IV (alpha1) chain genes. Significant increases in vascular cell adhesion molecule-1 (VCAM-1) and inducible nitric oxide synthase (iNOS) expression, nitrosative stress, myeloperoxidase activity and CD68 staining for macrophages were evident in post-ischaemic kidneys of Eng(+/+), but not Eng(+/-) mice, suggesting that impaired endothelial activation and macrophage maturation may account for the reduced injury in post-ischaemic kidneys of Eng(+/-) mice. Endoglin is up-regulated in the post-ischaemic kidney and endoglin-haploinsufficient mice are protected from renal I-R injury. Endoglin may play a primary role in promoting inflammatory responses following renal I-R.

Sub-inhibitory tigecycline concentrations induce extracellular matrix binding protein Embp dependent Staphylococcus epidermidis biofilm formation and immune evasion.

PubMed

Weiser, Julian; Henke, Hanae A; Hector, Nina; Both, Anna; Christner, Martin; Büttner, Henning; Kaplan, Jeffery B; Rohde, Holger

2016-09-01

Biofilm-associated Staphylococcus epidermidis implant infections are notoriously reluctant to antibiotic treatment. Here we studied the effect of sub-inhibitory concentrations of penicillin, oxacillin, vancomycin, daptomycin, linezolid and tigecycline on S. epidermidis 1585 biofilm formation, expression of extracellular matrix binding protein (Embp) and potential implications for S. epidermidis - macrophage interactions. Penicillin, vancomycin, daptomycin, and linezolid had no biofilm augmenting effect at any of the concentrations tested. In contrast, at sub-inhibitory concentrations tigecycline and oxacillin exhibited significant biofilm inducing activity. In S. epidermidis 1585, SarA is a negative regulator of giant 1 MDa Embp, and down regulation of sarA induces Embp-dependent assembly of a multi-layered biofilm architecture. Dot blot immune assays, confocal laser scanning microscopy, and qPCR showed that under biofilm inducing conditions, tigecycline augmented embp expression compared to the control grown without antibiotics. Conversely, expression of regulator sarA was suppressed, suggesting that tigecycline exerts its effects on embp expression through SarA. Tigecycline failed to induce biofilm formation in embp transposon mutant 1585-M135, proving that under these conditions Embp up-regulation is necessary for biofilm accumulation. As a functional consequence, tigecycline induced biofilm formation significantly impaired the up-take of S. epidermidis by mouse macrophage-like cell line J774A.1. Our data provide novel evidence for the molecular basis of antibiotic induced biofilm formation, a phenotype associated with inherently increased antimicrobial tolerance. While this could explain failure of antimicrobial therapies, persistence of S. epidermidis infections in the presence of sub-inhibitory antimicrobials is additionally propelled by biofilm-related impairment of macrophage-mediated pathogen eradication. Copyright © 2016 Elsevier GmbH. All rights reserved.

MicroRNA-155 silencing enhances inflammatory response and lipid uptake in oxidized low-density lipoprotein-stimulated human THP-1 macrophages.

PubMed

Huang, Ri-sheng; Hu, Guan-qiong; Lin, Bin; Lin, Zhi-yi; Sun, Cheng-chao

2010-12-01

It has been proposed that the inflammatory response of monocytes/macrophages induced by oxidized low-density lipoprotein (oxLDL) is a key event in the pathogenesis of atherosclerosis. MicroRNA-155 (miR-155) is an important regulator of the immune system and has been shown to be involved in acute inflammatory response. However, the function of miR-155 in oxLDL-stimulated inflammation and atherosclerosis remains unclear. Here, we show that the exposure of human THP-1 macrophages to oxLDL led to a marked up-regulation of miR-155 in a dose-dependent manner. Silencing of endogenous miR-155 in THP-1 cells using locked nucleic acid-modified antisense oligonucleotides significantly enhanced oxLDL-induced lipid uptake, up-regulated the expression of scavenger receptors (lectinlike oxidized LDL receptor-1, cluster of differentiation 36 [CD36], and CD68), and promoted the release of several cytokines including interleukin (IL)-6, -8, and tumor necrosis factor α (TNF-α). Luciferase reporter assay showed that targeting miR-155 promoted nuclear factor-kappa B (NF-κB) nuclear translocation and potentiated the NF-κB-driven transcription activity. Moreover, miR-155 knockdown resulted in a marked increase in the protein amount of myeloid differentiation primary response gene 88 (MyD88), an important adapter protein used by Toll-like receptors to activate the NF-κB pathway. Our data demonstrate that miR-155 serves as a negative feedback regulator in oxLDL-stimulated THP-1 inflammatory responses and lipid uptake and thus might have potential therapeutic implications in atherosclerosis.

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

PubMed Central

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

1992-01-01

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

The decidua of preeclamptic-like BPH/5 mice exhibits an exaggerated inflammatory response during early pregnancy.

PubMed

Heyward, C Y; Sones, J L; Lob, H E; Yuen, L C; Abbott, K E; Huang, W; Begun, Z R; Butler, S D; August, A; Leifer, C A; Davisson, R L

2017-04-01

Preeclampsia is a devastating complication of pregnancy characterized by late-gestation hypertension and proteinuria. Because the only definitive treatment is delivery of the fetus and placenta, preeclampsia contributes to increased morbidity and mortality of both mother and fetus. The BPH/5 mouse model, which spontaneously develops a syndrome strikingly similar to preeclampsia, displays excessive inflammation and suppression of inflammation improves pregnancy outcomes. During early pregnancy, decidual macrophages play an important role in promoting maternal tolerance to fetal antigens and regulating tissue remodeling, two functions that are critical for normal placental development. BPH/5 pregnancies are characterized by abnormal placentation; therefore, we hypothesized that macrophage localization and/or function is altered during early pregnancy at the site of placental formation (the decidua) compared to C57BL/6 controls. At early gestation time points, before the onset of maternal hypertension or proteinuria, there was a reduction in the number of macrophages in BPH/5 decidua and a concomitant increase in activated T cells compared with C57BL/6. BPH/5 decidua also exhibited decreased expression of the immunosuppressive cytokine, IL-10, and increased expression of pro-inflammatory, inducible nitric oxide synthase. Together, these data suggest that a reduction in decidual macrophages during pregnancy is associated with immune activation in BPH/5 mice, inadequate placental development and may contribute to adverse pregnancy outcomes in this model. Copyright © 2017 Elsevier B.V. All rights reserved.

Macrophages: contributors to allograft dysfunction, repair, or innocent bystanders?

PubMed

Mannon, Roslyn B

2012-02-01

Macrophages are members of the innate immune response. However, their role in the adaptive immune response is not known. The purpose of this review is to highlight our current understanding of macrophage structure and function and how they may participate in allograft injury. Studies in acute kidney injury models identify macrophages as key mediators of inflammatory injury, while more recent studies indicate that they may play a reparative role, depending on phenotype - M1 or M2 type macrophages. Mregs, generated in vitro, appear to have immune suppressive abilities and a unique phenotype. In solid-organ transplant, the emphasis of studies has been on acute or chronic injury. These data are derived from animal models using depletion of macrophages or antagonizing their activation and inflammatory responses. The relative contribution of macrophage phenotype in transplantation has not been explored. These studies suggest that macrophages play an injurious role in acute cellular allograft rejection, as well as in chronic injury. Infiltration of an allograft with macrophages is also associated with worse graft function and poor prognosis. Further studies are needed to understand the mechanisms of macrophage-mediated injury, explore their potential reparative role, and determine if they or their functional products are biomarkers of poor graft outcomes.

Co-culture of human bone marrow mesenchymal stem cells and macrophages attenuates lipopolysaccharide-induced inflammation in human corneal epithelial cells.

PubMed

Jeong, Won-Yong; Kim, Ji-Hye; Kim, Chan-Wha

2018-05-01

Dry eye syndrome (DES) is considered as an ocular surface inflammatory disease. Previous studies have shown inflammation plays an important role in the progression and onset of DES. Co-culture of human bone marrow mesenchymal stem cells (HBMSCs) and macrophages showed immunomodulatory effects via regulation of cytokine regulation. Thus, the aim of this study was to investigate the effect of the interaction of these cells on in vitro DES model. The conditioned media (CM) from macrophages, HBMSCs, and HBMSCs + macrophages were treated to human corneal epithelial cells, which showed significant reduction in IL-1α and IL-1β expression levels in HBMSCs + macrophages group. Moreover, the IL-1 Receptor Antagonist (IL-1RA) was highly expressed in the CM from the HBMSCs + macrophages group. Wounded eyes of mice were treated with IL-1RA at 0-100 ng/mL for 16 h, the wound size was reduced. The results of this study might lead to the identification of new therapeutic targets for DES.

CYLD Proteolysis Protects Macrophages from TNF-Mediated Auto-necroptosis Induced by LPS and Licensed by Type I IFN.

PubMed

Legarda, Diana; Justus, Scott J; Ang, Rosalind L; Rikhi, Nimisha; Li, Wenjing; Moran, Thomas M; Zhang, Jianke; Mizoguchi, Emiko; Zelic, Matija; Kelliher, Michelle A; Blander, J Magarian; Ting, Adrian T

2016-06-14

Tumor necrosis factor (TNF) induces necroptosis, a RIPK3/MLKL-dependent form of inflammatory cell death. In response to infection by Gram-negative bacteria, multiple receptors on macrophages, including TLR4, TNF, and type I IFN receptors, are concurrently activated, but it is unclear how they crosstalk to regulate necroptosis. We report that TLR4 activates CASPASE-8 to cleave and remove the deubiquitinase cylindromatosis (CYLD) in a TRIF- and RIPK1-dependent manner to disable necroptosis in macrophages. Inhibiting CASPASE-8 leads to CYLD-dependent necroptosis caused by the TNF produced in response to TLR4 ligation. While lipopolysaccharides (LPS)-induced necroptosis was abrogated in Tnf(-/-) macrophages, a soluble TNF antagonist was not able to do so in Tnf(+/+) macrophages, indicating that necroptosis occurs in a cell-autonomous manner. Surprisingly, TNF-mediated auto-necroptosis of macrophages requires type I IFN, which primes the expression of key necroptosis-signaling molecules, including TNFR2 and MLKL. Thus, the TNF necroptosis pathway is regulated by both negative and positive crosstalk. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Suppressive effect of delta 9-tetrahydrocannabinol in vitro on phagocytosis by murine macrophages.

PubMed

Friedman, M; Cepero, M L; Klein, T; Friedman, H

1986-06-01

Incubation of normal mouse peritoneal cells consisting of over 90% phagocytizing macrophages with delta 9-tetrahydrocannabinol (THC) resulted in a inhibition of phagocytic function. The THC in a dose-related manner suppressed the percentage of macrophages per culture which ingested yeast and the average number of yeast particles ingested by the phagocytizing macrophages. The vehicle used to suspend the THC in vitro, i.e., DMSO, had no detectable effect on macrophage function. Suppression of phagocytosis with no effects on viability or cell number occurred with doses of 10 micrograms or less THC per milliliter culture medium. Measurable suppression also occurred after 24- to 48-hr treatment of the macrophages with the THC. This compound had little if any detectable effect on phagocytosis when added directly to the cultures shortly before testing for phagocytosis. Further studies concerning the effects of THC on macrophage function appear warranted.

Downregulation of monocytic differentiation via modulation of CD147 by 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors.

PubMed

Sasidhar, Manda V; Chevooru, Sai Krishnaveni; Eickelberg, Oliver; Hartung, Hans-Peter; Neuhaus, Oliver

2017-01-01

CD147 is an activation induced glycoprotein that promotes the secretion and activation of matrix metalloproteinases (MMPs) and is upregulated during the differentiation of macrophages. Interestingly, some of the molecular functions of CD147 rely on its glycosylation status: the highly glycosylated forms of CD147 induce MMPs whereas the lowly glycosylated forms inhibit MMP activation. Statins are hydroxy-methylglutaryl coenzyme A reductase inhibitors that block the synthesis of mevalonate, thereby inhibiting all mevalonate-dependent pathways, including isoprenylation, N-glycosylation and cholesterol synthesis. In this study, we investigated the role of statins in the inhibition of macrophage differentiation and the associated process of MMP secretion through modulation of CD147. We observed that differentiation of the human monocytic cell line THP-1 to a macrophage phenotype led to upregulation of CD147 and CD14 and that this effect was inhibited by statins. At the molecular level, statins altered CD147 expression, structure and function by inhibiting isoprenylation and N-glycosylation. In addition, statins induced a shift of CD147 from its highly glycosylated form to its lowly glycosylated form. This shift in N-glycosylation status was accompanied by a decrease in the production and functional activity of MMP-2 and MMP-9. In conclusion, these findings describe a novel molecular mechanism of immune regulation by statins, making them interesting candidates for autoimmune disease therapy.

Dissecting engineered cell types and enhancing cell fate conversion via CellNet

PubMed Central

Morris, Samantha A.; Cahan, Patrick; Li, Hu; Zhao, Anna M.; San Roman, Adrianna K.; Shivdasani, Ramesh A.; Collins, James J.; Daley, George Q.

2014-01-01

SUMMARY Engineering clinically relevant cells in vitro holds promise for regenerative medicine, but most protocols fail to faithfully recapitulate target cell properties. To address this, we developed CellNet, a network biology platform that determines whether engineered cells are equivalent to their target tissues, diagnoses aberrant gene regulatory networks, and prioritizes candidate transcriptional regulators to enhance engineered conversions. Using CellNet, we improved B cell to macrophage conversion, transcriptionally and functionally, by knocking down predicted B cell regulators. Analyzing conversion of fibroblasts to induced hepatocytes (iHeps), CellNet revealed an unexpected intestinal program regulated by the master regulator Cdx2. We observed long-term functional engraftment of mouse colon by iHeps, thereby establishing their broader potential as endoderm progenitors and demonstrating direct conversion of fibroblasts into intestinal epithelium. Our studies illustrate how CellNet can be employed to improve direct conversion and to uncover unappreciated properties of engineered cells. PMID:25126792

Diabetes Mellitus-Induced Long Noncoding RNA Dnm3os Regulates Macrophage Functions and Inflammation via Nuclear Mechanisms.

PubMed

Das, Sadhan; Reddy, Marpadga A; Senapati, Parijat; Stapleton, Kenneth; Lanting, Linda; Wang, Mei; Amaram, Vishnu; Ganguly, Rituparna; Zhang, Lingxiao; Devaraj, Sridevi; Schones, Dustin E; Natarajan, Rama

2018-06-21

Macrophages play key roles in inflammation and diabetic vascular complications. Emerging evidence implicates long noncoding RNAs in inflammation, but their role in macrophage dysfunction associated with inflammatory diabetic complications is unclear and was therefore investigated in this study. RNA-sequencing and real-time quantitative PCR demonstrated that a long noncoding RNA Dnm3os (dynamin 3 opposite strand) is upregulated in bone marrow-derived macrophages from type 2 diabetic db/db mice, diet-induced insulin-resistant mice, and diabetic ApoE -/ - mice, as well as in monocytes from type 2 diabetic patients relative to controls. Diabetic conditions (high glucose and palmitic acid) induced Dnm3os in mouse and human macrophages. Promoter reporter analysis and chromatin immunoprecipitation assays demonstrated that diabetic conditions induce Dnm3os via NF-κB activation. RNA fluorescence in situ hybridization and real-time quantitative PCRs of subcellular fractions demonstrated nuclear localization and chromatin enrichment of Dnm3os in macrophages. Stable overexpression of Dnm3os in macrophages altered global histone modifications and upregulated inflammation and immune response genes and phagocytosis. Conversely, RNAi-mediated knockdown of Dnm3os attenuated these responses. RNA pull-down assays with macrophage nuclear lysates identified nucleolin and ILF-2 (interleukin enhancer-binding factor 2) as protein binding partners of Dnm3os , which was further confirmed by RNA immunoprecipitation and RNA fluorescence in situ hybridization immunofluorescence. Furthermore, nucleolin levels were decreased in diabetic conditions, and its knockdown enhanced Dnm3os -induced inflammatory gene expression and histone H3K9-acetylation at their promoters. These results demonstrate novel mechanisms involving upregulation of long noncoding RNA Dnm3os , disruption of its interaction with nucleolin, and epigenetic modifications at target genes that promote macrophage inflammatory phenotype in diabetes mellitus. The data could lead to long noncoding RNA-based therapies for inflammatory diabetes mellitus complications. © 2018 American Heart Association, Inc.

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

PubMed

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

2017-04-28

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

Centrifugation of Cultured Osteoblasts And Macrophages as a Model To Study How Gravity Regulates The Function of Skeletal Cells

NASA Technical Reports Server (NTRS)

Globus, Ruth K.; Searby, Nancy D.; Almeida, Eduardo A. C.; Sutijono, Darrell; Yu, Joon-Ho; Malouvier, Alexander; Doty, Steven B.; Morey-Holton, Emily; Weinstein, Steven L.; Dalton, Bonnie P. (Technical Monitor)

2000-01-01

Mechanical loading helps define the architecture of weight-bearing bone via the tightly regulated process of skeletal turnover. Turnover occurs by the concerted activity of osteoblasts, responsible for bone formation. and osteoclasts, responsible for bone resorption. Osteoclasts are specialized megakaryon macrophages, which differentiate from monocytes in response to resorption stimuli, such as reduced weight-bearing. Habitation in space dramatically alters musculoskeletal loading, which modulates both cell function and bone structure. Our long-term objective is to define the molecular and cellular mechanisms that mediate skeletal adaptations to altered gravity environments. Our experimental approach is to apply hypergravity loads by centrifugation to rodents and cultured cells. As a first step, we examined the influence of centrifugation on the structure of cancellous bone in rats to test the ability of hypergravity to change skeletal architecture. Since cancellous bone undergoes rapid turnover we expected the most dramatic structural changes to occur in the shape of trabeculae of weight-bearing, cancellous bone. To define the cellular responses to hypergravity loads, we exposed cultured osteoblasts and macrophages to centrifugation. The intraosseous and intramedullary pressures within long bones in vivo reportedly range from 12-40 mm Hg, which would correspond to 18-59 gravity (g) in our cultures. We assumed that hydrostatic pressure from the medium above the cell layer is at least one major component of the mechanical load generated by centrifuging cultured cells. and therefore we exposed the cells to 10-50g. In osteoblasts, we examined the structure of their actin and microtubule networks, production of prostaglandin E2 (PGE2), and cell survival. Analysis of the shape of the cytoskeletal networks provides evidence for the ability of centrifugation to affect cell structure, while the production of PGE2 serves as a convenient marker for mechanical stimulation. We examined cell survival, reasoning that osteoblasts might mold skeletal structure in a hypergravity environment in part by regulating apoptosis and thus the duration of osteoblast productivity. Finally, we tested the influence of centrifugation on microbial activation of a macrophage cell line (RAW264.7). In response to the appropriate hormonal stimulation, this cell line is reportedly capable of undergoing differentiation to express osteoclast markers. In addition, a component of the cell wall of gram-negative bacteria, lipopolysaccaride (LPS), stimulates the formation of osteoclasts in vivo. Thus we tested the influence on centrifugation on RAW264.7 cells stimulated with LPS to provide an index of the function of osteoclast precursors.

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

PubMed Central

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

2016-01-01

Differentiated macrophages can self-renew in tissues and expand long-term in culture, but the gene regulatory mechanisms that accomplish self-renewal in the differentiated state have remained unknown. Here we show that in mice, the transcription factors MafB and c-Maf repress a macrophage-specific enhancer repertoire associated with a gene network controlling self-renewal. Single cell analysis revealed that, in vivo, proliferating resident macrophages can access this network by transient down-regulation of Maf transcription factors. The network also controls embryonic stem cell self-renewal but is associated with distinct embryonic stem cell-specific enhancers. This indicates that distinct lineage-specific enhancer platforms regulate a shared network of genes that control self-renewal potential in both stem and mature cells. PMID:26797145

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

PubMed

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.

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

PubMed

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

2015-10-01

What are the actions of soluble human leukocyte antigen G5 (sHLAG5) on macrophage differentiation? sHLAG5 polarizes the differentiation of macrophages toward a decidual macrophage-like phenotype, which could regulate fetomaternal tolerance and placental development. sHLAG5 is a full-length soluble isoform of human leukocyte antigen implicated in immune tolerance during pregnancy. Low or undetectable circulating level of sHLAG5 in first trimester of pregnancy is associated with pregnancy complications such as pre-eclampsia and spontaneous abortion. Decidual macrophages are located in close proximity to invasive trophoblasts, and are involved in regulating fetomaternal tolerance and placental development. Human peripheral blood monocytes were differentiated into macrophages by treatment with granulocyte macrophage colony-stimulating factor in the presence or absence of recombinant sHLAG5 during the differentiation process. The phenotypes and the biological activities of the resulting macrophages were compared. Recombinant sHLAG5 was produced in Escherichia coli BL21 and the protein identity was verified by tandem mass spectrometry. The expression of macrophage markers were analyzed by flow cytometry and quantitative PCR. Phagocytosis was determined by flow cytometry. Indoleamine 2,3-dioxygenase 1 expression and activity were measured by western blot analysis and kynurenine assay, respectively. Cell proliferation and cell cycling were determined by fluorometric cell proliferation assay and flow cytometry, respectively. Cytokine secretion was determined by cytokine array and ELISA kits. Intracellular cytokine expression was measured by flow cytometry. Cell invasion and migration were determined by trans-well invasion and migration assay, respectively. sHLAG5 drove the differentiation of macrophages with 'immuno-modulatory' characteristics, including reduced expression of M1 macrophage marker CD86 and increased expression of M2 macrophage marker CD163. sHLAG5-polarized macrophages showed enhanced phagocytic activity. They also had higher expression and activity of indoleamine 2,3-dioxygenase 1, a phenotypic marker of decidual macrophages, which inhibited proliferation of autologous T-cells via induction of G0/G1 cell cycle arrest. In addition, sHLAG5-polarized macrophages had an increased secretion of interleukin-6 and C-X-C motif ligand 1, which inhibited interferon-γ production in T-cells and induction of trophoblast invasion, respectively. Most information on the phenotypes and biological activities of human decidual macrophages are based on past literatures. A direct comparison between sHLAG5-polarized macrophages and primary decidual macrophages is required to verify the present observations. This is the first study on the role of sHLAG5 in macrophage differentiation. Further study on the mechanism that regulates the differentiation process of macrophages would enhance our understanding on the physiology of early pregnancy. This work was supported in part by the Hong Kong Research Grant Council Grant HKU774212 and the University of Hong Kong Grant 201309176126. The authors have no competing interests to declare. Nil. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Epigenetic Regulation of the Nitrosative Stress Response and Intracellular Macrophage Survival by Extraintestinal Pathogenic Escherichia coli

PubMed Central

Bateman, Stacey L.; Seed, Patrick C.

2013-01-01

Summary Extraintestinal pathogenic Escherichia coli (ExPEC) reside in the enteric tract as a commensal reservoir, but can transition to a pathogenic state by invading normally sterile niches, establishing infection, and disseminating to invasive sites like the bloodstream. Macrophages are required for ExPEC dissemination, suggesting the pathogen has developed mechanisms to persist within professional phagocytes. Here, we report that FimX, an ExPEC-associated DNA invertase that regulates the major virulence factor type 1 pili (T1P), is also an epigenetic regulator of a LuxR-like response regulator HyxR. FimX regulated hyxR expression through bidirectional phase inversion of its promoter region at sites different from the type 1 pili promoter and independent of integration host factor IHF. In vitro, transition from high to low HyxR expression produced enhanced tolerance of reactive nitrogen intermediates (RNI), primarily through de-repression of hmpA, encoding a nitric oxide detoxifying flavohemoglobin. However, in the macrophage, HyxR produced large effects on intracellular survival in the presence and absence of RNI and independent of Hmp. Collectively, we have shown that the ability of ExPEC to survive in macrophages is contingent upon the proper transition from high to low HyxR expression through epigenetic regulatory control by FimX. PMID:22221182

Impaired structural and functional regeneration of skeletal muscles from β2-adrenoceptor knockout mice

PubMed Central

Silva, M T; Wensing, L A; Brum, P C; Câmara, N O; Miyabara, E H

2014-01-01

Aims β2-adrenergic stimulation causes beneficial effects on structure and function of regenerating muscles; thus, the β2-adrenoceptor may play an important role in the muscle regenerative process. Here, we investigated the role of the β2-adrenoceptor in skeletal muscle regeneration. Methods Tibialis anterior (TA) muscles from β2-adrenoceptor knockout (β2KO) mice were cryolesioned and analysed after 1, 3, 10 and 21 days. The role of β2-adrenoceptor on regenerating muscles was assessed through the analysis of morphological and contractile aspects, M1 and M2 macrophage profile, cAMP content, and activation of TGF-β signalling elements. Results Regenerating muscles from β2KO mice showed decreased calibre of regenerating myofibres and reduced muscle contractile function at 10 days when compared with those from wild type. The increase in cAMP content in muscles at 10 days post-cryolesion was attenuated in the absence of the β2-adrenoceptor. Furthermore, there was an increase in inflammation and in the number of macrophages in regenerating muscles lacking the β2-adrenoceptor at 3 and 10 days, a predominance of M1 macrophage phenotype, a decrease in TβR-I/Smad2/3 activation, and in the Smad4 expression at 3 days, while akirin1 expression increased at 10 days in muscles from β2KO mice when compared to those from wild type. Conclusions Our results suggest that the β2-adrenoceptor contributes to the regulation of the initial phases of muscle regeneration, especially in the control of macrophage recruitment in regenerating muscle through activation of TβR-I/Smad2/3 and reduction in akirin1 expression. These findings have implications for the future development of better therapeutic approaches to prevent or treat muscle injuries. PMID:24938737

The Role of Siglec-1 and SR-BI Interaction in the Phagocytosis of Oxidized Low Density Lipoprotein by Macrophages

PubMed Central

Li, Chang; Zhu, Lin; Wu, Li-juan; Zhong, Ren-qian

2013-01-01

Background Macrophages play a proatherosclerotic role in atherosclerosis via oxLDL uptake. As an adhesion molecular of I-type lectins, Siglec-1 is highly expressed on circulating monocytes and plaque macrophages of atherosclerotic patients, but the exact role of Siglec-1 has not been elucidated. Methods In this study, oxLDL was used to stimulate Siglec-1 and some oxLDL receptors (SR-BI, CD64, CD32B, LOX-1 and TLR-4) expression on bone marrow-derived macrophages, whereas small interfering RNA was used to down-regulate Siglec-1. Meanwhile, an ELISA-based assay for Siglec-1-oxLDL interaction was performed, and co-immunoprecipitation (co-IP) and laser scanning confocal microscopy (LSCM) were used to determine the role of Siglec-1 in oxLDL uptake by macrophages. Results We found that oxLDL could up-regulate the expression of various potential oxLDL receptors, including Siglec-1, in a dose-dependent manner. Moreover, down-regulation of Siglec-1 could attenuate oxLDL uptake by Oil red O staining. LSCM revealed that Siglec-1 and CD64/SR-BI may colocalize on oxLDL-stimulated macrophage surface, whereas co-IP showed that Siglec-1 and SR-BI can be immunoprecipitated by each other. However, no direct interaction between Siglec-1 and oxLDL was found in the in vitro protein interaction system. Conclusions Thus, Siglec-1 can interact with SR-BI in the phagocytosis of oxLDL by macrophages, rather than act as an independent receptor for oxLDL. PMID:23520536

Modulation of Ocular Inflammation by Mesenchymal Stem Cells

DTIC Science & Technology

2017-03-01

mature myeloid cells in 64 host defense and resolution of inflammation, excessive innate immune response can have 65 deleterious effects on tissue...that MSCs can regulate 69 functions of mature innate immune cells , including polarization of inflammatory macrophages 70 into an anti-inflammatory... cells 191 As immune cells are primarily developed in lymphoid organs, single cell suspensions from bone 192 marrow, spleen, and submandibular lymph

T Cells Encountering Myeloid Cells Programmed for Amino Acid-dependent Immunosuppression Use Rictor/mTORC2 Protein for Proliferative Checkpoint Decisions*

PubMed Central

Van de Velde, Lee-Ann; Subramanian, Chitra; Smith, Amber M.; Barron, Luke; Qualls, Joseph E.; Neale, Geoffrey; Alfonso-Pecchio, Adolfo; Jackowski, Suzanne; Rock, Charles O.; Wynn, Thomas A.; Murray, Peter J.

2017-01-01

Modulation of T cell proliferation and function by immunoregulatory myeloid cells are an essential means of preventing self-reactivity and restoring tissue homeostasis. Consumption of amino acids such as arginine and tryptophan by immunoregulatory macrophages is one pathway that suppresses local T cell proliferation. Using a reduced complexity in vitro macrophage-T cell co-culture system, we show that macrophage arginase-1 is the only factor required by M2 macrophages to block T cells in G1, and this effect is mediated by l-arginine elimination rather than metabolite generation. Tracking how T cells adjust their metabolism when deprived of arginine revealed the significance of macrophage-mediated arginine deprivation to T cells. We found mTORC1 activity was unaffected in the initial G1 block. After 2 days of arginine deprivation, mTORC1 activity declined paralleling a selective down-regulation of SREBP target gene expression, whereas mRNAs involved in glycolysis, gluconeogenesis, and T cell activation were unaffected. Cell cycle arrest was reversible at any point by exogenous arginine, suggesting starved T cells remain poised awaiting nutrients. Arginine deprivation-induced cell cycle arrest was mediated in part by Rictor/mTORC2, providing evidence that this nutrient recognition pathway is a central component of how T cells measure environmental arginine. PMID:27903651

Inflammasome-driven catecholamine catabolism in macrophages blunts lipolysis in the aged

PubMed Central

Camell, Christina D.; Sander, Jil; Spadaro, Olga; Lee, Aileen; Nguyen, Kim Y.; Wing, Allison; Goldberg, Emily L.; Youm, Yun-Hee; Brown, Chester W.; Elsworth, John; Rodeheffer, Matthew S.; Schultze, Joachim L.; Dixit, Vishwa Deep

2017-01-01

Catecholamine-induced lipolysis, the first step in generation of energy substrates through hydrolysis of triglycerides (TGs) 1, declines with age 2,3. The defect in mobilization of free fatty acids (FFA) in elderly is accompanied with increased visceral adiposity, lower exercise capacity, failure to maintain core body temperature during cold stress, and reduced ability to survive starvation. While catecholamine signaling in adipocytes is normal in elderly, how lipolysis is impaired in aging remains unknown 2,4. Here we uncover that the adipose tissue macrophages (ATMs) regulate age-related reduction in adipocyte lipolysis by lowering the bioavailability of norepinephrine (NE). Unexpectedly, unbiased whole transcriptome analyses of adipose macrophages revealed that aging upregulates genes controlling catecholamine degradation in an NLRP3 inflammasome-dependent manner. Deletion of NLRP3 in aging restored catecholamine-induced lipolysis through downregulation of growth differentiation factor-3 (GDF3) and monoamine oxidase-a (MAOA) that is known to degrade NE. Consistent with this, deletion of GDF3 in inflammasome-activated macrophages improved lipolysis by decreasing MAOA and caspase-1. Furthermore, inhibition of MAOA reversed age-related reduction in adipose tissue NE concentration and restored lipolysis with increased levels of key lipolytic enzymes, adipose triglyceride lipase (ATGL) and hormone sensitive lipase (HSL). Our study reveals that targeting neuro-innate signaling between sympathetic nervous system and macrophages may offer new approaches to mitigate chronic inflammation-induced metabolic impairment and functional decline. PMID:28953873

Reconstitution of recycling from the phagosomal compartment in streptolysin O-permeabilized macrophages: role of Rab11.

PubMed

Leiva, Natalia; Pavarotti, Martín; Colombo, María I; Damiani, María T

2006-06-10

By phagocytosis, macrophages engulf large particles, microorganisms and senescent cells in vesicles called phagosomes. Many internalized proteins rapidly shuttle back to the plasma membrane following phagosome biogenesis. Here, we report a new approach to the study of recycling from the phagosomal compartment: streptolysin O- (SLO) permeabilized macrophages. In this semi-intact cell system, energy and cytosol are required to efficiently reconstitute recycling transport. Addition of GDPbetaS strongly inhibits this transport step, suggesting that a GTP-binding protein modulates the dynamics of cargo exit from the phagosomal compartment. GTPases of the Rab family control vesicular trafficking, and Rab11 is involved in transferrin receptor recycling. To unravel the role of Rab11 in the phagocytic pathway, we added recombinant proteins to SLO-permeabilized macrophages. Rab11:S25N, a negative mutant, strongly diminishes the release of recycled proteins from phagosomes. In contrast, wild type Rab11 and its positive mutant (Rab11:Q70L) favor this vesicular transport event. Using biochemical and morphological assays, we confirm that overexpression of Rab11:S25N substantially decreases recycling from phagosomes in intact cells. These findings show the requirement of a functional Rab11 for the retrieval to the plasma membrane of phagosomal content. SLO-permeabilized macrophages likely constitute a useful tool to identify new molecules involved in regulating transport along the phagocytic pathway.

T Cells Encountering Myeloid Cells Programmed for Amino Acid-dependent Immunosuppression Use Rictor/mTORC2 Protein for Proliferative Checkpoint Decisions.

PubMed

Van de Velde, Lee-Ann; Subramanian, Chitra; Smith, Amber M; Barron, Luke; Qualls, Joseph E; Neale, Geoffrey; Alfonso-Pecchio, Adolfo; Jackowski, Suzanne; Rock, Charles O; Wynn, Thomas A; Murray, Peter J

2017-01-06

Modulation of T cell proliferation and function by immunoregulatory myeloid cells are an essential means of preventing self-reactivity and restoring tissue homeostasis. Consumption of amino acids such as arginine and tryptophan by immunoregulatory macrophages is one pathway that suppresses local T cell proliferation. Using a reduced complexity in vitro macrophage-T cell co-culture system, we show that macrophage arginase-1 is the only factor required by M2 macrophages to block T cells in G 1 , and this effect is mediated by l-arginine elimination rather than metabolite generation. Tracking how T cells adjust their metabolism when deprived of arginine revealed the significance of macrophage-mediated arginine deprivation to T cells. We found mTORC1 activity was unaffected in the initial G 1 block. After 2 days of arginine deprivation, mTORC1 activity declined paralleling a selective down-regulation of SREBP target gene expression, whereas mRNAs involved in glycolysis, gluconeogenesis, and T cell activation were unaffected. Cell cycle arrest was reversible at any point by exogenous arginine, suggesting starved T cells remain poised awaiting nutrients. Arginine deprivation-induced cell cycle arrest was mediated in part by Rictor/mTORC2, providing evidence that this nutrient recognition pathway is a central component of how T cells measure environmental arginine. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.