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Sample records for activated m2 macrophages

  1. Alternatively activated macrophages (M2 macrophages) in the skin of patient with localized scleroderma.

    PubMed

    Higashi-Kuwata, Nobuyo; Makino, Takamitsu; Inoue, Yuji; Takeya, Motohiro; Ihn, Hironobu

    2009-08-01

    Localized scleroderma is a connective tissue disorder that is limited to the skin and subcutaneous tissue. Macrophages have been reported to be particularly activated in patients with skin disease including systemic sclerosis and are potentially important sources for fibrosis-inducing cytokines, such as transforming growth factor beta. To clarify the features of immunohistochemical characterization of the immune cell infiltrates in localized scleroderma focusing on macrophages, skin biopsy specimens were analysed by immunohistochemistry. The number of cells stained with monoclonal antibodies, CD68, CD163 and CD204, was calculated. An evident macrophage infiltrate and increased number of alternatively activated macrophages (M2 macrophages) in their fibrotic areas were observed along with their severity of inflammation. This study revealed that alternatively activated macrophages (M2 macrophages) may be a potential source of fibrosis-inducing cytokines in localized scleroderma, and may play a crucial role in the pathogenesis of localized scleroderma.

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

    PubMed Central

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

    2016-01-01

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

  3. 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.

  4. Roles of alternatively activated M2 macrophages in allergic contact dermatitis.

    PubMed

    Suzuki, Kotaro; Meguro, Kazuyuki; Nakagomi, Daiki; Nakajima, Hiroshi

    2017-03-17

    Alternatively activated macrophages (M2 macrophages) play key roles in the suppression of Th1 cell responses and the orchestration of tissue repair. However, recent studies have shown that M2 macrophages have potentials to produce high levels of proinflammatory cytokines such as IL-1β, IL-6, and TNF-α, suggesting that M2 macrophages may exacerbate inflammation in some settings. In this regard, we have recently shown that large numbers of M2 macrophages accumulate in the sites of hapten-induced contact hypersensitivity (CHS), an animal model of allergic contact dermatitis, and that M2 macrophages exacerbate hapten-induced CHS by producing matrix metalloproteinase 12 (MMP12). We have also shown that suppressor of cytokine signaling-3 (SOCS3), a member of SOCS family proteins that are cytokine-inducible negative regulators of the JAK/STAT signaling pathways, is highly and preferentially expressed in M2 macrophages in hapten-induced CHS and that SOCS3 expressed in M2 macrophages is involved in the attenuation of CHS by suppressing MMP12 production. These findings underscore the importance of M2 macrophage-derived MMP12 in the development of CHS, and suggest that inhibition of M2 macrophages or MMP12 could be a potential therapeutic strategy for the treatment of allergic contact dermatitis.

  5. HMGB1 enhances the protumoral activities of M2 macrophages by a RAGE-dependent mechanism.

    PubMed

    Rojas, Armando; Delgado-López, Fernando; Perez-Castro, Ramón; Gonzalez, Ileana; Romero, Jacqueline; Rojas, Israel; Araya, Paulina; Añazco, Carolina; Morales, Erik; Llanos, Jorge

    2016-03-01

    The monocyte-macrophage lineage shows a high degree of diversity and plasticity. Once they infiltrate tissues, they may acquire two main functional phenotypes, being known as the classically activated type 1 macrophages (M1) and the alternative activated type 2 macrophages (M2). The M1 phenotype can be induced by bacterial products and interferon-γ and exerts a cytotoxic effect on cancer cells. Conversely, the alternatively activated M2 phenotype is induced by Il-4/IL13 and promotes tumor cell growth and vascularization. Although receptor for advanced glycation end-products (RAGE) engagement in M1 macrophages has been reported by several groups to promote inflammation, nothing is known about the functionality of RAGE in M2 macrophages. In the current study, we demonstrate that RAGE is equally expressed in both macrophage phenotypes and that RAGE activation by high-mobility group protein box1 (HMGB1) promotes protumoral activities of M2 macrophages. MKN45 cells co-cultured with M2 macrophages treated with HMGB1 at different times displayed higher invasive abilities. Additionally, conditioned medium from HMGB1-treated M2 macrophages promotes angiogenesis in vitro. RAGE-targeting knockdown abrogates these activities. Overall, the present findings suggest that HMGB1 may contribute, by a RAGE-dependent mechanism, to the protumoral activities of the M2 phenotype.

  6. Understanding the Mysterious M2 Macrophage through Activation Markers and Effector Mechanisms

    PubMed Central

    Rőszer, Tamás

    2015-01-01

    The alternatively activated or M2 macrophages are immune cells with high phenotypic heterogeneity and are governing functions at the interface of immunity, tissue homeostasis, metabolism, and endocrine signaling. Today the M2 macrophages are identified based on the expression pattern of a set of M2 markers. These markers are transmembrane glycoproteins, scavenger receptors, enzymes, growth factors, hormones, cytokines, and cytokine receptors with diverse and often yet unexplored functions. This review discusses whether these M2 markers can be reliably used to identify M2 macrophages and define their functional subdivisions. Also, it provides an update on the novel signals of the tissue environment and the neuroendocrine system which shape the M2 activation. The possible evolutionary roots of the M2 macrophage functions are also discussed. PMID:26089604

  7. Chronic hepatitis C infection–induced liver fibrogenesis is associated with M2 macrophage activation

    PubMed Central

    Bility, Moses T.; Nio, Kouki; Li, Feng; McGivern, David R.; Lemon, Stanley M.; Feeney, Eoin R.; Chung, Raymond T.; Su, Lishan

    2016-01-01

    The immuno-pathogenic mechanisms of chronic hepatitis C virus (HCV) infection remain to be elucidated and pose a major hurdle in treating or preventing chronic HCV-induced advanced liver diseases such as cirrhosis. Macrophages are a major component of the inflammatory milieu in chronic HCV–induced liver disease, and are generally derived from circulating inflammatory monocytes; however very little is known about their role in liver diseases. To investigate the activation and role of macrophages in chronic HCV–induced liver fibrosis, we utilized a recently developed humanized mouse model with autologous human immune and liver cells, human liver and blood samples and cell culture models of monocyte/macrophage and/or hepatic stellate cell activation. We showed that M2 macrophage activation was associated with liver fibrosis during chronic HCV infection in the livers of both humanized mice and patients, and direct-acting antiviral therapy attenuated M2 macrophage activation and associated liver fibrosis. We demonstrated that supernatant from HCV-infected liver cells activated human monocytes/macrophages with M2-like phenotypes. Importantly, HCV-activated monocytes/macrophages promoted hepatic stellate cell activation. These results suggest a critical role for M2 macrophage induction in chronic HCV-associated immune dysregulation and liver fibrosis. PMID:28000758

  8. Chronic hepatitis C infection-induced liver fibrogenesis is associated with M2 macrophage activation.

    PubMed

    Bility, Moses T; Nio, Kouki; Li, Feng; McGivern, David R; Lemon, Stanley M; Feeney, Eoin R; Chung, Raymond T; Su, Lishan

    2016-12-21

    The immuno-pathogenic mechanisms of chronic hepatitis C virus (HCV) infection remain to be elucidated and pose a major hurdle in treating or preventing chronic HCV-induced advanced liver diseases such as cirrhosis. Macrophages are a major component of the inflammatory milieu in chronic HCV-induced liver disease, and are generally derived from circulating inflammatory monocytes; however very little is known about their role in liver diseases. To investigate the activation and role of macrophages in chronic HCV-induced liver fibrosis, we utilized a recently developed humanized mouse model with autologous human immune and liver cells, human liver and blood samples and cell culture models of monocyte/macrophage and/or hepatic stellate cell activation. We showed that M2 macrophage activation was associated with liver fibrosis during chronic HCV infection in the livers of both humanized mice and patients, and direct-acting antiviral therapy attenuated M2 macrophage activation and associated liver fibrosis. We demonstrated that supernatant from HCV-infected liver cells activated human monocytes/macrophages with M2-like phenotypes. Importantly, HCV-activated monocytes/macrophages promoted hepatic stellate cell activation. These results suggest a critical role for M2 macrophage induction in chronic HCV-associated immune dysregulation and liver fibrosis.

  9. Serum from patients with systemic vasculitis induces alternatively activated macrophage M2c polarization.

    PubMed

    Ohlsson, Susanne M; Linge, Carl Petrus; Gullstrand, Birgitta; Lood, Christian; Johansson, Asa; Ohlsson, Sophie; Lundqvist, Andrea; Bengtsson, Anders A; Carlsson, Fredric; Hellmark, Thomas

    2014-01-01

    Anti-neutrophil cytoplasmic antibody associated vasculitides (AAV) are conditions defined by an autoimmune small vessel inflammation. Dying neutrophils are found around the inflamed vessels and the balance between infiltrating neutrophils and macrophages is important to prevent autoimmunity. Here we investigate how sera from AAV patients may regulate macrophage polarization and function. Macrophages from healthy individuals were differentiated into M0, M1, M2a, M2b or M2c macrophages using a standardized protocol, and phenotyped according to their expression surface markers and cytokine production. These phenotypes were compared with those of macrophages stimulated with serum from AAV patients or healthy controls. While the healthy control sera induced a M0 macrophage, AAV serum promoted polarization towards the M2c subtype. No sera induced M1, M2a or M2b macrophages. The M2c subtype showed increased phagocytosis capacity compared with the other subtypes. The M2c polarization found in AAV is consistent with previous reports of increased levels of M2c-associated cytokines.

  10. IKKβ Activity Drives Fetal Lung Macrophage Maturation Along a Non-M1/M2 Paradigm

    PubMed Central

    Stouch, Ashley N.; Zaynagetdinov, Rinat; Barham, Whitney J.; Stinnett, Amanda M.; Slaughter, James C.; Yull, Fiona E.; Hoffman, Hal M.; Blackwell, Timothy S.; Prince, Lawrence S.

    2014-01-01

    In preterm infants, exposure to inflammation increases the risk of bronchopulmonary dysplasia, a chronic, developmental lung disease. While macrophages are the key cells that initiate lung inflammation, less is known about lung macrophage phenotype and maturation. We hypothesized that fetal lung macrophages mature into distinct subpopulations during mouse development, and that activation could influence macrophage maturation. Expression of the fetal macrophage markers CD68, CD86, CD206, Ym1, fibrinogen-like protein 2 (FGL2), and indolamine-2, 3-dioxygenase (Ido1) were developmentally regulated, with each marker having different temporal patterns. Flow cytometry analysis showed macrophages within the fetal lung were less diverse than the distinctly separate subpopulations in newborn and adult lungs. Similar to adult alveolar macrophages, fetal lung macrophages responded to the TLR4 agonist LPS and the alternative activation cytokines IL-4 and IL-13. Using a macrophage-specific constitutively active IKKβ transgenic model (IKFM), we demonstrated that macrophage activation increased proinflammatory gene expression and reduced the response of fetal lung macrophages to IL-4 and IL-13. Activation also increased fetal lung macrophage proliferation. Fetal IKFM lungs contained increased percentages of more mature, CD11bloF4/80hi cells that also expressed higher levels of the alternative activation markers CD204 and CD206. Development of fetal lung macrophages into mature alveolar macrophages may therefore include features of both proinflammatory and alternative activation paradigms. PMID:24981452

  11. Mesenchymal stem cells ameliorate rhabdomyolysis-induced acute kidney injury via the activation of M2 macrophages.

    PubMed

    Geng, Yanqiu; Zhang, Li; Fu, Bo; Zhang, Jianrong; Hong, Quan; Hu, Jie; Li, Diangeng; Luo, Congjuan; Cui, Shaoyuan; Zhu, Fei; Chen, Xiangmei

    2014-06-24

    The mortality of rhabdomyolysis-induced acute kidney injury (AKI) is still high, as there is no effective therapy. It has been shown that bone marrow-derived mesenchymal stem cells (MSCs) can induce M2 macrophages, which mediate MSC protection in other experimental inflammation-related organ injury. This study was designed to investigate the protective effects of macrophage activation in MSC therapy of rhabdomyolysis-induced AKI. MSCs were injected into glycerol-induced rhabdomyolysis mice. Renal injury was evaluated using the serum creatinine, urea nitrogen, renal pathology and acute tubular necrosis score. The distribution of MSCs was detected using two-photon fluorescence confocal imaging. Immunofluorescence of anti-F4/80 and anti-CD206 was performed to determine macrophages and M2 macrophages in the tissues of the kidney, and M2 macrophage infiltration was also evaluated using western blotting analyses. After depletion of macrophages using clodronate liposomes at the phase of kidney repair, renal injury was re-evaluated. RAW 264.7 macrophages were incubated with lipopolysaccharide and co-cultured with MSCs and subsequently visualised using immunofluorescence staining and flow cytometry analysis. Finally, disparate phenotype macrophages, including normal macrophages (M0), lipopolysaccharide-stimulated macrophages (M1), and MSC-co-cultured macrophages (M2), were infused into mice with AKI, which were pre-treated with liposomal clodronate. In vivo infusion of MSCs protected AKI mice from renal function impairment and severe tubular injury, which was accompanied by a time-dependent increase in CD206-positive M2 macrophage infiltration. In addition, depleting macrophages with clodronate delayed restoration of AKI. In vitro, macrophages co-cultured with MSCs acquired an anti-inflammatory M2 phenotype, which was characterised by an increased expression of CD206 and the secretory cytokine interleukin (IL)-10. The concentrations of IL-10, IL-6 and tumor necrosis factor

  12. [Recombinant Trichinella spiralis-53000 protein alleviates liver damage due to lipopolysaccharides via M2 macrophage activation].

    PubMed

    Chen, Zhibin; Tang, Hao; Li, Zhenyu; Liang, Yanbing; Wu, Jingguo; Zeng, Lijin; Yang, Qing; Liang, Huaping; Ma, Zhongfu

    2014-08-01

    To evaluate if recombinant Trichinella spiralis-53 000 protein (rTsP53) could alleviate liver damage caused by lipopolysaccharides (LPS) via M2 macrophage activation. Sixty male BALB/c mice were randomly divided into LPS group, LPS + phosphate buffer saline (PBS) group and rTsP53 intervention group by random number table, with 20 mice in each group. Intraperitoneal injection of 15 μg/kg LPS was performed for all the mice in the three groups after 8 hours of fasting. The mice in LPS + PBS group were injected with PBS after 1 hour of LPS injection. The mice in the rTsP53 intervention group were injected with rTsP53 (5 mg/kg) after 1 hour of LPS injection. After 48 hours all the mice were sacrificed. Peritoneal macrophages were harvested and flow cytometry (FCM) was used to detect markers CCR7 (M1) and CD206 (M2) of macrophages. Hepatic tissue was harvested for pathological study after hematoxylin-eosin (HE) staining, and double staining immunofluorescence was used to detect F4/80⁺ HLA-DR⁺ and F4/80⁺ CD163⁺. Peripheral blood serum was harvested to detect the levels of aspartate transaminase (AST) and alanine transaminase (ALT). Compared with LPS and LPS + PBS groups, survival rate of mice of rTsP53 intervention group was significantly elevated (90% vs. 25%, 30%, both P<0.01), and the pathological injury of the liver was significantly ameliorated, and the hepatic structure was better preserved. The transaminase in rTsP53 intervention group was significantly lower than that of LPS and LPS + PBS groups (ALT: 97.7 ± 8.5 U/L vs. 181.7 ± 19.5 U/L, 173.7 ± 17.2 U/L; AST: 142.7 ± 12.1 U/L vs. 235.7 ± 9.9 U/L, 213.7 ± 6.7 U/L, all P<0.05), FITC-CD206⁺ proportion of peritoneal macrophage was significantly higher [(17.75 ± 0.30)% vs. (1.38 ± 0.13)%, (1.36 ± 0.05)%, both P<0.05] while PE-CCR7(+) proportion [(6.89 ± 0.11)% vs. (15.30 ± 0.64)%, (14.96 ± 0.93)%, both P<0.05] was significantly lower. Fluorescence intensity of macrophages with F4/80⁺ CD163

  13. Haemophilus ducreyi-induced interleukin-10 promotes a mixed M1 and M2 activation program in human macrophages.

    PubMed

    Li, Wei; Katz, Barry P; Spinola, Stanley M

    2012-12-01

    During microbial infection, macrophages are polarized to classically activated (M1) or alternatively activated (M2) cells in response to microbial components and host immune mediators. Proper polarization of macrophages is critical for bacterial clearance. To study the role of macrophage polarization during Haemophilus ducreyi infection, we analyzed a panel of macrophage surface markers in skin biopsy specimens of pustules obtained from experimentally infected volunteers. Lesional macrophages expressed markers characteristic of both M1 and M2 polarization. Monocyte-derived macrophages (MDM) also expressed a mixed M1 and M2 profile of surface markers and cytokines/chemokines upon infection with H. ducreyi in vitro. Endogenous interleukin 10 (IL-10) produced by infected MDM downregulated and enhanced expression of several M1 and M2 markers, respectively. Bacterial uptake, mediated mainly by class A scavenger receptors, and activation of mitogen-activated protein kinase and phosphoinositide 3-kinase signaling pathways were required for H. ducreyi-induced IL-10 production in MDM. Compared to M1 cells, IL-10-polarized M2 cells displayed enhanced phagocytic activity against H. ducreyi and similar bacterial killing. Thus, IL-10-modulated macrophage polarization may contribute to H. ducreyi clearance during human infection.

  14. Glucagon-like peptide-1 (GLP-1) induces M2 polarization of human macrophages via STAT3 activation.

    PubMed

    Shiraishi, Daisuke; Fujiwara, Yukio; Komohara, Yoshihiro; Mizuta, Hiroshi; Takeya, Motohiro

    2012-08-24

    It is known that glucagon-like peptide-1 (GLP-1) is a hormone secreted postprandially from the L-cells of the small intestine and regulates glucose homeostasis. GLP-1 is now used for the treatment of diabetes because of its beneficial role against insulin resistance. The GLP-1 receptor (GLP-1R) is expressed on many cell types, including macrophages, and GLP-1 suppresses the development of atherosclerosis by inhibiting macrophage function. However, there have so far been few studies that have investigated the significance of GLP-1/GLP-1R signaling in macrophage activation. In the present study, we examined the effect of GLP-1 and exenatide, a GLP-1R agonist, on human monocyte-derived macrophage (HMDM) activation. We found that GLP-1 induced signal transducer and activator of transcription 3 (STAT3) activation. Silencing of GLP-1R suppressed the GLP-1-induced STAT3 activation. In addition, alternatively activated (M2) macrophage-related molecules, such as IL-10, CD163, and CD204 in HMDM, were significantly upregulated by GLP-1. Furthermore, the co-culture of 3T3-L1 adipocytes with GLP-1-treated RAW 264.7 macrophages increased the secretion of adiponectin compared to co-culture of the 3T3-L1 adipocytes with untreated RAW 264.7 macrophages. Our results demonstrate that GLP-1 induces macrophage polarization toward the M2 phenotype, which may contribute to the protective effects of GLP-1 against diabetes and cardiovascular diseases.

  15. Atorvastatin promotes human monocyte differentiation toward alternative M2 macrophages through p38 mitogen-activated protein kinase-dependent peroxisome proliferator-activated receptor γ activation.

    PubMed

    Zhang, Ou; Zhang, Jinying

    2015-05-01

    M1 and M2 macrophages are detectable in human atherosclerotic lesions, and M2 macrophages are present at locations distant from the lipid core in more stable zones of the plaque and appear to exert anti-inflammatory properties on M1 macrophages. Peroxisome proliferator-activated receptor (PPAR) γ promotes the differentiation of monocytes into anti-inflammatory M2 macrophages. Although both statins and PPARγ ligands have been reported to protect against the progression of atherosclerosis, no data are currently available regarding the implication of statins in the alternative differentiation of human monocytes. In the present study, we hypothesized that atorvastatin may exert novel effects to prime human monocytes toward an anti-inflammatory alternative M2 phenotype. To this aim, we first found that abundant M2 markers were expressed in human circulating monocytes after atorvastatin treatment. Moreover, atorvastatin was able to induce PPARγ expression and activation in human monocytes in vivo and in vitro, resulting in priming primary human monocytes differentiation into M2 macrophages with a more pronounced paracrine anti-inflammatory activity in M1 macrophages. Additional data with molecular approaches revealed that p38 mitogen-activated protein kinase (MAPK) but not extracellular signal-regulated kinase (ERK) 1/2 activation was involved in atorvastatin-mediated PPARγ activation and enhanced alternative M2 macrophage phenotype. Collectively, our data demonstrated that atorvastatin promotes human monocyte differentiation toward alternative M2 macrophages via p38 MAPK-dependent PPARγ activation.

  16. Carbon nanotubes activate macrophages into a M1/M2 mixed status: recruiting naïve macrophages and supporting angiogenesis.

    PubMed

    Meng, Jie; Li, Xiaojin; Wang, Chuan; Guo, Hua; Liu, Jian; Xu, Haiyan

    2015-02-11

    The potential of carbon nanotubes (CNTs) in medical applications has been attracting constant research interest as well as raising concerns related to toxicity. The immune system serves as the first line of defense against invasion. In this work, interactions of oxidized multiwalled carbon nanotubes (MWCNT) with macrophages were investigated to unravel the activation profile of macrophages, using cytokine array, ELISA assay, transwell assay, confocal microscopy, and reactive oxygen species examination. Results show that MWCNT initiate phagocytosis of macrophages and upregulate CD14, CD11b, TLR-4/MD2, and CD206, which does not alter the MHCII expression of the macrophages. The macrophages engulfing MWCNT (MWCNT-RAW) secrete a large amount of MIP-1α and MIP-2 to recruit naïve macrophages and produce angiogenesis-related cytokines MMP-9 and VEGF, while inducing much lower levels of proinflammatory cytokines than those activated by LPS. In conclusion, MWCNT activate macrophages into a M1/M2 mixed status, which allows the cells to recruit naïve macrophages and support angiogenesis.

  17. Negative Immune Regulator TIPE2 Promotes M2 Macrophage Differentiation through the Activation of PI3K-AKT Signaling Pathway

    PubMed Central

    Geng, Wenwen; Chen, Youhai H.; Zhang, Cui

    2017-01-01

    Macrophages play important roles in the regulation of the innate and adaptive immune responses. Classically activated macrophages and alternatively activated macrophages are the two major forms of macrophages and have opposing functionalities. Tumor necrosis factor-α-induced protein 8–2 is expressed primarily by immune cells and negatively regulates type 1 innate and adaptive immune responses to maintain immune tolerance. While previous studies indicate that TIPE2 promotes M2 but inhibits M1 macrophage differentiation, the underlying molecular mechanism by which TIPE2 promotes M2 macrophage differentiation remains unclear. Our current study shows that TIPE2-deficient bone-marrow cells are defective in IL-4 induced M2 macrophage differentiation in vitro. Mechanistic studies revealed that TIPE2 promotes phosphoinositide metabolism and the activation of the down-stream AKT signaling pathway, which in turn leads to the expression of markers specific for M2 macrophages. In addition, our results showed that Tipe2-deficiency does not affect the activation of the JAK-STAT6 signaling pathway that also plays an important role during M2 macrophage differentiation. Taken together, these results indicate that TIPE2 promotes M2 macrophage differentiation through the activation of PI3K-AKT signaling pathway, and may play an important role during the resolution of inflammation, parasite control, as well as tissue repair. PMID:28122045

  18. In utero nicotine exposure promotes M2 activation in neonatal mouse alveolar macrophages.

    PubMed

    Wongtrakool, Cherry; Grooms, Kora; Ping, Xiao-Du; Rivera, Hilda; Ward, Janine; Roser-Page, Susanne; Roman, Jesse; Brown, Lou Ann S; Gauthier, Theresa W

    2012-08-01

    Maternal smoking in utero has been associated with adverse health outcomes including lower respiratory tract infections in infants and children, but the mechanisms underlying these associations continue to be investigated. We hypothesized that nicotine plays a significant role in mediating the effects of maternal tobacco smoke on the function of the neonatal alveolar macrophage (AM), the resident immune cell in the neonatal lung. Primary AMs were isolated at postnatal day 7 from a murine model of in utero nicotine exposure. The murine AM cell line MH-S was used for additional in vitro studies. In utero nicotine increased interleukin-13 and transforming growth factor-β1 (TGFβ1) in the neonatal lung. Nicotine-exposed AMs demonstrated increased TGFβ1 and increased markers of alternative activation with diminished phagocytic function. However, AMs from mice deficient in the α7 nicotinic acetylcholine receptor (α7 nAChR) had less TGFβ1, reduced alternative activation, and improved phagocytic functioning despite similar in utero nicotine exposure. In utero nicotine exposure, mediated in part via the α7 nAChR, may increase the risk of lower respiratory tract infections in neonates by changing the resting state of AM toward alternative activation. These findings have important implications for immune responses in the nicotine-exposed neonatal lung.

  19. The effect of mineral trioxide aggregate on phagocytic activity and production of reactive oxygen, nitrogen species and arginase activity by M1 and M2 macrophages.

    PubMed

    Rezende, T M B; Vieira, L Q; Cardoso, F P; Oliveira, R R; de Oliveira Mendes, S T; Jorge, M L R; Ribeiro Sobrinho, A P

    2007-08-01

    To assess the influence of co-culture with mineral trioxide aggregate (MTA) on phagocytosis and the production of reactive oxygen intermediates (ROI) and nitrogen (NO) species and the arginase activity by M1 and M2 peritoneal macrophages. Cellular viability, adherence and phagocytosis of Saccharomyces boulardii were assayed in the presence of MTA. Macrophages were stimulated with zymosan for ROI assays and with Fusobacterium nucleatum and Peptostreptococcus anaerobius and IFN-gamma for NO production and arginase activity, when in contact with capillaries containing MTA. Data were analysed by T, anova, Kruskall-Wallis and Mann-Whitney tests. M2 macrophages displayed greater cellular viability in polypropylene tubes, greater ability to ingest yeast and smaller production of ROI and higher arginase activity when compared with M1 macrophages. Both macrophages, M1 and M2, presented similar cell adherence and NO production. The addition of bacterial preparations to macrophages interfered with NO and arginase productions. MTA did not interfere with any of the parameters measured. Phagocytosis and the ability of the two macrophage subtypes to eliminate microbes were not affected by MTA.

  20. Adrenergic receptor β2 activation by stress promotes breast cancer progression through macrophages M2 polarization in tumor microenvironment.

    PubMed

    Qin, Jun-fang; Jin, Feng-jiao; Li, Ning; Guan, Hai-tao; Lan, Lan; Ni, Hong; Wang, Yue

    2015-05-01

    Stress and its related hormones epinephrine (E) and norepinephrine (NE) play a crucial role in tumor progression. Macrophages in the tumor microenvironment (TME) polarized to M2 is also a vital pathway for tumor deterioration. Here, we explore the underlying role of macrophages in the effect of stress and E promoting breast cancer growth. It was found that the weight and volume of tumor in tumor bearing mice were increased, and dramatically accompanied with the rising E level after chronic stress using social isolation. What is most noteworthy, the number of M2 macrophages inside tumor was up-regulated with it. The effects of E treatment appear to be directly related to the change of M2 phenotype is reproduced in vitro. Moreover, E receptor ADRβ2 involved in E promoting M2 polarization was comprehended simultaneously. Our results imply psychological stress is influential on specific immune system, more essential for the comprehensive treatment against tumors.

  1. Serum Stability and Affinity Optimization of an M2 Macrophage-Targeting Peptide (M2pep)

    PubMed Central

    Ngambenjawong, Chayanon; Gustafson, Heather H.; Pineda, Julio M.; Kacherovsky, Nataly A.; Cieslewicz, Maryelise; Pun, Suzie H.

    2016-01-01

    Tumor associated macrophages (TAMs) are a major stromal component of the tumor microenvironment in several cancers. TAMs are a potential target for adjuvant cancer therapies due to their established roles in promoting proliferation of cancer cells, angiogenesis, and metastasis. We previously discovered an M2 macrophage-targeting peptide (M2pep) which was successfully used to target and deliver a pro-apoptotic KLA peptide to M2-like TAMs in a CT-26 colon carcinoma model. However, the effectiveness of in vivo TAM-targeting using M2pep is limited by its poor serum stability and low binding affinity. In this study, we synthesized M2pep derivatives with the goals of increasing serum stability and binding affinity. Serum stability evaluation of M2pepBiotin confirmed its rapid degradation attributed to exolytic cleavage from the N-terminus and endolytic cleavages at the W10/W11 and S16/K17 sites. N-terminal acetylation of M2pepBiotin protected the peptide against the exolytic degradation while W10w and K(17,18,19)k substitutions were able to effectively protect endolytic degradation at their respective cleavage sites. However, no tested amino acid changes at the W10 position resulted in both protease resistance at that site and retention of binding activity. Therefore, cyclization of M2pep was investigated. Cyclized M2pep better resisted serum degradation without compromising binding activity to M2 macrophages. During the serum stability optimization process, we also discovered that K9R and W10Y substitutions significantly enhanced binding affinity of M2pep. In an in vitro binding study of different M2pep analogs pre-incubated in mouse serum, cyclic M2pep with K9R and W10Y modifications (cyclic M2pep(RY)) retained the highest binding activity to M2 macrophages over time due to its improved serum stability. Finally, we evaluated the in vivo accumulation of sulfo-Cy5-labeled M2pep and cyclic M2pep(RY) in both the CT-26 and 4T1 breast carcinoma models. Cyclic M2pep

  2. HCV core protein inhibits polarization and activity of both M1 and M2 macrophages through the TLR2 signaling pathway

    PubMed Central

    Zhang, Qianqian; Wang, Yang; Zhai, Naicui; Song, Hongxiao; Li, Haijun; Yang, Yang; Li, Tianyang; Guo, Xiaolin; Chi, Baorong; Niu, Junqi; Crispe, Ian Nicholas; Su, Lishan; Tu, Zhengkun

    2016-01-01

    Hepatitis C virus (HCV) establishes persistent infection in most infected patients, and eventually causes chronic hepatitis, cirrhosis, and hepatocellular carcinoma in some patients. Monocytes and macrophages provide the first line of defense against pathogens, but their roles in HCV infection remains unclear. We have reported that HCV core protein (HCVc) manipulates human blood-derived dendritic cell development. In the present study, we tested whether HCVc affects human blood-derived monocyte differentiating into macrophages. Results showed that HCVc inhibits monocyte differentiation to either M1 or M2 macrophages through TLR2, associated with impaired STATs signaling pathway. Moreover, HCVc inhibits phagocytosis activity of M1 and M2 macrophages, M1 macrophage-induced autologous and allogeneic CD4+ T cell activation, but promotes M2 macrophage-induced autologous and allogeneic CD4+ T cell activation. In conclusion, HCVc inhibits monocyte-derived macrophage polarization via TLR2 signaling, leading to dysfunctions of both M1 and M2 macrophages in chronic HCV infected patients. This may contribute to the mechanism of HCV persistent infection, and suggest that blockade of HCVc might be a novel therapeutic approach to treating HCV infection. PMID:27786268

  3. Microbial metabolite butyrate facilitates M2 macrophage polarization and function.

    PubMed

    Ji, Jian; Shu, Dingming; Zheng, Mingzhu; Wang, Jie; Luo, Chenglong; Wang, Yan; Guo, Fuyou; Zou, Xian; Lv, Xiaohui; Li, Ying; Liu, Tianfei; Qu, Hao

    2016-04-20

    Metabolites from intestinal microbes modulate the mucosal immune system by regulating the polarization and expansion of T cells. Whether the microbial metabolites influence macrophage polarization, however, is poorly understood. Here, we show that the large bowel microbial fermentation product, butyrate, facilitates M2 macrophage polarization, in vitro and in vivo. The supernatant from butyrate-treated M2 macrophage increased the migration and enhanced the wound closure rate of MLE-12 cells. Butyrate attenuated intestinal inflammation in mice with dextran sulfate sodium (DSS)-induced colitis, with a significant increase in colonic expression of the M2 macrophage-associated protein, Arg1. M2 macrophage treated with butyrate, had increased activation of the H3K9/STAT6 signaling pathway, suggesting a mechanism for butyrate facilitated M2 macrophage polarization. Collectively, our study indicated that commensal microbe-derived butyrate is a novel activator of STAT6-mediated transcription through H3K9 acetylation driving M2 macrophage polarization, and delineated new insights into the immune interplay underlying inflammatory bowel disease.

  4. Adrenergic receptor β2 activation by stress promotes breast cancer progression through macrophages M2 polarization in tumor microenvironment

    PubMed Central

    Qin, Jun-fang; Jin, Feng-jiao; Li, Ning; Guan, Hai-tao; Lan, Lan; Ni, Hong; Wang, Yue

    2015-01-01

    Stress and its related hormones epinephrine (E) and norepinephrine (NE) play a crucial role in tumor progression. Macrophages in the tumor microenvironment (TME) polarized to M2 is also a vital pathway for tumor deterioration. Here, we explore the underlying role of macrophages in the effect of stress and E promoting breast cancer growth. It was found that the weight and volume of tumor in tumor bearing mice were increased, and dramatically accompanied with the rising E level after chronic stress using social isolation. What is most noteworthy, the number of M2 macrophages inside tumor was up-regulated with it. The effects of E treatment appear to be directly related to the change of M2 phenotype is reproduced in vitro. Moreover, E receptor ADRβ2 involved in E promoting M2 polarization was comprehended simultaneously. Our results imply psychological stress is influential on specific immune system, more essential for the comprehensive treatment against tumors. [BMB Reports 2015; 48(5): 295-300] PMID:25748171

  5. Tumor hypoxia enhances non-small cell lung cancer metastasis by selectively promoting macrophage M2 polarization through the activation of ERK signaling

    PubMed Central

    Ma, Shenglin; Dong, Rong; Meng, Wen; Ying, Meidan; Weng, Qinjie; Chen, Zibo; Ma, Jian; Fang, Qingxia; He, Qiaojun; Yang, Bo

    2014-01-01

    Hypoxia is a common phenomenon occurring in the majority of human tumors and has been proved to play an important role in tumor progression. However, it remains unclear that whether the action of hypoxia on macrophages is a main driving force of hypoxia-mediated aggressive tumor behaviors. In the present study, we observe that high density of M2 macrophages is associated with metastasis in adenocarcinoma Non-Small Cell Lung Cancer (NSCLC) patients. By applying the in vivo hypoxia model, the results suggest that intermittent hypoxia significantly promotes the metastasis of Lewis lung carcinoma (LLC), accompanied with more CD209+ macrophages infiltrated in primary tumor tissue. More intriguingly, by skewing macrophages polarization away from the M1- to a tumor-promoting M2-like phenotype, hypoxia and IL-6 cooperate to enhance the LLC metastasis both in vitro and in vivo. In addition, we also demonstrate that skewing of macrophage M2 polarization by hypoxia relies substantially on activation of ERK signaling. Collectively, these observations unveil a novel tumor hypoxia concept involving the macrophage phenotype shift and provide direct evidence for lung cancer intervention through modulating the phenotype of macrophages. PMID:25313135

  6. TNF Counterbalances the Emergence of M2 Tumor Macrophages.

    PubMed

    Kratochvill, Franz; Neale, Geoffrey; Haverkamp, Jessica M; Van de Velde, Lee-Ann; Smith, Amber M; Kawauchi, Daisuke; McEvoy, Justina; Roussel, Martine F; Dyer, Michael A; Qualls, Joseph E; Murray, Peter J

    2015-09-22

    Cancer can involve non-resolving, persistent inflammation where varying numbers of tumor-associated macrophages (TAMs) infiltrate and adopt different activation states between anti-tumor M1 and pro-tumor M2 phenotypes. Here, we resolve a cascade causing differential macrophage phenotypes in the tumor microenvironment. Reduction in TNF mRNA production or loss of type I TNF receptor signaling resulted in a striking pattern of enhanced M2 mRNA expression. M2 gene expression was driven in part by IL-13 from eosinophils co-recruited with inflammatory monocytes, a pathway that was suppressed by TNF. Our data define regulatory nodes within the tumor microenvironment that balance M1 and M2 populations. Our results show macrophage polarization in cancer is dynamic and dependent on the balance between TNF and IL-13, thus providing a strategy for manipulating TAMs. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  7. IFN-γ promotes muscle damage in the mdx mouse model of Duchenne muscular dystrophy by suppressing M2 macrophage activation and inhibiting muscle cell proliferation.

    PubMed

    Villalta, S Armando; Deng, Bo; Rinaldi, Chiara; Wehling-Henricks, Michelle; Tidball, James G

    2011-11-15

    Duchenne muscular dystrophy is a degenerative disorder that leads to death by the third decade of life. Previous investigations have shown that macrophages that invade dystrophic muscle are a heterogeneous population consisting of M1 and M2 macrophages that promote injury and repair, respectively. In the present investigation, we tested whether IFN-γ worsens the severity of mdx dystrophy by activating macrophages to a cytolytic M1 phenotype and by suppressing the activation of proregenerative macrophages to an M2 phenotype. IFN-γ is a strong inducer of the M1 phenotype and is elevated in mdx dystrophy. Contrary to our expectations, null mutation of IFN-γ caused no reduction of cytotoxicity of macrophages isolated from mdx muscle and did not reduce muscle fiber damage in vivo or improve gross motor function of mdx mice at the early, acute peak of pathology. In contrast, ablation of IFN-γ reduced muscle damage in vivo during the regenerative stage of the disease and increased activation of the M2 phenotype and improved motor function of mdx mice at that later stage of the disease. IFN-γ also inhibited muscle cell proliferation and differentiation in vitro, and IFN-γ mutation increased MyoD expression in mdx muscle in vivo, showing that IFN-γ can have direct effects on muscle cells that could impair repair. Taken together, the findings show that suppression of IFN-γ signaling in muscular dystrophy reduces muscle damage and improves motor performance by promoting the M2 macrophage phenotype and by direct actions on muscle cells.

  8. Higenamine promotes M2 macrophage activation and reduces Hmgb1 production through HO-1 induction in a murine model of spinal cord injury.

    PubMed

    Zhang, Zhenyu; Li, Mingchao; Wang, Yan; Wu, Jian; Li, Jiaping

    2014-12-01

    Spinal cord injury (SCI) is considered to be primarily associated with loss of motor function and leads to the activation of diverse cellular mechanisms in the central nervous system to attempt to repair the damaged spinal cord tissue. Higenamine (HG) (1-[(4-hydroxyphenyl) methyl]-1,2,3,4-tetrahydroisoquinoline-6,7-diol), an active ingredient of Aconiti Lateralis Radix Praeparata, has been traditionally used as a heart stimulant and anti-inflammatory agent in oriental countries. However, the function and related mechanism of HG on SCI have never been investigated. In our current study, HG treatment displayed increased myelin sparring and enhanced spinal cord repair process. The numbers of CD4(+) T cells, CD8(+) T cells, Ly6G(+) neutrophils and CD11b(+) macrophages were all significantly lower in the HG-treated group than that in the control group after SCI. HG administration increased the expression of IL-4 and IL-10 and promoted M2 macrophage activation. Significantly reduced Hmgb1 expression was also observed in HG-treated mice with SCI. Furthermore, HG treatment promoted HO-1 production. The increased number of M2 macrophages, decreased expression of Hmgb1 and promoted locomotor recovery induced by HG were all reversed with additional HO-1 inhibitor treatment. In conclusion, HG promotes M2 macrophage activation and reduces Hmgb1 expression dependent on HO-1 induction and then promotes locomotor function after SCI.

  9. Modulation of macrophage activity by aflatoxins B1 and B2 and their metabolites aflatoxins M1 and M2.

    PubMed

    Bianco, G; Russo, R; Marzocco, S; Velotto, S; Autore, G; Severino, L

    2012-05-01

    Aflatoxins are natural contaminants frequently found both in food and feed. Many of them exert immunomodulatory properties in mammals; therefore, the aim of the current study was to investigate immune-effects of AFB1, AFB2, AFM1 and AFM2, alone and differently combined, in J774A.1 murine macrophages. MTT assay showed that AFB1, alone and combined with AFB2, possess antiproliferative activity only at the highest concentration; such effect was not shown by their hydroxylated metabolites, AFM1 and AFM2, respectively. However, the immunotoxic effects of the aflatoxins evaluated in the current study may be due to the inhibition of production of active oxygen metabolites such as NO. Cytofluorimetric assay in macrophages exposed to aflatoxins (10-100 μM) revealed that their cytoxicity is not related to apoptotic pathways. Nevertheless, a significant increase of the S phase cell population accompanied by a decrease in G0/G1 phase cell population was observed after AFB1 treatment. In conclusion, the results of the current study suggest that aflatoxins could compromise the macrophages functions; in particular, co-exposure to AFB1, AFB2, AFM1 and AFM2 may exert interactions which can significantly affect immunoreactivity.

  10. Liposomal targeting of prednisolone phosphate to synovial lining macrophages during experimental arthritis inhibits M1 activation but does not favor M2 differentiation.

    PubMed

    Hofkens, Wouter; Schelbergen, Rik; Storm, Gert; van den Berg, Wim B; van Lent, Peter L

    2013-01-01

    To determine the effects of liposomal targeting of prednisolone phosphate (Lip-PLP) to synovial lining macrophages on M1 and M2 polarization in vitro and during experimental arthritis. Experimental arthritis (antigen and immune complex induced) was elicited in mice and prednisolone containing liposomes were given systemically. Synovium was investigated using microarray analysis, RT-PCR and histology. Bone-marrow macrophages were stimulated towards M1 using LPS and IFNγ before treatment by PLP-liposomes. M1 and M2 markers were determined using RT-PCR. Microarray analysis of biopsies of inflamed synovium during antigen induced arthritis (AIA) showed an increased M1 signature characterized by upregulation of IL-1β, IL-6 and FcγRI starting from day 1 and lasting up until day 7 after arthritis induction. The M2 signature remained low throughout the 7 day course of arthritis. Treatment of AIA with intravenously delivered Lip-PLP strongly suppressed joint swelling and synovial infiltration whereas colloidal gold containing liposomes exclusively targeted the macrophages within the inflamed synovial intima layer. In vitro studies showed that Lip-PLP phagocytosed by M1 macrophages resulted in a suppression of the M1 phenotype and induction of M2 markers (IL-10, TGF-β, IL-1RII, CD163, CD206 and Ym1). In vivo, Lip-PLP treatment strongly suppressed M1 markers (TNF-α, IL-1β, IL-6, IL-12p40, iNOS, FcγRI, Ciita and CD86) after local M1 activation of lining macrophages with LPS and IFN-γ and during experimental AIA and immune complex arthritis (ICA). In contrast, M2 markers were not significantly upregulated in antigen-induced arthritis and down regulated in immune complex arthritis. This study clearly shows that systemic treatment with PLP-liposomes selectively targets synovial lining macrophages and inhibits M1 activation. In contrast to in vitro findings, PLP-liposomes do not cause a shift of synovial lining macrophages towards M2.

  11. Liposomal Targeting of Prednisolone Phosphate to Synovial Lining Macrophages during Experimental Arthritis Inhibits M1 Activation but Does Not Favor M2 Differentiation

    PubMed Central

    Hofkens, Wouter; Schelbergen, Rik; Storm, Gert; van den Berg, Wim B.; van Lent, Peter L.

    2013-01-01

    Background To determine the effects of liposomal targeting of prednisolone phosphate (Lip-PLP) to synovial lining macrophages on M1 and M2 polarization in vitro and during experimental arthritis. Material and Methods Experimental arthritis (antigen and immune complex induced) was elicited in mice and prednisolone containing liposomes were given systemically. Synovium was investigated using microarray analysis, RT-PCR and histology. Bone–marrow macrophages were stimulated towards M1 using LPS and IFNγ before treatment by PLP-liposomes. M1 and M2 markers were determined using RT-PCR. Results Microarray analysis of biopsies of inflamed synovium during antigen induced arthritis (AIA) showed an increased M1 signature characterized by upregulation of IL-1β, IL-6 and FcγRI starting from day 1 and lasting up until day 7 after arthritis induction. The M2 signature remained low throughout the 7 day course of arthritis. Treatment of AIA with intravenously delivered Lip-PLP strongly suppressed joint swelling and synovial infiltration whereas colloidal gold containing liposomes exclusively targeted the macrophages within the inflamed synovial intima layer. In vitro studies showed that Lip-PLP phagocytosed by M1 macrophages resulted in a suppression of the M1 phenotype and induction of M2 markers (IL-10, TGF-β, IL-1RII, CD163, CD206 and Ym1). In vivo, Lip-PLP treatment strongly suppressed M1 markers (TNF-α, IL-1β, IL-6, IL-12p40, iNOS, FcγRI, Ciita and CD86) after local M1 activation of lining macrophages with LPS and IFN-γ and during experimental AIA and immune complex arthritis (ICA). In contrast, M2 markers were not significantly upregulated in antigen-induced arthritis and down regulated in immune complex arthritis. Conclusion This study clearly shows that systemic treatment with PLP-liposomes selectively targets synovial lining macrophages and inhibits M1 activation. In contrast to in vitro findings, PLP-liposomes do not cause a shift of synovial lining

  12. Cervical Cancer Cell Supernatants Induce a Phenotypic Switch from U937-Derived Macrophage-Activated M1 State into M2-Like Suppressor Phenotype with Change in Toll-Like Receptor Profile

    PubMed Central

    Sánchez-Reyes, Karina; Bravo-Cuellar, Alejandro; Hernández-Flores, Georgina; Lerma-Díaz, José Manuel; Jave-Suárez, Luis Felipe; Gómez-Lomelí, Paulina; de Celis, Ruth; Aguilar-Lemarroy, Adriana; Domínguez-Rodríguez, Jorge Ramiro; Ortiz-Lazareno, Pablo Cesar

    2014-01-01

    Cervical cancer (CC) is the second most common cancer among women worldwide. Infection with human papillomavirus (HPV) is the main risk factor for developing CC. Macrophages are important immune effector cells; they can be differentiated into two phenotypes, identified as M1 (classically activated) and M2 (alternatively activated). Macrophage polarization exerts profound effects on the Toll-like receptor (TLR) profile. In this study, we evaluated whether the supernatant of human CC cells HeLa, SiHa, and C-33A induces a shift of M1 macrophage toward M2 macrophage in U937-derived macrophages. Results. The results showed that soluble factors secreted by CC cells induce a change in the immunophenotype of macrophages from macrophage M1 into macrophage M2. U937-derived macrophages M1 released proinflammatory cytokines and nitric oxide; however, when these cells were treated with the supernatant of CC cell lines, we observed a turnover of M1 toward M2. These cells increased CD163 and IL-10 expression. The expression of TLR-3, -7, and -9 is increased when the macrophages were treated with the supernatant of CC cells. Conclusions. Our result strongly suggests that CC cells may, through the secretion of soluble factors, induce a change of immunophenotype M1 into M2 macrophages. PMID:25309919

  13. Cervical cancer cell supernatants induce a phenotypic switch from U937-derived macrophage-activated M1 state into M2-like suppressor phenotype with change in Toll-like receptor profile.

    PubMed

    Sánchez-Reyes, Karina; Bravo-Cuellar, Alejandro; Hernández-Flores, Georgina; Lerma-Díaz, José Manuel; Jave-Suárez, Luis Felipe; Gómez-Lomelí, Paulina; de Celis, Ruth; Aguilar-Lemarroy, Adriana; Domínguez-Rodríguez, Jorge Ramiro; Ortiz-Lazareno, Pablo Cesar

    2014-01-01

    Cervical cancer (CC) is the second most common cancer among women worldwide. Infection with human papillomavirus (HPV) is the main risk factor for developing CC. Macrophages are important immune effector cells; they can be differentiated into two phenotypes, identified as M1 (classically activated) and M2 (alternatively activated). Macrophage polarization exerts profound effects on the Toll-like receptor (TLR) profile. In this study, we evaluated whether the supernatant of human CC cells HeLa, SiHa, and C-33A induces a shift of M1 macrophage toward M2 macrophage in U937-derived macrophages. The results showed that soluble factors secreted by CC cells induce a change in the immunophenotype of macrophages from macrophage M1 into macrophage M2. U937-derived macrophages M1 released proinflammatory cytokines and nitric oxide; however, when these cells were treated with the supernatant of CC cell lines, we observed a turnover of M1 toward M2. These cells increased CD163 and IL-10 expression. The expression of TLR-3, -7, and -9 is increased when the macrophages were treated with the supernatant of CC cells. Our result strongly suggests that CC cells may, through the secretion of soluble factors, induce a change of immunophenotype M1 into M2 macrophages.

  14. Novel Markers to Delineate Murine M1 and M2 Macrophages

    PubMed Central

    Jablonski, Kyle A.; Amici, Stephanie A.; Webb, Lindsay M.; Ruiz-Rosado, Juan de Dios; Popovich, Phillip G.; Partida-Sanchez, Santiago; Guerau-de-Arellano, Mireia

    2015-01-01

    Classically (M1) and alternatively activated (M2) macrophages exhibit distinct phenotypes and functions. It has been difficult to dissect macrophage phenotypes in vivo, where a spectrum of macrophage phenotypes exists, and also in vitro, where low or non-selective M2 marker protein expression is observed. To provide a foundation for the complexity of in vivo macrophage phenotypes, we performed a comprehensive analysis of the transcriptional signature of murine M0, M1 and M2 macrophages and identified genes common or exclusive to either subset. We validated by real-time PCR an M1-exclusive pattern of expression for CD38, G-protein coupled receptor 18 (Gpr18) and Formyl peptide receptor 2 (Fpr2) whereas Early growth response protein 2 (Egr2) and c-Myc were M2-exclusive. We further confirmed these data by flow cytometry and show that M1 and M2 macrophages can be distinguished by their relative expression of CD38 and Egr2. Egr2 labeled more M2 macrophages (~70%) than the canonical M2 macrophage marker Arginase-1, which labels 24% of M2 macrophages. Conversely, CD38 labeled most (71%) in vitro M1 macrophages. In vivo, a similar CD38+ population greatly increased after LPS exposure. Overall, this work defines exclusive and common M1 and M2 signatures and provides novel and improved tools to distinguish M1 and M2 murine macrophages. PMID:26699615

  15. Heme oxygenase-1 and anti-inflammatory M2 macrophages.

    PubMed

    Naito, Yuji; Takagi, Tomohisa; Higashimura, Yasuki

    2014-12-15

    Heme oxygenase-1 (HO-1) catalyzes the first and rate-limiting enzymatic step of heme degradation and produces carbon monoxide, free iron, and biliverdin. HO-1, a stress-inducible protein, is induced by various oxidative and inflammatory signals. Consequently, HO-1 expression has been regarded as an adaptive cellular response against inflammatory response and oxidative injury. Although several transcriptional factors and signaling cascades are involved in HO-1 regulation, the two main pathways of Nrf2/Bach1 system and IL-10/HO-1 axis exist in monocyte/macrophage. Macrophages are broadly divisible into two groups: pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages. More recently, several novel macrophage subsets have been identified including Mhem, Mox, and M4 macrophages. Of these, M2 macrophages, Mhem, and Mox are HO-1 highly expressing macrophages. HO-1 has been recognized as having major immunomodulatory and anti-inflammatory properties, which have been demonstrated in HO-1 deficient mice and human cases of genetic HO-1 deficiency. However, the mechanism underlying the immunomodulatory actions of HO-1 remains poorly defined. This review specifically addresses macrophage polarization. The present current evidence indicates that HO-1 induction mediated by multiple pathways can drive the phenotypic shift to M2 macrophages and suggests that HO-1 induction in macrophages is a potential therapeutic approach to immunomodulation in widely diverse human diseases.

  16. M2A and M2C Macrophage Subsets Ameliorate Inflammation and Fibroproliferation in Acute Lung Injury Through Interleukin 10 Pathway.

    PubMed

    Tang, Lunxian; Zhang, Hua; Wang, Chunmei; Li, Hongqiang; Zhang, Qian; Bai, Jianwen

    2016-12-09

    The role of M2 macrophages in the resolution and fibroproliferation of acute lung injury (ALI) is poorly understood. In this study, we investigated the effects of two M2 macrophage subtypes, M2a induced by interleukin (IL)-4/IL-13 and M2c induced by IL-10/transforming growth factor (TGF)-β, on the pathogenesis of ALI. M2a and M2c were adoptively transferred into LPS-induced ALI mice model. Data showed that Vybrant-labeled macrophages appeared in the lungs of ALI mice. Subsequently, we observed that both subsets significantly reduced lung inflammation and injury including a reduction of neutrophil influx into the lung and an augmentation of apoptosis. Interestingly, M2c macrophages more effectively suppressed indices of lung injury than M2a macrophages. M2c macrophages were also more effective than M2a in reduction of lung fibrosis. In addition, we found that M2c but not M2a macrophages increased IL-10 level in lung tissues of the recipient ALI mice partially mediated by activating the JAK1/STAT3/SOCS3 signaling pathway. After blocking IL-10, these superior effects of M2c over M2a were abolished. These data imply that M2c are more potent than M2a macrophages in protecting against lung injury and subsequent fibrosis due to their ability to produce IL-10. Therefore, reprogramming macrophages to M2c subset may be a novel treatment modality with transitional potential.

  17. Transcriptome analysis of IL-10-stimulated (M2c) macrophages by next-generation sequencing.

    PubMed

    Lurier, Emily B; Dalton, Donald; Dampier, Will; Raman, Pichai; Nassiri, Sina; Ferraro, Nicole M; Rajagopalan, Ramakrishan; Sarmady, Mahdi; Spiller, Kara L

    2017-02-20

    Alternatively activated "M2" macrophages are believed to function during late stages of wound healing, behaving in an anti-inflammatory manner to mediate the resolution of the pro-inflammatory response caused by "M1" macrophages. However, the differences between two main subtypes of M2 macrophages, namely interleukin-4 (IL-4)-stimulated "M2a" macrophages and IL-10-stimulated "M2c" macrophages, are not well understood. M2a macrophages are characterized by their ability to inhibit inflammation and contribute to the stabilization of angiogenesis. However, the role and temporal profile of M2c macrophages in wound healing are not known. Therefore, we performed next generation sequencing (RNA-seq) to identify biological functions and gene expression signatures of macrophages polarized in vitro with IL-10 to the M2c phenotype in comparison to M1 and M2a macrophages and an unactivated control (M0). We then explored the expression of these gene signatures in a publicly available data set of human wound healing. RNA-seq analysis showed that hundreds of genes were upregulated in M2c macrophages compared to the M0 control, with thousands of alternative splicing events. Following validation by Nanostring, 39 genes were found to be upregulated by M2c macrophages compared to the M0 control, and 17 genes were significantly upregulated relative to the M0, M1, and M2a phenotypes (using an adjusted p-value cutoff of 0.05 and fold change cutoff of 1.5). Many of the identified M2c-specific genes are associated with angiogenesis, matrix remodeling, and phagocytosis, including CD163, MMP8, TIMP1, VCAN, SERPINA1, MARCO, PLOD2, PCOCLE2 and F5. Analysis of the macrophage-conditioned media for secretion of matrix-remodeling proteins showed that M2c macrophages secreted higher levels of MMP7, MMP8, and TIMP1 compared to the other phenotypes. Interestingly, temporal gene expression analysis of a publicly available microarray data set of human wound healing showed that M2c-related genes were

  18. The chemotaxis of M1 and M2 macrophages is regulated by different chemokines.

    PubMed

    Xuan, Wenjuan; Qu, Qing; Zheng, Biao; Xiong, Sidong; Fan, Guo-Huang

    2015-01-01

    The homing of proinflammatory (M1) and the "alternatively activated" anti-inflammatory (M2) macrophages plays a different role in the process of inflammation. Chemokines are the major mediators of macrophage chemotaxis, but how they differentially regulate M1 and M2 macrophages remains largely unclear. In the present study, we attempted to screen chemokines that differentially induce chemotaxis of M1 and M2 macrophages and to explore the underlying mechanism. Among the 41 chemokines that specifically bind to 20 chemokine receptors, CCL19, CCL21, CCL24, CCL25, CXCL8, CXCL10, and XCL2 specifically induced M1 macrophage chemotaxis, whereas CCL7 induced chemotaxis of both M1 and M2 macrophages. Whereas the differential effects of these chemokines on M1/M2 macrophage chemotaxis could be attributable to the predominant expression of their cognate receptors on the macrophage subsets, CCR7, the receptor for CCL19/CCL21, appeared to be an exception. Immunoblot analysis indicated an equivalent level of CCR7 in the whole cell lysate of M1 and M2 macrophages, but CCL19 and CCL21 only induced M1 macrophage chemotaxis. Both immunoblot and confocal microscopy analyses demonstrated that CCR7 was predominantly expressed on the cell surface of M1 but in the cytosol of M2 macrophages before ligand stimulation. As a result, CCL19 or CCL21 induced activation of both MEK1-ERK1/2 and PI3K-AKT cascades in M1 but not in M2 macrophages. Intriguingly, CCL19/CCL21-mediated M1 macrophage chemotaxis was blocked by specific inhibition of PI3K rather than MEK1. Together, these findings suggest that recruitment of M1 and M2 macrophages is fine tuned by different chemokines with the involvement of specific signaling pathways. © Society for Leukocyte Biology.

  19. Unprimed, M1 and M2 Macrophages Differentially Interact with Porphyromonas gingivalis

    PubMed Central

    Lenzo, Jason C.; Fong, Shao B.; Reynolds, Eric C.

    2016-01-01

    Porphyromonas gingivalis is a keystone pathogen in the development of chronic periodontitis. Tissue macrophages are amongst the first immune cells to respond to bacteria and depending on the cytokine profile at the infection site, macrophages are primed to react to infection in different ways. Priming of naive macrophages with IFN-γ produces a classical pro-inflammatory, antibacterial M1 macrophage after TLR ligation, whereas priming with IL-4 induces an anti-inflammatory tissue-repair M2 phenotype. Previous work has shown that M1 are preferentially generated in gingival tissue following infection with P. gingivalis. However, few studies have investigated the interactions of macrophage subsets with P. gingivalis cells. The aim of this study was to determine the ability of naive, M1 and M2 macrophages to phagocytose P. gingivalis and investigate how this interaction affects both the bacterial cell and the macrophage. M1 and M2 macrophages were both found to have enhanced phagocytic capacity compared with that of naive macrophages, however only the naive and M1 macrophages were able to produce a respiratory burst in order to clear the bacteria from the phagosome. P. gingivalis was found to persist in naive and M2, but not M1 macrophages for 24 hours. Phagocytosis of P. gingivalis also induced high levels of TNF-α, IL-12 and iNOS in M1 macrophages, but not in naive or M2 macrophages. Furthermore, infection of macrophages with P. gingivalis at high bacteria to macrophage ratios, while inducing an inflammatory response, was also found to be deleterious to macrophage longevity, with high levels of apoptotic cell death found in macrophages after infection. The activation of M1 macrophages observed in this study may contribute to the initiation and maintenance of a pro-inflammatory state during chronic periodontitis. PMID:27383471

  20. Adrenomedullin 2 Enhances Beiging in White Adipose Tissue Directly in an Adipocyte-autonomous Manner and Indirectly through Activation of M2 Macrophages.

    PubMed

    Lv, Ying; Zhang, Song-Yang; Liang, Xianyi; Zhang, Heng; Xu, Zhi; Liu, Bo; Xu, Ming-Jiang; Jiang, Changtao; Shang, Jin; Wang, Xian

    2016-11-04

    Adrenomedullin 2 (ADM2) is an endogenous bioactive peptide belonging to the calcitonin gene-related peptide family. Our previous studies showed that overexpression of ADM2 in mice reduced obesity and insulin resistance by increasing thermogenesis in brown adipose tissue. However, the effects of ADM2 in another type of thermogenic adipocyte, beige adipocytes, remain to be understood. The plasma ADM2 levels were inversely correlated with obesity in humans, and adipo-ADM2-transgenic (tg) mice displayed resistance to high-fat diet-induced obesity with increased energy expenditure. Beiging of subcutaneous white adipose tissues (WAT) was more noticeably induced in high-fat diet-fed transgenic mice with adipocyte-ADM2 overexpression (adipo-ADM2-tg mice) than in WT animals. ADM2 treatment in primary rat subcutaneous adipocytes induced beiging with up-regulation of UCP1 and beiging-related marker genes and increased mitochondrial uncoupling respiration, which was mainly mediated by activation of the calcitonin receptor-like receptor (CRLR)·receptor activity-modifying protein 1 (RAMP1) complex and PKA and p38 MAPK signaling pathways. Importantly, this adipocyte-autonomous beiging effect by ADM2 was translatable to human primary adipocytes. In addition, M2 macrophage activation also contributed to the beiging effects of ADM2 through catecholamine secretion. Therefore, our study reveals that ADM2 enhances subcutaneous WAT beiging via a direct effect by activating the CRLR·RAMP1-cAMP/PKA and p38 MAPK pathways in white adipocytes and via an indirect effect by stimulating alternative M2 polarization in macrophages. Through both mechanisms, beiging of WAT by ADM2 results in increased energy expenditure and reduced obesity, suggesting ADM2 as a novel anti-obesity target. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Anatomy of a Discovery: M1 and M2 Macrophages

    PubMed Central

    Mills, Charles Dudley

    2015-01-01

    M1 and M2 macrophage-type responses kill or repair in vivo. The unique ability of macrophages to make these polar opposite type of responses provides primary host protection and maintains tissue homeostasis throughout the animal kingdom. In humans and other higher animals, M1 and M2-type macrophage responses also initiate and direct T cells/adaptive immunity to provide additional protection such as Th1 (cytotoxic) or Th2 (antibody-mediated) type responses. Hence, macrophages were renamed M1 and M2 to indicate the central role of macrophages/innate immunity in immune systems. These findings indicate that the long held notion that adaptive immunity controls innate immunity was backward: a sea change in understanding how immune responses occur. The clinical impact of M1/kill and M2/repair responses is immense playing pivotal roles in curing (or causing) many diseases including infections, cancer, autoimmunity, and atherosclerosis. How M1/M2 came to be is an interesting story that, like life, involved Direction, Determination, Discouragement, and Discovery. PMID:25999950

  2. M2 polarization of macrophages facilitates arsenic-induced cell transformation of lung epithelial cells.

    PubMed

    Cui, Jiajun; Xu, Wenhua; Chen, Jian; Li, Hui; Dai, Lu; Frank, Jacqueline A; Peng, Shaojun; Wang, Siying; Chen, Gang

    2017-03-28

    The alterations in microenvironment upon chronic arsenic exposure may contribute to arsenic-induced lung carcinogenesis. Immune cells, such as macrophages, play an important role in mediating the microenvironment in the lungs. Macrophages carry out their functions after activation. There are two activation status for macrophages: classical (M1) or alternative (M2); the latter is associated with tumorigenesis. Our previous work showed that long-term arsenic exposure induces transformation of lung epithelial cells. However, the crosstalk between epithelial cells and macrophages upon arsenic exposure has not been investigated. In this study, using a co-culture system in which human lung epithelial cells are cultured with macrophages, we determined that long-term arsenic exposure polarizes macrophages towards M2 status through ROS generation. Co-culture with epithelial cells further enhanced the polarization of macrophages as well as transformation of epithelial cells, while blocking macrophage M2 polarization decreased the transformation. In addition, macrophage M2 polarization decreased autophagy activity, which may account for increased cell transformation of epithelial cells with co-culture of macrophages.

  3. M2 polarization of macrophages facilitates arsenic-induced cell transformation of lung epithelial cells

    PubMed Central

    Li, Hui; Dai, Lu; Frank, Jacqueline A.; Peng, Shaojun; Wang, Siying; Chen, Gang

    2017-01-01

    The alterations in microenvironment upon chronic arsenic exposure may contribute to arsenic-induced lung carcinogenesis. Immune cells, such as macrophages, play an important role in mediating the microenvironment in the lungs. Macrophages carry out their functions after activation. There are two activation status for macrophages: classical (M1) or alternative (M2); the latter is associated with tumorigenesis. Our previous work showed that long-term arsenic exposure induces transformation of lung epithelial cells. However, the crosstalk between epithelial cells and macrophages upon arsenic exposure has not been investigated. In this study, using a co-culture system in which human lung epithelial cells are cultured with macrophages, we determined that long-term arsenic exposure polarizes macrophages towards M2 status through ROS generation. Co-culture with epithelial cells further enhanced the polarization of macrophages as well as transformation of epithelial cells, while blocking macrophage M2 polarization decreased the transformation. In addition, macrophage M2 polarization decreased autophagy activity, which may account for increased cell transformation of epithelial cells with co-culture of macrophages. PMID:28423485

  4. Superparamagnetic iron oxide nanoparticle uptake alters M2 macrophage phenotype, iron metabolism, migration and invasion.

    PubMed

    Rojas, José M; Sanz-Ortega, Laura; Mulens-Arias, Vladimir; Gutiérrez, Lucía; Pérez-Yagüe, Sonia; Barber, Domingo F

    2016-05-01

    Superparamagnetic iron oxide nanoparticles (SPIONs) have shown promise as contrast agents and nanocarriers for drug delivery. Their impact on M2-polarised macrophages has nonetheless not been well studied. Here we explored the effects of SPIONs coated with dimercaptosuccinic acid, aminopropyl silane or aminodextran in two M2 macrophage models (murine primary IL-4-activated bone marrow-derived macrophages and human M2-like differentiated THP-1 cells). All SPIONs were internalised and no cell toxicity was observed. SPION treatment produced reactive oxygen species and activated the extracellular signal-regulated kinase and AKT pathways. After 24-h SPION incubation, M2 macrophages switched their iron metabolism towards an iron-replete state. SPION treatment in both M2 macrophage models altered their M2 activation profiles, promoted IL-10 production, and stimulated protease-dependent invasion. These results highlight the need to evaluate the interactions between SPIONs and cells to take full advantage of the intrinsic properties of these nanoparticles in biological systems. Superparamagnetic iron oxide nanoparticles (SPIONs) have been used as an MRI contrast agent in many experimental studies. The authors here investigated the effects of these nanoparticles on M2 macrophages after cellular uptake. The findings of cell activation further enhanced our current knowledge on the interaction of SPIONS with macrophages. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Gut dysbiosis promotes M2 macrophage polarization and allergic airway inflammation via fungi-induced PGE₂.

    PubMed

    Kim, Yun-Gi; Udayanga, Kankanam Gamage Sanath; Totsuka, Naoya; Weinberg, Jason B; Núñez, Gabriel; Shibuya, Akira

    2014-01-15

    Although imbalances in gut microbiota composition, or "dysbiosis," are associated with many diseases, the effects of gut dysbiosis on host systemic physiology are less well characterized. We report that gut dysbiosis induced by antibiotic (Abx) treatment promotes allergic airway inflammation by shifting macrophage polarization in the lung toward the alternatively activated M2 phenotype. Adoptive transfer of alveolar macrophages derived from Abx-treated mice was sufficient to increase allergic airway inflammation. Abx treatment resulted in the overgrowth of a commensal fungal Candida species in the gut and increased plasma concentrations of prostaglandin E₂ (PGE₂), which induced M2 macrophage polarization in the lung. Suppression of PGE₂ synthesis by the cyclooxygenase inhibitors aspirin and celecoxib suppressed M2 macrophage polarization and decreased allergic airway inflammatory cell infiltration in Abx-treated mice. Thus, Abx treatment can cause overgrowth of particular fungal species in the gut and promote M2 macrophage activation at distant sites to influence systemic responses including allergic inflammation.

  6. Differential S1P Receptor Profiles on M1- and M2-Polarized Macrophages Affect Macrophage Cytokine Production and Migration

    PubMed Central

    Müller, Jan; von Bernstorff, Wolfram; Heidecke, Claus-Dieter

    2017-01-01

    Introduction. Macrophages are key players in complex biological processes. In response to environmental signals, macrophages undergo polarization towards a proinflammatory (M1) or anti-inflammatory (M2) phenotype. Sphingosine 1-phosphate (S1P) is a bioactive lysophospholipid that acts via 5 G-protein coupled receptors (S1P1–5) in order to influence a broad spectrum of biological processes. This study assesses S1P receptor expression on macrophages before and after M1 and M2 polarization and performs a comparative analysis of S1P signalling in the two activational states of macrophages. Methods. Bone marrow derived macrophages (BMDM) from C57 BL/6 mice were cultured under either M1- or M2-polarizing conditions. S1P-receptor expression was determined by quantitative RT-PCR. Influence of S1P on macrophage activation, migration, phagocytosis, and cytokine secretion was assessed in vitro. Results. All 5 S1P receptor subclasses were expressed in macrophages. Culture under both M1- and M2-polarizing conditions led to significant downregulation of S1P1. In contrast, M1-polarized macrophages significantly downregulated S1P4. The expression of the remaining three S1P receptors did not change. S1P increased expression of iNOS under M2-polarizing conditions. Furthermore, S1P induced chemotaxis in M1 macrophages and changed cytokine production in M2 macrophages. Phagocytosis was not affected by S1P-signalling. Discussion. The expression of different specific S1P receptor profiles may provide a possibility to selectively influence M1- or M2-polarized macrophages. PMID:28367448

  7. Tramadol differentially regulates M1 and M2 macrophages from human umbilical cord blood.

    PubMed

    Zhang, Jun; Chen, Liang; Sun, Yunyun; Li, Yuanhai

    2017-03-17

    Tramadol is an analgesic drug and relieves pain through activating μ-opioid receptors and inhibiting serotonin and noradrenaline reuptake. Emerging evidence shows that it also stimulates immune cells, including NK cells, splenocytes, and lymphocytes, and elevates IL-2 production. However, it remains unknown whether and how tramadol directly affects macrophages. To answer these questions, we collected human umbilical cord blood, isolated macrophages, and examined their responses to tramadol. Although tramadol did not alter resting macrophages and the antigen-presenting function in lipopolysaccharide-activated macrophages, it regulated M1 and M2 macrophages, which are, respectively, transformed by IFN-γ and IL-4. Interestingly, tramadol inhibits production and secretion of cytokines in M1 macrophages, but facilitates the production of inflammation-responding molecules, synthesized in M2 macrophages. We also found that STAT6 cascade pathway in M2 macrophages was significantly enhanced by tramadol. Therefore, this study reveals that tramadol regulates inflammation by inhibiting M1 macrophages (killing process), but promoting the function of M2 macrophages (healing process).

  8. Macrophage activation and polarization.

    PubMed

    Martinez, Fernando Oneissi; Sica, Antonio; Mantovani, Alberto; Locati, Massimo

    2008-01-01

    Macrophages are widely distributed immune system cells that play an indispensable role in homeostasis and defense. They can be phenotypically polarized by the microenvironment to mount specific functional programs. Polarized macrophages can be broadly classified in two main groups: classically activated macrophages (or M1), whose prototypical activating stimuli are IFNgamma and LPS, and alternatively activated macrophages (or M2), further subdivided in M2a (after exposure to IL-4 or IL-13), M2b (immune complexes in combination with IL-1beta or LPS) and M2c (IL-10, TGFbeta or glucocorticoids). M1 exhibit potent microbicidal properties and promote strong IL-12-mediated Th1 responses, whilst M2 support Th2-associated effector functions. Beyond infection M2 polarized macrophages play a role in resolution of inflammation through high endocytic clearance capacities and trophic factor synthesis, accompanied by reduced pro-inflammatory cytokine secretion. Similar functions are also exerted by tumor-associated macrophages (TAM), which also display an alternative-like activation phenotype and play a detrimental pro-tumoral role. Here we review the main functions of polarized macrophages and discuss the perspectives of this field.

  9. Structural environment built by AKAP12+ colon mesenchymal cells drives M2 macrophages during inflammation recovery

    PubMed Central

    Yang, Jun-Mo; Lee, Hye Shin; Seo, Ji Hae; Park, Ji-Hyeon; Gelman, Irwin H.; Lo, Eng H.; Kim, Kyu-Won

    2017-01-01

    Macrophages exhibit phenotypic plasticity, as they have the ability to switch their functional phenotypes during inflammation and recovery. Simultaneously, the mechanical environment actively changes. However, how these dynamic alterations affect the macrophage phenotype is unknown. Here, we observed that the extracellular matrix (ECM) constructed by AKAP12+ colon mesenchymal cells (CMCs) generated M2 macrophages by regulating their shape during recovery. Notably, rounded macrophages were present in the linear and loose ECM of inflamed colons and polarized to the M1 phenotype. In contrast, ramified macrophages emerged in the contracted ECM of recovering colons and mainly expressed M2 macrophage markers. These contracted structures were not observed in the inflamed colons of AKAP12 knockout (KO) mice. Consequently, the proportion of M2 macrophages in inflamed colons was lower in AKAP12 KO mice than in WT mice. In addition, clinical symptoms and histological damage were more severe in AKAP12 KO mice than in WT mice. In experimentally remodeled collagen gels, WT CMCs drove the formation of a more compacted structure than AKAP12 KO CMCs, which promoted the polarization of macrophages toward an M2 phenotype. These results demonstrated that tissue contraction during recovery provides macrophages with the physical cues that drive M2 polarization. PMID:28205544

  10. Structural environment built by AKAP12+ colon mesenchymal cells drives M2 macrophages during inflammation recovery.

    PubMed

    Yang, Jun-Mo; Lee, Hye Shin; Seo, Ji Hae; Park, Ji-Hyeon; Gelman, Irwin H; Lo, Eng H; Kim, Kyu-Won

    2017-02-16

    Macrophages exhibit phenotypic plasticity, as they have the ability to switch their functional phenotypes during inflammation and recovery. Simultaneously, the mechanical environment actively changes. However, how these dynamic alterations affect the macrophage phenotype is unknown. Here, we observed that the extracellular matrix (ECM) constructed by AKAP12+ colon mesenchymal cells (CMCs) generated M2 macrophages by regulating their shape during recovery. Notably, rounded macrophages were present in the linear and loose ECM of inflamed colons and polarized to the M1 phenotype. In contrast, ramified macrophages emerged in the contracted ECM of recovering colons and mainly expressed M2 macrophage markers. These contracted structures were not observed in the inflamed colons of AKAP12 knockout (KO) mice. Consequently, the proportion of M2 macrophages in inflamed colons was lower in AKAP12 KO mice than in WT mice. In addition, clinical symptoms and histological damage were more severe in AKAP12 KO mice than in WT mice. In experimentally remodeled collagen gels, WT CMCs drove the formation of a more compacted structure than AKAP12 KO CMCs, which promoted the polarization of macrophages toward an M2 phenotype. These results demonstrated that tissue contraction during recovery provides macrophages with the physical cues that drive M2 polarization.

  11. ICAM-1 suppresses tumor metastasis by inhibiting macrophage M2 polarization through blockade of efferocytosis

    PubMed Central

    Yang, M; Liu, J; Piao, C; Shao, J; Du, J

    2015-01-01

    Efficient clearance of apoptotic cells (efferocytosis) can profoundly influence tumor-specific immunity. Tumor-associated macrophages are M2-polarized macrophages that promote key processes in tumor progression. Efferocytosis stimulates M2 macrophage polarization and contributes to cancer metastasis, but the signaling mechanism underlying this process is unclear. Intercellular cell adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein member of the immunoglobulin superfamily, which has been implicated in mediating cell–cell interaction and outside-in cell signaling during the immune response. We report that ICAM-1 expression is inversely associated with macrophage infiltration and the metastasis index in human colon tumors by combining Oncomine database analysis and immunohistochemistry for ICAM-1. Using a colon cancer liver metastasis model in ICAM-1-deficient (ICAM-1−/−) mice and their wild-type littermates, we found that loss of ICAM-1 accelerated liver metastasis of colon carcinoma cells. Moreover, ICAM-1 deficiency increased M2 macrophage polarization during tumor progression. We further demonstrated that ICAM-1 deficiency in macrophages led to promotion of efferocytosis of apoptotic tumor cells through activation of the phosphatidylinositol 3 kinase/Akt signaling pathway. More importantly, coculture of ICAM-1−/− macrophages with apoptotic cancer cells resulted in an increase of M2-like macrophages, which was blocked by an efferocytosis inhibitor. Our findings demonstrate a novel role for ICAM-1 in suppressing M2 macrophage polarization via downregulation of efferocytosis in the tumor microenvironment, thereby inhibiting metastatic tumor progression. PMID:26068788

  12. Fra-1 protooncogene regulates IL-6 expression in macrophages and promotes the generation of M2d macrophages.

    PubMed

    Wang, Qingshan; Ni, Hong; Lan, Lan; Wei, Xiaoli; Xiang, Rong; Wang, Yue

    2010-06-01

    The tumor microenvironment (TME) plays a prominent role in the growth of tumor cells. As the major inflammatory component of the TME, M2d macrophages are educated by the TME such that they adopt an immunosuppressive role that promotes tumor metastasis and progression. Fra-1 forms activator protein-1 heterodimers with Jun partners and drives gene transcription. Fra-1 is thought to drastically induce tumorigenesis and progression. However, the functional role of Fra-1 in the generation of M2d macrophages is poorly understood to date. Here, we demonstrate that 4T1 mammary carcinoma cells, when co-cultured with RAW264.7 macrophage cells, skew the RAW264.7 macrophage cell differentiation into M2d macrophages. The 4T1 cells stimulate de novo overexpression of Fra-1 in RAW264.7 cells, and then Fra-1 binds to the interleukin 6 (IL-6) promoter to increase the production of the cytokine IL-6 in RAW264.7 cells. IL-6 acts in an autocrine fashion to skew RAW264.7 macrophage cell differentiation into M2d macrophages. These findings open new insights into how to reverse M2d macrophage-induced immune tolerance to improve the efficacy of immunotherapeutic approaches.

  13. M1 and M2 macrophages: the chicken and the egg of immunity.

    PubMed

    Mills, Charles D; Ley, Klaus

    2014-01-01

    The purpose of this perspective is to describe a critical advance in understanding how immune responses work. Macrophages are required for all animal life: 'Inhibit' type macrophages in all animals (called M1) can rapidly kill pathogens, and are thus the primary host defense, and 'Heal' type macrophages (M2) routinely repair and maintain tissue integrity. Macrophages perform these activities in all animals without T cells, and also in T cell-deficient vertebrates. Although adaptive immunity can amplify macrophage polarization, the long-held notion that macrophages need to be 'activated' or 'alternatively activated' by T cells is incorrect; indeed, immunology has had it backward. M1/M2-type macrophages necessarily direct T cells toward Th1- or Th2-like activities, respectively. That such macrophage-innate activities are the central directing element in immune responses is a dramatic change in understanding how immune systems operate. Most important, this revelation is opening up whole new approaches to immunotherapy. For example, many modern diseases, such as cancer and atherosclerosis, may not display 'foreign' antigens. However, there are clear imbalances in M1/M2-type responses. Correcting such innate imbalances can result in better health. Macrophages are the chicken and the egg of immunity.

  14. Discrete functions of M2a and M2c macrophage subsets determine their relative efficacy in treating chronic kidney disease.

    PubMed

    Lu, Junyu; Cao, Qi; Zheng, Dong; Sun, Yan; Wang, Changqi; Yu, Xiao; Wang, Ya; Lee, Vincent W S; Zheng, Guoping; Tan, Thian K; Wang, Xin; Alexander, Stephen I; Harris, David C H; Wang, Yiping

    2013-10-01

    Two types of alternatively activated macrophages, M(2a) induced by IL-4/IL-13 and M(2c) by IL-10/TGF-β, exhibit anti-inflammatory functions in vitro and protect against renal injury in vivo. Since their relative therapeutic efficacy is unclear, we compared the effects of these two macrophage subsets in murine adriamycin nephrosis. Both subsets significantly reduced renal inflammation and renal injury; however, M(2c) macrophages more effectively reduced glomerulosclerosis, tubular atrophy, interstitial expansion, and proteinuria than M(2a) macrophages. The M(2c) macrophages were also more effective than M(2a) in reduction of macrophage and CD4(+) T-cell infiltration in kidney. Moreover, nephrotic mice treated with M(2c) had a greater reduction in renal fibrosis than those treated with M(2a). M(2c) but not M(2a) macrophages induced regulatory T cells (Tregs) from CD4(+)CD25(-) T cells in vitro, and increased Treg numbers in local draining lymph nodes of nephrotic mice. To determine whether the greater protection with M(2c) was due to their capability to induce Tregs, the Tregs were depleted by PC61 antibody in nephrotic mice treated with M(2a) or M(2c). Treg depletion diminished the superior effects of M(2c) compared to M(2a) in protection against renal injury, inflammatory infiltrates, and renal fibrosis. Thus, M(2c) are more potent than M(2a) macrophages in protecting against renal injury due to their ability to induce Tregs.

  15. RBP-J is required for M2 macrophage polarization in response to chitin and mediates expression of a subset of M2 genes.

    PubMed

    Foldi, Julia; Shang, Yingli; Zhao, Baohong; Ivashkiv, Lionel B; Hu, Xiaoyu

    2016-03-01

    Development of alternatively activated (M2) macrophage phenotypes is a complex process that is coordinately regulated by a plethora of pathways and factors. Here, we report that RBP-J, a DNA-binding protein that integrates signals from multiple pathways including the Notch pathway, is critically involved in polarization of M2 macrophages. Mice deficient in RBP-J in the myeloid compartment exhibited impaired M2 phenotypes in vivo in a chitin-induced model of M2 polarization. Consistent with the in vivo findings, M2 polarization was partially compromised in vitro in Rbpj-deficient macrophages as demonstrated by reduced expression of a subset of M2 effector molecules including arginase 1. Functionally, myeloid Rbpj deficiency impaired M2 effector functions including recruitment of eosinophils and suppression of T cell proliferation. Collectively, we have identified RBP-J as an essential regulator of differentiation and function of alternatively activated macrophages.

  16. M1 and M2 Macrophages: The Chicken and the Egg of Immunity

    PubMed Central

    Mills, Charles D.; Ley, Klaus

    2015-01-01

    The purpose of this perspective is to describe a critical advance in understanding how immune responses work. Macrophages are required for all animal life: ‘Inhibit’ type macrophages in all animals (called M1) can rapidly kill pathogens, and are thus the primary host defense, and ‘Heal’ type macrophages (M2) routinely repair and maintain tissue integrity. Macrophages perform these activities in all animals without T cells, and also in T cell-deficient vertebrates. Although adaptive immunity can amplify macrophage polarization, the long-held notion that macrophages need to be ‘activated’ or ‘alternatively activated’ by T cells is incorrect; indeed, immunology has had it backward. M1/M2-type macrophages necessarily direct T cells toward Th1- or Th2-like activities, respectively. That such macrophage-innate activities are the central directing element in immune responses is a dramatic change in understanding how immune systems operate. Most important, this revelation is opening up whole new approaches to immunotherapy. For example, many modern diseases, such as cancer and atherosclerosis, may not display ‘foreign’ antigens. However, there are clear imbalances in M1/M2-type responses. Correcting such innate imbalances can result in better health. Macrophages are the chicken and the egg of immunity. PMID:25138714

  17. M2 macrophages induce EMT through the TGF-β/Smad2 signaling pathway.

    PubMed

    Zhu, Liangying; Fu, Xiao; Chen, Xiang; Han, Xiaodong; Dong, Ping

    2017-09-01

    IPF is characterized by fibroblast accumulation, collagen deposition, and ECM remodeling, with myofibroblasts believed to be the effector cell type. Myofibroblasts develop due to EMT of lung alveolar epithelial cells, which can be induced by TGF-β. M2 macrophages, a macrophage subpopulation, secrete large amounts of TGF-β. To clarify the relationship between IPF, EMT, TGF-β, and M2 macrophages, a bleomycin-induced pulmonary fibrosis mouse model was used. Seventeen days after mice were treated with bleomycin, the successful establishment of a pulmonary fibrosis model was confirmed by HE stain and Masson's trichrome stain. We found evidence in support of EMT, such as elevated protein levels of α-SMA in lung tissue and decreased levels of E-cadherin and CK-18. Additionally, increased TGF-β levels and TGF-β/Smad2 signaling activation was observed. Macrophages were recruited to pulmonary alveoli. Alveolar macrophages were phenotyped and identified as M2 macrophages, with up-regulated CD206 on the cell surfaces. For in vitro studies, we treated RAW 264.7 cells with IL-4 for 24 h, and the cells were then utilized as M2 macrophages. TGF-β levels increased significantly in the culture supernatant. Forty-eight hours after lung epithelial cells (MLE-12) were co-cultured with the M2 macrophages, the expression of α-SMA increased, and E-cadherin and CK-18 decreased. When a TGF-β receptor inhibitor, LY2109761 was used, the EMT induced by M2 macrophages was blocked. In conclusion, we demonstrated that M2 macrophages induce EMT through the TGF-β/Smad2 signaling pathway. © 2017 International Federation for Cell Biology.

  18. Adenosine augments IL-10-induced STAT3 signaling in M2c macrophages.

    PubMed

    Koscsó, Balázs; Csóka, Balázs; Kókai, Endre; Németh, Zoltán H; Pacher, Pál; Virág, László; Leibovich, S Joseph; Haskó, György

    2013-12-01

    The alternatively activated macrophage phenotype induced by IL-10 is called M2c. Adenosine is an endogenous purine nucleoside that accumulates in the extracellular space in response to metabolic disturbances, hypoxia, inflammation, physical damage, or apoptosis. As adenosine is known to regulate classically activated M1 and IL4- and IL-13-activated M2a macrophages, the goal of the present study was to explore its effects on M2c macrophages. We found that adenosine augmented the IL-10-induced expression of TIMP-1 and arginase-1 by the mouse macrophage cell line RAW 264.7 and by mouse BMDMs. The effects of AR stimulation on IL-10-induced TIMP-1 or arginase-1 expression were lacking in A2BAR KO macrophages. The role of A2BAR on TIMP-1 production of RAW 264.7 cells was confirmed with specific agonist BAY606583 and antagonist PSB0788. AR stimulation augmented IL-10-induced STAT3 phosphorylation in macrophages, and pharmacological inhibition or silencing of STAT3 using siRNA reduced the stimulatory effect of AR stimulation on TIMP-1 production. In contrast to its stimulatory effect on IL-10-induced STAT3 activation, adenosine inhibited IL-6-induced STAT3 phosphorylation and SAA3 expression. In conclusion, adenosine enhances IL-10-induced STAT3 signaling and M2c macrophage activation.

  19. From Monocytes to M1/M2 Macrophages: Phenotypical vs. Functional Differentiation

    PubMed Central

    Italiani, Paola; Boraschi, Diana

    2014-01-01

    Studies on monocyte and macrophage biology and differentiation have revealed the pleiotropic activities of these cells. Macrophages are tissue sentinels that maintain tissue integrity by eliminating/repairing damaged cells and matrices. In this M2-like mode, they can also promote tumor growth. Conversely, M1-like macrophages are key effector cells for the elimination of pathogens, virally infected, and cancer cells. Macrophage differentiation from monocytes occurs in the tissue in concomitance with the acquisition of a functional phenotype that depends on microenvironmental signals, thereby accounting for the many and apparently opposed macrophage functions. Many questions arise. When monocytes differentiate into macrophages in a tissue (concomitantly adopting a specific functional program, M1 or M2), do they all die during the inflammatory reaction, or do some of them survive? Do those that survive become quiescent tissue macrophages, able to react as naïve cells to a new challenge? Or, do monocyte-derived tissue macrophages conserve a “memory” of their past inflammatory activation? This review will address some of these important questions under the general framework of the role of monocytes and macrophages in the initiation, development, resolution, and chronicization of inflammation. PMID:25368618

  20. Chlorogenic acid inhibits glioblastoma growth through repolarizating macrophage from M2 to M1 phenotype

    PubMed Central

    Xue, Nina; Zhou, Qin; Ji, Ming; Jin, Jing; Lai, Fangfang; Chen, Ju; Zhang, Mengtian; Jia, Jing; Yang, Huarong; Zhang, Jie; Li, Wenbin; Jiang, Jiandong; Chen, Xiaoguang

    2017-01-01

    Glioblastoma is an aggressive tumor that is associated with distinctive infiltrating microglia/macrophages populations. Previous studies demonstrated that chlorogenic acid (5-caffeoylquinic acid, CHA), a phenolic compound with low molecular weight, has an anti-tumor effect in multiple malignant tumors. In the present study, we focused on the macrophage polarization to investigate the molecular mechanisms behind the anti-glioma response of CHA in vitro and in vivo. We found that CHA treatment increased the expression of M1 markers induced by LPS/IFNγ, including iNOS, MHC II (I-A/I-E subregions) and CD11c, and reduced the expression of M2 markers Arg and CD206 induced by IL-4, resulting in promoting the production of apoptotic-like cancer cells and inhibiting the growth of tumor cells by co-culture experiments. The activations of STAT1 and STAT6, which are two crucial signaling events in M1 and M2-polarization, were significantly promoted and suppressed by CHA in macrophages, respectively. Furthermore, In G422 xenograft mice, CHA increased the proportion of CD11c-positive M1 macrophages and decreased the distribution of CD206-positive M2 macrophages in tumor tissue, consistent with the reduction of tumor weight observed in CHA-treated mice. Overall these findings indicated CHA as a potential therapeutic approach to reduce glioma growth through promoting M1-polarized macrophage and inhibiting M2 phenotypic macrophage. PMID:28045028

  1. M1/M2 Macrophage Polarity in Normal and Complicated Pregnancy

    PubMed Central

    Brown, Mary B.; von Chamier, Maria; Allam, Ayman B.; Reyes, Leticia

    2014-01-01

    Tissue macrophages play an important role in all stages of pregnancy, including uterine stromal remodeling (decidualization) before embryo implantation, parturition, and post-partum uterine involution. The activation state and function of utero-placental macrophages are largely dependent on the local tissue microenvironment. Thus, macrophages are involved in a variety of activities such as regulation of immune cell activities, placental cell invasion, angiogenesis, and tissue remodeling. Disruption of the uterine microenvironment, particularly during the early stages of pregnancy (decidualization, implantation, and placentation) can have profound effects on macrophage activity and subsequently impact pregnancy outcome. In this review, we will provide an overview of the temporal and spatial regulation of utero-placental macrophage activation during normal pregnancy in human beings and rodents with a focus on more recent findings. We will also discuss the role of M1/M2 dysregulation within the intrauterine environment during adverse pregnancy outcomes. PMID:25505471

  2. Endoplasmic Reticulum Stress Controls M2 Macrophage Differentiation and Foam Cell Formation*

    PubMed Central

    Oh, Jisu; Riek, Amy E.; Weng, Sherry; Petty, Marvin; Kim, David; Colonna, Marco; Cella, Marina; Bernal-Mizrachi, Carlos

    2012-01-01

    Macrophages are essential in atherosclerosis progression, but regulation of the M1 versus M2 phenotype and their role in cholesterol deposition are unclear. We demonstrate that endoplasmic reticulum (ER) stress is a key regulator of macrophage differentiation and cholesterol deposition. Macrophages from diabetic patients were classically or alternatively stimulated and then exposed to oxidized LDL. Alternative stimulation into M2 macrophages lead to increased foam cell formation by inducing scavenger receptor CD36 and SR-A1 expression. ER stress induced by alternative stimulation was necessary to generate the M2 phenotype through JNK activation and increased PPARγ expression. The absence of CD36 or SR-A1 signaling independently of modified cholesterol uptake decreased ER stress and prevented the M2 differentiation typically induced by alternative stimulation. Moreover, suppression of ER stress shifted differentiated M2 macrophages toward an M1 phenotype and subsequently suppressed foam cell formation by increasing HDL- and apoA-1-induced cholesterol efflux indicating suppression of macrophage ER stress as a potential therapy for atherosclerosis. PMID:22356914

  3. mTORC2 signalling regulates M2 macrophage differentiation in response to helminth infection and adaptive thermogenesis.

    PubMed

    Hallowell, R W; Collins, S L; Craig, J M; Zhang, Y; Oh, M; Illei, P B; Chan-Li, Y; Vigeland, C L; Mitzner, W; Scott, A L; Powell, J D; Horton, M R

    2017-01-27

    Alternatively activated macrophages (M2) have an important function in innate immune responses to parasitic helminths, and emerging evidence also indicates these cells are regulators of systemic metabolism. Here we show a critical role for mTORC2 signalling in the generation of M2 macrophages. Abrogation of mTORC2 signalling in macrophages by selective conditional deletion of the adaptor molecule Rictor inhibits the generation of M2 macrophages while leaving the generation of classically activated macrophages (M1) intact. Selective deletion of Rictor in macrophages prevents M2 differentiation and clearance of a parasitic helminth infection in mice, and also abrogates the ability of mice to regulate brown fat and maintain core body temperature. Our findings define a role for mTORC2 in macrophages in integrating signals from the immune microenvironment to promote innate type 2 immunity, and also to integrate systemic metabolic and thermogenic responses.

  4. mTORC2 signalling regulates M2 macrophage differentiation in response to helminth infection and adaptive thermogenesis

    PubMed Central

    Hallowell, R. W.; Collins, S. L.; Craig, J. M.; Zhang, Y.; Oh, M.; Illei, P. B.; Chan-Li, Y.; Vigeland, C. L.; Mitzner, W.; Scott, A. L.; Powell, J. D.; Horton, M. R.

    2017-01-01

    Alternatively activated macrophages (M2) have an important function in innate immune responses to parasitic helminths, and emerging evidence also indicates these cells are regulators of systemic metabolism. Here we show a critical role for mTORC2 signalling in the generation of M2 macrophages. Abrogation of mTORC2 signalling in macrophages by selective conditional deletion of the adaptor molecule Rictor inhibits the generation of M2 macrophages while leaving the generation of classically activated macrophages (M1) intact. Selective deletion of Rictor in macrophages prevents M2 differentiation and clearance of a parasitic helminth infection in mice, and also abrogates the ability of mice to regulate brown fat and maintain core body temperature. Our findings define a role for mTORC2 in macrophages in integrating signals from the immune microenvironment to promote innate type 2 immunity, and also to integrate systemic metabolic and thermogenic responses. PMID:28128208

  5. Identification of M2 macrophages in anterior pituitary glands of normal rats and rats with estrogen-induced prolactinoma.

    PubMed

    Fujiwara, Ken; Yatabe, Megumi; Tofrizal, Alimuddin; Jindatip, Depicha; Yashiro, Takashi; Nagai, Ryozo

    2017-01-24

    Macrophages are present throughout the anterior pituitary gland. However, the features and function of macrophages in the gland are poorly understood. Recent studies have indicated that there are two main macrophage classes: M1 (classically activated) and M2 (alternatively activated). In this study, we examine whether both M1 and M2 macrophages are present in the anterior pituitary gland of rats. Our findings indicate that macrophages that are positive for CD68 (a pan-macrophage marker) were localized near capillaries in rat anterior pituitary gland. These macrophages were positive for iNOS or mannose receptor (MR), which are markers of M1 and M2 macrophages, respectively. To determine the morphological characteristics of M2 macrophages under pathological conditions, diethylstilbestrol (DES)-treated rats were used as an animal model of prolactinoma. After 2 weeks of DES treatment, a number of MR-immunopositive cells were present in the gland. Immunoelectron microscopy revealed that MR-immunopositive M2 macrophages had many small vesicles and moderately large vacuoles in cytoplasm. Phagosomes were sometimes present in cytoplasm. Interestingly, M2 macrophages in prolactinoma tissues did not usually exhibit distinct changes or differences during the normal, hyperplasia and adenoma stages. This study is the first to confirm that both M1 and M2 macrophages are present in the anterior pituitary gland of rats. Moreover, the number of M2 macrophages was greatly increased in rats with DES-induced prolactinoma. Future studies should attempt to characterize the functional role of M2 macrophages in the gland.

  6. Polarization of M2 macrophages requires Lamtor1 that integrates cytokine and amino-acid signals.

    PubMed

    Kimura, Tetsuya; Nada, Shigeyuki; Takegahara, Noriko; Okuno, Tatsusada; Nojima, Satoshi; Kang, Sujin; Ito, Daisuke; Morimoto, Keiko; Hosokawa, Takashi; Hayama, Yoshitomo; Mitsui, Yuichi; Sakurai, Natsuki; Sarashina-Kida, Hana; Nishide, Masayuki; Maeda, Yohei; Takamatsu, Hyota; Okuzaki, Daisuke; Yamada, Masaki; Okada, Masato; Kumanogoh, Atsushi

    2016-10-12

    Macrophages play crucial roles in host defence and tissue homoeostasis, processes in which both environmental stimuli and intracellularly generated metabolites influence activation of macrophages. Activated macrophages are classified into M1 and M2 macrophages. It remains unclear how intracellular nutrition sufficiency, especially for amino acid, influences on macrophage activation. Here we show that a lysosomal adaptor protein Lamtor1, which forms an amino-acid sensing complex with lysosomal vacuolar-type H(+)-ATPase (v-ATPase), and is the scaffold for amino acid-activated mTORC1 (mechanistic target of rapamycin complex 1), is critically required for M2 polarization. Lamtor1 deficiency, amino-acid starvation, or inhibition of v-ATPase and mTOR result in defective M2 polarization and enhanced M1 polarization. Furthermore, we identified liver X receptor (LXR) as the downstream target of Lamtor1 and mTORC1. Production of 25-hydroxycholesterol is dependent on Lamtor1 and mTORC1. Our findings demonstrate that Lamtor1 plays an essential role in M2 polarization, coupling immunity and metabolism.

  7. Polarization of M2 macrophages requires Lamtor1 that integrates cytokine and amino-acid signals

    PubMed Central

    Kimura, Tetsuya; Nada, Shigeyuki; Takegahara, Noriko; Okuno, Tatsusada; Nojima, Satoshi; Kang, Sujin; Ito, Daisuke; Morimoto, Keiko; Hosokawa, Takashi; Hayama, Yoshitomo; Mitsui, Yuichi; Sakurai, Natsuki; Sarashina-Kida, Hana; Nishide, Masayuki; Maeda, Yohei; Takamatsu, Hyota; Okuzaki, Daisuke; Yamada, Masaki; Okada, Masato; Kumanogoh, Atsushi

    2016-01-01

    Macrophages play crucial roles in host defence and tissue homoeostasis, processes in which both environmental stimuli and intracellularly generated metabolites influence activation of macrophages. Activated macrophages are classified into M1 and M2 macrophages. It remains unclear how intracellular nutrition sufficiency, especially for amino acid, influences on macrophage activation. Here we show that a lysosomal adaptor protein Lamtor1, which forms an amino-acid sensing complex with lysosomal vacuolar-type H+-ATPase (v-ATPase), and is the scaffold for amino acid-activated mTORC1 (mechanistic target of rapamycin complex 1), is critically required for M2 polarization. Lamtor1 deficiency, amino-acid starvation, or inhibition of v-ATPase and mTOR result in defective M2 polarization and enhanced M1 polarization. Furthermore, we identified liver X receptor (LXR) as the downstream target of Lamtor1 and mTORC1. Production of 25-hydroxycholesterol is dependent on Lamtor1 and mTORC1. Our findings demonstrate that Lamtor1 plays an essential role in M2 polarization, coupling immunity and metabolism. PMID:27731330

  8. The adult murine heart has a sparse, phagocytically active macrophage population that expands through monocyte recruitment and adopts an ‘M2’ phenotype in response to Th2 immunologic challenge

    PubMed Central

    Mylonas, Katie J.; Jenkins, Stephen J.; Castellan, Raphael F.P.; Ruckerl, Dominik; McGregor, Kieran; Phythian-Adams, Alexander T.; Hewitson, James P.; Campbell, Sharon M.; MacDonald, Andrew S.; Allen, Judith E.; Gray, Gillian A.

    2015-01-01

    Tissue resident macrophages have vital homeostatic roles in many tissues but their roles are less well defined in the heart. The present study aimed to identify the density, polarisation status and distribution of macrophages in the healthy murine heart and to investigate their ability to respond to immune challenge. Histological analysis of hearts from CSF-1 receptor (csf1-GFP; MacGreen) and CX3CR1 (Cx3cr1GFP/+) reporter mice revealed a sparse population of GFP positive macrophages that were evenly distributed throughout the left and right ventricular free walls and septum. F4/80+CD11b+ cardiac macrophages, sorted from myocardial homogenates, were able to phagocytose fluorescent beads in vitro and expressed markers typical of both ‘M1’ (IL-1β, TNF and CCR2) and ‘M2activation (Ym1, Arg 1, RELMα and IL-10), suggesting no specific polarisation in healthy myocardium. Exposure to Th2 challenge by infection of mice with helminth parasites Schistosoma mansoni, or Heligmosomoides polygyrus, resulted in an increase in cardiac macrophage density, adoption of a stellate morphology and increased expression of Ym1, RELMα and CD206 (mannose receptor), indicative of ‘M2’ polarisation. This was dependent on recruitment of Ly6ChighCCR2+ monocytes and was accompanied by an increase in collagen content. In conclusion, in the healthy heart resident macrophages are relatively sparse and have a phagocytic role. Following Th2 challenge this population expands due to monocyte recruitment and adopts an ‘M2’ phenotype associated with increased tissue fibrosis. PMID:25700973

  9. The adult murine heart has a sparse, phagocytically active macrophage population that expands through monocyte recruitment and adopts an 'M2' phenotype in response to Th2 immunologic challenge.

    PubMed

    Mylonas, Katie J; Jenkins, Stephen J; Castellan, Raphael F P; Ruckerl, Dominik; McGregor, Kieran; Phythian-Adams, Alexander T; Hewitson, James P; Campbell, Sharon M; MacDonald, Andrew S; Allen, Judith E; Gray, Gillian A

    2015-07-01

    Tissue resident macrophages have vital homeostatic roles in many tissues but their roles are less well defined in the heart. The present study aimed to identify the density, polarisation status and distribution of macrophages in the healthy murine heart and to investigate their ability to respond to immune challenge. Histological analysis of hearts from CSF-1 receptor (csf1-GFP; MacGreen) and CX3CR1 (Cx3cr1(GFP/+)) reporter mice revealed a sparse population of GFP positive macrophages that were evenly distributed throughout the left and right ventricular free walls and septum. F4/80+CD11b+ cardiac macrophages, sorted from myocardial homogenates, were able to phagocytose fluorescent beads in vitro and expressed markers typical of both 'M1' (IL-1β, TNF and CCR2) and 'M2' activation (Ym1, Arg 1, RELMα and IL-10), suggesting no specific polarisation in healthy myocardium. Exposure to Th2 challenge by infection of mice with helminth parasites Schistosoma mansoni, or Heligmosomoides polygyrus, resulted in an increase in cardiac macrophage density, adoption of a stellate morphology and increased expression of Ym1, RELMα and CD206 (mannose receptor), indicative of 'M2' polarisation. This was dependent on recruitment of Ly6ChighCCR2+ monocytes and was accompanied by an increase in collagen content. In conclusion, in the healthy heart resident macrophages are relatively sparse and have a phagocytic role. Following Th2 challenge this population expands due to monocyte recruitment and adopts an 'M2' phenotype associated with increased tissue fibrosis.

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

    PubMed Central

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

    2015-01-01

    Macrophages dictate both initiation and resolution of inflammation. During acute inflammation classically activated macrophages (M1) predominate, and during the resolution phase alternative macrophages (M2) are dominant. The molecular mechanisms involved in macrophage polarization are understudied. MicroRNAs are differentially expressed in M1 and M2 macrophages that influence macrophage polarization. We identified a role of miR-21 in macrophage polarization, and found that cross-talk between miR-21 and the lipid mediator prostaglandin E2 (PGE2) is a determining factor in macrophage polarization. miR-21 inhibition impairs expression of M2 signature genes but not M1 genes. PGE2 and its downstream effectors PKA and Epac inhibit miR-21 expression and enhance expression of M2 genes, and this effect is more pronounced in miR-21-/- cells. Among potential targets involved in macrophage polarization, we found that STAT3 and SOCS1 were enhanced in miR-21-/- cells and further enhanced by PGE2. We found that STAT3 was a direct target of miR-21 in macrophages. Silencing the STAT3 gene abolished PGE2-mediated expression of M2 genes in miR-21-/- macrophages. These data shed light on the molecular brakes involved in homeostatic macrophage polarization and suggest new therapeutic strategies to prevent inflammatory responses. PMID:25706647

  11. Tumor-derived interleukin-1 promotes lymphangiogenesis and lymph node metastasis through M2-type macrophages.

    PubMed

    Watari, Kosuke; Shibata, Tomohiro; Kawahara, Akihiko; Sata, Ken-ichi; Nabeshima, Hiroshi; Shinoda, Ai; Abe, Hideyuki; Azuma, Koichi; Murakami, Yuichi; Izumi, Hiroto; Takahashi, Takashi; Kage, Masayoshi; Kuwano, Michihiko; Ono, Mayumi

    2014-01-01

    Tumors formed by a highly metastatic human lung cancer cell line are characterized by activated signaling via vascular endothelial growth factor (VEGF)-C through its receptor (VEGFR-3) and aggressive lymph node metastasis. In this study, we examined how these highly metastatic cancers acquired aggressive lymph node metastasis. Compared with their lower metastatic counterparts, the highly metastatic tumors formed by this cell line expressed higher amounts of interleukin (IL)-1α, with similarly augmented expression of IL-1α and IL-1β by tumor stromal cells and of VEGF-A and VEGF-C by tumor-associated macrophages. These tumor-associated macrophages were mainly of the M2 type. Administration of a macrophage-targeting drug suppressed the production of these potent angiogenic and lymphangiogenic factors, resulting in decreased tumor growth, angiogenesis, lymphangiogenesis, and lymph node metastasis. In Matrigel plug assays, the highly metastatic cells formed tumors that were extensively infiltrated by M2-type macrophages and exhibited enhanced angiogenesis and lymphangiogenesis. All of these responses were suppressed by the IL-1 receptor (IL-1R) antagonist anakinra. Thus, the IL-1α-driven inflammatory activation of angiogenesis and lymphangiogenesis seems to provide a highly metastatic tumor microenvironment favorable for lymph node metastasis through cross-talk with macrophages. Accordingly, the IL-1R/M2-type macrophage axis may be a good therapeutic target for patients with this form of lung cancer.

  12. Medroxyprogesterone acetate drives M2 macrophage differentiation toward a phenotype of decidual macrophage.

    PubMed

    Tsai, Yung-Chieh; Tseng, Joseph T; Wang, Chia-Yih; Su, Mei-Tsz; Huang, Jyun-Yuan; Kuo, Pao-Lin

    2017-09-05

    M1 macrophage differentiation plays a crucial role in enhanced inflammation during pregnancy, which may lead to pregnancy complications. Therefore, modulation of macrophage differentiation toward the M2 phenotype is desirable to ensure a successful pregnancy. Medroxyprogesterone acetate (MPA) is a potent progestin with an anti-inflammatory property, but its effect on macrophage differentiation is unknown. This study aimed to examine whether MPA can induce an M2 macrophage differentiation by using the human monocytes cell line THP-1 or primary monocytes. THP-1 cells were primed with phorbol-12-myristate-13 acetate (PMA) to initiate macrophage differentiation. By incubating with MPA, the cells (denoted as MPA-pTHP-1) underwent M2 macrophage differentiation with downregulations of CD11c, IL-1β and TNF-α, and upregulations of CD163 and IL-10; while cells incubated with progesterone (P4) did not show the M2 phenotype. Primary monocytes treated with MPA also had the same M2 phenotype. Moreover, M1 macrophages derived from IFN-γ/LPS-treated THP-1 cells, which had high levels of IL-1b and iNOS, and low levels of IL-10 and IDO, were reversed to the M2 phenotype by the MPA treatment. We also found that the MPA-pTHP-1 promoted the decidualization of endometrial stromal cells and the invasion of trophoblast cells. To mimic conditions of exposure to various pathogens, MPA-pTHP-1 cells were stimulated by different types of TLR ligands. We found they produced lower levels of IL-1β and TNF-α, as well as a higher level of IL-10, compared to untreated cells. Finally, we found the level of phosphorylated ERK in the MPA-pTHP-1 cells was increased, but its IL-10 production was suppressed by either the progesterone/glucocorticoid antagonist (Mifepristone) or MEK inhibitor (U0126). Taken together, MPA could drive monocyte differentiation toward an M2 phenotype that mimics decidual macrophages. This finding holds great potential to combat chronic endometrial inflammation

  13. Chronic Opisthorchis viverrini infection and associated hepatobiliary disease is associated with iron loaded M2-like macrophages.

    PubMed

    Bility, Moses T; Sripa, Banchob

    2014-12-01

    Chronic Opisthorchis viverrini-induced hepatobiliary disease is associated with significant leukocyte infiltration, including activated macrophages; however, the polarization of infiltrating macrophages remains to be fully characterized. In this study, we characterized macrophage polarization and phenotype in chronic O. viverrini-induced hepatobiliary disease in humans and hamsters using gene expression and histochemical analysis. Chronic O. viverrini infection and associated hepatobiliary diseases were associated with iron loaded M2-like macrophages in both humans and hamsters. This study provides suggestive evidence that iron loaded M2-like macrophages promote hepatobiliary disease in chronic O. viverrini infection.

  14. Fasciola hepatica tegumental antigens indirectly induce an M2 macrophage-like phenotype in vivo.

    PubMed

    Adams, P N; Aldridge, A; Vukman, K V; Donnelly, S; O'Neill, S M

    2014-10-01

    The M2 subset of macrophages has a critical role to play in host tissue repair, tissue fibrosis and modulation of adaptive immunity during helminth infection. Infection with the helminth, Fasciola hepatica, is associated with M2 macrophages in its mammalian host, and this response is mimicked by its excretory-secretory products (FhES). The tegumental coat of F. hepatica (FhTeg) is another major source of immune-modulatory molecules; we have previously shown that FhTeg can modulate the activity of both dendritic cells and mast cells inhibiting their ability to prime a Th1 immune response. Here, we report that FhTeg does not induce Th2 immune responses but can induce M2-like phenotype in vivo that modulates cytokine production from CD4(+) cells in response to anti-CD3 stimulation. FhTeg induces a RELMα expressing macrophage population in vitro, while in vivo, the expression of Arg1 and Ym-1/2 but not RELMα in FhTeg-stimulated macrophages was STAT6 dependent. To support this finding, FhTeg induces RELMα expression in vivo prior to the induction of IL-13. FhTeg can induce IL-13-producing peritoneal macrophages following intraperitoneal injection This study highlights the important role of FhTeg as an immune-modulatory source during F. hepatica infection and sheds further light on helminth-macrophage interactions.

  15. Pharmacological effects of mitraphylline from Uncaria tomentosa in primary human monocytes: Skew toward M2 macrophages.

    PubMed

    Montserrat-de la Paz, S; de la Puerta, R; Fernandez-Arche, A; Quilez, A M; Muriana, F J G; Garcia-Gimenez, M D; Bermudez, B

    2015-07-21

    Uncaria tomentosa (Willdenow ex Roemer & Schultes) DC. (Rubiaceae) is a Peruvian thorny liana, commonly known as "cat׳s claw", and traditionally used in folk medicine to deal with several inflammatory diseases. Mitraphylline (MTP) is the most abundant pentacyclic oxindolic alkaloid (POA) from U. Tomentosa and has been reported to modify the inflammatory response. Herein, we have sought to identify the mechanisms underlying this modulatory effect of MTP on primary human monocytes and its ability to regulate differentiation processes on human primary monocyte and monocyte-derived macrophages. In vitro studies with human primary monocytes and monocyte-derived macrophages were performed. Monocytes and M0 macrophages were exposed to MTP (25μM) and LPS (100ng/mL). M0 macrophages were polarized to M1 and M2 phenotypes in the absence or presence of MTP. The activation state of monocytes/macrophages was assessed by flow cytometry, gene expression and protein analysis of different specific markers. In human primary monocytes, the incubation of MTP for 24h reduced the number of classical (CD14(++)CD16(-)) and intermediate (CD14(++)CD16(+)) subsets when compared to untreated or LPS-treated cells. MTP also reduced the chemotactic capacity of human primary monocytes. In addition, MTP promoted the polarization of M0 macrophages toward an anti-inflammatory M2 phenotype, the abrogation of the release of pro-inflammatory cytokines such as TNFα, IL-6 or IL-1β, as well as the restoration of markers for M2 macrophages in LPS-treated M1 macrophages. Our results suggest that MTP may be a key modulator for regulating the plasticity of monocytes/macrophages and the attenuation of the inflammatory response. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

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

    PubMed Central

    Bi, Jia; Zeng, Xianxin; Zhao, Lin; Wei, Qian; Yu, Lifeng; Wang, Xinnan; Yu, Zhaojin; Cao, Yaming; Shan, Fengping; Wei, Minjie

    2016-01-01

    Macrophages can acquire a variety of polarization status and functions: classically activated macrophages (M1 macrophages); alternatively activated macrophages (M2 macrophages). However, the molecular basis of the process is still unclear. Here, this study addresses that microRNA-181a (miR-181a) is a key molecule controlling macrophage polarization. We found that miR-181a is overexpressed in M2 macrophages than in M1 macrophages. miR-181a expression was decreased when M2 phenotype converted to M1, whereas it increased when M1 phenotype converted to M2. Overexpression of miR-181a in M1 macrophages diminished M1 phenotype expression while promoting polarization to the M2 phenotype. In contrast, knockdown of miR-181a in M2 macrophages promoted M1 polarization and diminished M2 phenotype expression. Mechanistically, Bioinformatic analysis revealed that Kruppel-like factor 6 (KLF6) and CCAAT/enhancer binding protein-α (C/EBPα) is a potential target of miR-181a and luciferase assay confirmed that KLF6 and C/EBPα translation is suppressed by miR-181a through interaction with the 3′UTR of KLF6 and C/EBPα mRNA. Further analysis showed that induction of miR-181a suppressed KLF6 and C/EBPα protein expression. Importantly, miR-181a also diminishes M2 macrophages-mediated migration and invasion capacity of tumor cells. Collectively, our results suggest that miR-181a plays a significant role in regulating macrophage polarization through directly target KLF6 and C/EBPα. PMID:27673564

  17. Schistosomal-derived lysophosphatidylcholine triggers M2 polarization of macrophages through PPARγ dependent mechanisms.

    PubMed

    Assunção, Leonardo Santos; Magalhães, Kelly G; Carneiro, Alan Brito; Molinaro, Raphael; Almeida, Patrícia E; Atella, Georgia C; Castro-Faria-Neto, Hugo C; Bozza, Patrícia T

    2017-02-01

    Mansonic schistosomiasis is a disease caused by the trematode Schistosoma mansoni, endemic to tropical countries. S. mansoni infection induces the formation of granulomas and potent polarization of Th2-type immune response. There is great interest in understanding the mechanisms used by this parasite that causes a modulation of the immune system. Recent studies from our group demonstrated that lipids of S. mansoni, including lysophosphatidylcholine (LPC) have immunomodulatory activity. In the present study, our aim was to investigate the role of lipids derived from S. mansoni in the activation and polarization of macrophages and to characterize the mechanisms involved in this process. Peritoneal macrophages obtained from wild type C57BL/6mice or bone marrow derived macrophages were stimulated in vitro with lipids extracted from adult worms of S. mansoni. We demonstrated that total schistosomal-derived lipids as well as purified LPC induced alternatively activated macrophages/M2 profile observed by increased expression of arginase-1, mannose receptor, Chi3l3, TGFβ and production of IL-10 and PGE2 24h after stimulation. The involvement of the nuclear receptor PPARγ in macrophage response against LPC was investigated. Through Western blot and immunofluorescence confocal microscopy we demonstrated that schistosomal-derived LPC induces increased expression of PPARγ in macrophages. The LPC-induced increased expression of arginase-1 were significantly inhibited by the PPAR-γ antagonist GW9662. Together, these results demonstrate an immunomodulatory role of schistosomal-derived LPC in activating macrophages to a profile of the type M2 through PPARγ-dependent mechanisms, indicating a novel pathway for macrophage polarization triggered by parasite-derived LPC with potential implications to disease pathogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

  18. Opposite Effects of M1 and M2 Macrophage Subtypes on Lung Cancer Progression.

    PubMed

    Yuan, Ang; Hsiao, Yi-Jing; Chen, Hsuan-Yu; Chen, Huei-Wen; Ho, Chao-Chi; Chen, Yu-Yun; Liu, Yi-Chia; Hong, Tsai-Hsia; Yu, Sung-Liang; Chen, Jeremy J W; Yang, Pan-Chyr

    2015-09-24

    Macrophages in a tumor microenvironment have been characterized as M1- and M2-polarized subtypes. Here, we discovered the different macrophages' impacts on lung cancer cell A549. The M2a/M2c subtypes promoted A549 invasion and xenograft tumor growth. The M1 subtype suppressed angiogenesis. M1 enhanced the sensitivity of A549 to cisplatin and decreased the tube formation activity and cell viability of A549 cells by inducing apoptosis and senescence. Different macrophage subtypes regulated genes involved in the immune response, cytoskeletal remodeling, coagulation, cell adhesion, and apoptosis pathways in A549 cells, which was a pattern that correlated with the altered behaviors of the A549 cells. Furthermore, we found that the identified M1/M2 gene signatures were significantly correlated with the extended overall survival of lung cancer patients. These results suggest that M1/M2 gene expression signature may be used as a prognostic indicator for lung cancer patients, and M1/M2 polarization may be a target of investigation of immune-modulating therapies for lung cancer in the future.

  19. PPARγ Ligands Switched High Fat Diet-Induced Macrophage M2b Polarization toward M2a Thereby Improving Intestinal Candida Elimination

    PubMed Central

    Olagnier, David; Bernad, José; Perez, Laurence; Burcelin, Rémy; Valentin, Alexis; Auwerx, Johan; Pipy, Bernard; Coste, Agnès

    2010-01-01

    Obesity is associated with a chronic low-grade inflammation that predisposes to insulin resistance and the development of type 2 diabetes. In this metabolic context, gastrointestinal (GI) candidiasis is common. We recently demonstrated that the PPARγ ligand rosiglitazone promotes the clearance of Candida albicans through the activation of alternative M2 macrophage polarization. Here, we evaluated the impact of high fat diet (HFD)-induced obesity and the effect of rosiglitazone (PPARγ ligand) or WY14643 (PPARα ligand) both on the phenotypic M1/M2 polarization of peritoneal and cecal tissue macrophages and on the outcome of GI candidiasis. We demonstrated that the peritoneal macrophages and the cell types present in the cecal tissue from HF fed mice present a M2b polarization (TNF-αhigh, IL-10high, MR, Dectin-1). Interestingly, rosiglitazone induces a phenotypic M2b-to-M2a (TNF-αlow, IL-10low, MRhigh, Dectin-1high) switch of peritoneal macrophages and of the cells present in the cecal tissue. The incapacity of WY14643 to switch this polarization toward M2a state, strongly suggests the specific involvement of PPARγ in this mechanism. We showed that in insulin resistant mice, M2b polarization of macrophages present on the site of infection is associated with an increased susceptibility to GI candidiasis, whereas M2a polarization after rosiglitazone treatment favours the GI fungal elimination independently of reduced blood glucose. In conclusion, our data demonstrate a dual benefit of PPARγ ligands because they promote mucosal defence mechanisms against GI candidiasis through M2a macrophage polarization while regulating blood glucose level. PMID:20877467

  20. Contact-dependent carcinoma aggregate dispersion by M2a macrophages via ICAM-1 and β2 integrin interactions.

    PubMed

    Bai, Jing; Adriani, Giulia; Dang, Truong-Minh; Tu, Ting-Yuan; Penny, Hwei-Xian Leong; Wong, Siew-Cheng; Kamm, Roger D; Thiery, Jean-Paul

    2015-09-22

    Tumor-associated macrophages (TAMs) can constitute up to 50% of the tumor mass and have strong implications in tumor progression and metastasis. Macrophages are plastic and can polarize to various subtypes that differ in terms of surface receptor expression as well as cytokine and chemokine production and effector function. Conventionally, macrophages are grouped into two major subtypes: the classically activated M1 macrophages and the alternatively activated M2 macrophages. M1 macrophages are pro-inflammatory, promote T helper (Th) 1 responses, and show tumoricidal activity, whereas M2 macrophages contribute to tissue repair and promote Th2 responses. Herein, we present a microfluidic system integrating tumor cell aggregates and subtypes of human monocyte-derived macrophages in a three-dimensional hydrogel scaffold, in close co-culture with an endothelial monolayer to create an in vitro tumor microenvironment. This platform was utilized to study the role of individual subtypes of macrophages (M0, M1, M2a, M2b and M2c) in human lung adenocarcinoma (A549) aggregate dispersion, as a representation of epithelial-mesenchymal transition (EMT). A significant difference was observed when M2a macrophages were in direct contact with or separated from A549 aggregates, suggesting a possible mechanism for proximity-induced, contact-dependent dissemination via ICAM-1 and integrin β2 interactions. Indeed, M2a macrophages tended to infiltrate and release cells from carcinoma cell aggregates. These findings may help in the development of immunotherapies based on enhancing the tumor-suppressive properties of TAMs.

  1. Vitamin D prevents podocyte injury via regulation of macrophage M1/M2 phenotype in diabetic nephropathy rats.

    PubMed

    Zhang, Xiao-Liang; Guo, Yin-Feng; Song, Zhi-Xia; Zhou, Min

    2014-12-01

    Increasing evidence suggests the heterogeneity of macrophage phenotype and function ultimately determines the outcome of diabetic nephropathy (DN). This study aimed to investigate the effects of vitamin D on macrophage M1/M2 phenotype and its role in preventing podocyte impairment in streptozotocin-induced DN rats. Calcitriol, a bioactive 1,25-dihydroxyvitamin D3, ameliorated proteinuria and renal damage as well as reversed the decline of both nephrin and podocin, crucial structural proteins in podocytes. DN rats showed increased infiltrating macrophages with M1 phenotype characterized by elevated expression of inducible nitric oxide synthase and TNF-α in glomeruli and interstitium, which were inhibited after calcitriol treatment. Interestingly, calcitriol promoted M2 macrophage activation with enhanced expression of CD163, arginase-1, and mannose receptor at week 18 but not at week 8 or 14. The ratio of CD163 to CD68, considered as the proportion of M2 macrophages, was about 2.9-fold higher at week 18 after calcitriol treatment. Furthermore, the protein expression of inducible nitric oxide synthase, a crucial marker of M1 macrophages, was negatively correlated with the expression of either nephrin or podocin, whereas CD163, indicating M2 macrophages, was positively correlated. In vitro, 1,25-dihydroxyvitamin D3 switched high-glucose-induced M1 macrophages toward an M2 phenotype in either U937-derived macrophages or RAW264.7 cells. Our results suggest that vitamin D not only reduces macrophage infiltration and inhibits M1 macrophage activation but also enhances M2 macrophage phenotype to protect against podocyte injury.

  2. Hepatocellular carcinoma is accelerated by NASH involving M2 macrophage polarization mediated by hif-1αinduced IL-10.

    PubMed

    Ambade, Aditya; Satishchandran, Abhishek; Saha, Banishree; Gyongyosi, Benedek; Lowe, Patrick; Kodys, Karen; Catalano, Donna; Szabo, Gyongyi

    2016-01-01

    Obesity-related inflammation promotes cancer development. Tissue resident macrophages affect tumor progression and the tumor micro-environment favors polarization into alternatively activated macrophages (M2) that facilitate tumor invasiveness. Here, we dissected the role of western diet-induced NASH in inducing macrophage polarization in a carcinogen initiated model of hepatocellular carcinoma (HCC). Adult C57BL/6 male mice received diethyl nitrosamine (DEN) followed by 24 weeks of high fat-high cholesterol-high sugar diet (HF-HC-HSD). We assessed liver MRI and histology, serum ALT, AFP, liver triglycerides, and cytokines. Macrophage polarization was determined by IL-12/TNFα (M1) and CD163/CD206 (M2) expression using flow cytometry. Role of hif-1α-induced IL-10 was dissected in hepatocyte specific hif-1αKO and hif-1αdPA (over-expression) mice. The western diet-induced features of NASH and accelerated HCC development after carcinogen exposure. Liver fibrosis and serum AFP were significantly increased in DEN + HF-HC-HSD mice compared to controls. Western diet resulted in macrophage (F4/80(+)CD11b(+)) infiltration to liver and DEN + HF-HC-HSD mice showed preferential increase in M2 macrophages. Isolated hepatocytes from western diet fed mice showed significant upregulation of the hypoxia-inducible transcription factor, hif-1α, and livers from hif-1α over-expressing mice had increased proportion of M2 macrophages. Primary hepatocytes from wild-type mice treated with DEN and palmitic acid in vitro showed activation of hif-1α and induction of IL-10, a M2 polarizing cytokine. IL-10 neutralization in hepatocyte-derived culture supernatant prevented M2 macrophage polarization and silencing hif-1α in macrophages blocked their M2 polarization. Therefore, our data demonstrate that NASH accelerates HCC progression via upregulation of hif-1α mediated IL-10 polarizing M2 macrophages.

  3. Wound administration of M2-polarized macrophages does not improve murine cutaneous healing responses.

    PubMed

    Jetten, Nadine; Roumans, Nadia; Gijbels, Marion J; Romano, Andrea; Post, Mark J; de Winther, Menno P J; van der Hulst, Rene R W J; Xanthoulea, Sofia

    2014-01-01

    Macrophages play a crucial role in all stages of cutaneous wound healing responses and dysregulation of macrophage function can result in derailed wound repair. The phenotype of macrophages is influenced by the wound microenvironment and evolves during healing from a more pro-inflammatory (M1) profile in early stages, to a less inflammatory pro-healing (M2) phenotype in later stages of repair. The aim of the current study was to investigate the potential of exogenous administration of M2 macrophages to promote wound healing in an experimental mouse model of cutaneous injury. Bone marrow derived macrophages were stimulated in-vitro with IL-4 or IL-10 to obtain two different subsets of M2-polarized cells, M2a or M2c respectively. Polarized macrophages were injected into full-thickness excisional skin wounds of either C57BL/6 or diabetic db/db mice. Control groups were injected with non-polarized (M0) macrophages or saline. Our data indicate that despite M2 macrophages exhibit an anti-inflammatory phenotype in-vitro, they do not improve wound closure in wild type mice while they delay healing in diabetic mice. Examination of wounds on day 15 post-injury indicated delayed re-epithelialization and persistence of neutrophils in M2 macrophage treated diabetic wounds. Therefore, topical application of ex-vivo generated M2 macrophages is not beneficial and contraindicated for cell therapy of skin wounds.

  4. Parthenolide Relieves Pain and Promotes M2 Microglia/Macrophage Polarization in Rat Model of Neuropathy.

    PubMed

    Popiolek-Barczyk, Katarzyna; Kolosowska, Natalia; Piotrowska, Anna; Makuch, Wioletta; Rojewska, Ewelina; Jurga, Agnieszka M; Pilat, Dominika; Mika, Joanna

    2015-01-01

    Neuropathic pain treatment remains a challenge because pathomechanism is not fully understood. It is believed that glial activation and increased spinal nociceptive factors are crucial for neuropathy. We investigated the effect of parthenolide (PTL) on the chronic constriction injury to the sciatic nerve (CCI)-induced neuropathy in rat. We analyzed spinal changes in glial markers and M1 and M2 polarization factors, as well as intracellular signaling pathways. PTL (5 µg; i.t.) was preemptively and then daily administered for 7 days after CCI. PTL attenuated the allodynia and hyperalgesia and increased the protein level of IBA1 (a microglial/macrophage marker) but did not change GFAP (an astrocyte marker) on day 7 after CCI. PTL reduced the protein level of M1 (IL-1β, IL-18, and iNOS) and enhanced M2 (IL-10, TIMP1) factors. In addition, it downregulated the phosphorylated form of NF-κB, p38MAPK, and ERK1/2 protein level and upregulated STAT3. In primary microglial cell culture we have shown that IL-1β, IL-18, iNOS, IL-6, IL-10, and TIMP1 are of microglial origin. Summing up, PTL directly or indirectly attenuates neuropathy symptoms and promotes M2 microglia/macrophages polarization. We suggest that neuropathic pain therapies should be shifted from blanketed microglia/macrophage suppression toward maintenance of the balance between neuroprotective and neurotoxic microglia/macrophage phenotypes.

  5. Parthenolide Relieves Pain and Promotes M2 Microglia/Macrophage Polarization in Rat Model of Neuropathy

    PubMed Central

    Popiolek-Barczyk, Katarzyna; Kolosowska, Natalia; Makuch, Wioletta; Rojewska, Ewelina; Jurga, Agnieszka M.; Pilat, Dominika

    2015-01-01

    Neuropathic pain treatment remains a challenge because pathomechanism is not fully understood. It is believed that glial activation and increased spinal nociceptive factors are crucial for neuropathy. We investigated the effect of parthenolide (PTL) on the chronic constriction injury to the sciatic nerve (CCI)-induced neuropathy in rat. We analyzed spinal changes in glial markers and M1 and M2 polarization factors, as well as intracellular signaling pathways. PTL (5 µg; i.t.) was preemptively and then daily administered for 7 days after CCI. PTL attenuated the allodynia and hyperalgesia and increased the protein level of IBA1 (a microglial/macrophage marker) but did not change GFAP (an astrocyte marker) on day 7 after CCI. PTL reduced the protein level of M1 (IL-1β, IL-18, and iNOS) and enhanced M2 (IL-10, TIMP1) factors. In addition, it downregulated the phosphorylated form of NF-κB, p38MAPK, and ERK1/2 protein level and upregulated STAT3. In primary microglial cell culture we have shown that IL-1β, IL-18, iNOS, IL-6, IL-10, and TIMP1 are of microglial origin. Summing up, PTL directly or indirectly attenuates neuropathy symptoms and promotes M2 microglia/macrophages polarization. We suggest that neuropathic pain therapies should be shifted from blanketed microglia/macrophage suppression toward maintenance of the balance between neuroprotective and neurotoxic microglia/macrophage phenotypes. PMID:26090236

  6. Regulation of Macrophage Motility by the Water Channel Aquaporin-1: Crucial Role of M0/M2 Phenotype Switch

    PubMed Central

    Tyteca, Donatienne; Nishino, Tomoya; Debaix, Huguette; Van Der Smissen, Patrick; N'Kuli, Francisca; Hoffmann, Delia; Cnops, Yvette; Rabolli, Virginie; van Loo, Geert; Beyaert, Rudi; Huaux, François; Devuyst, Olivier; Courtoy, Pierre J.

    2015-01-01

    The water channel aquaporin-1 (AQP1) promotes migration of many cell types. Although AQP1 is expressed in macrophages, its potential role in macrophage motility, particularly in relation with phenotype polarization, remains unknown. We here addressed these issues in peritoneal macrophages isolated from AQP1-deficient mice, either undifferentiated (M0) or stimulated with LPS to orientate towards pro-inflammatory phenotype (classical macrophage activation; M1). In non-stimulated macrophages, ablation of AQP1 (like inhibition by HgCl2) increased by 2–3 fold spontaneous migration in a Src/PI3K/Rac-dependent manner. This correlated with cell elongation and formation of lamellipodia/ruffles, resulting in membrane lipid and F4/80 recruitment to the leading edge. This indicated that AQP1 normally suppresses migration of resting macrophages, as opposed to other cell types. Resting Aqp1-/- macrophages exhibited CD206 redistribution into ruffles and increased arginase activity like IL4/IL13 (alternative macrophage activation; M2), indicating a M0-M2 shift. In contrast, upon M1 orientation by LPS in vitro or peritoneal inflammation in vivo, migration of Aqp1-/- macrophages was reduced. Taken together, these data indicate that AQP1 oppositely regulates macrophage migration, depending on stimulation or not by LPS, and that macrophage phenotypic and migratory changes may be regulated independently of external cues. PMID:25719758

  7. Tumour cell conditioned medium reveals greater M2 skewing of macrophages in the absence of properdin

    PubMed Central

    Al‐Rayahi, Izzat A.M.; Browning, Michael J.

    2017-01-01

    Abstract Introduction The tumour microenvironment is shaped by the interaction of immune, non immune, and tumour cells present in close proximity. Tumour cells direct the development of a locally immune suppressed state, affecting the activity of anti tumour T cells and preparing the escape phase of tumour development. Macrophages in the tumour typically develop into so‐called tumour associated macrophages with a distinct profile of activities which lead to a reduction in inflammation and antigen presentation. The direct impact of tumour cell conditioned medium on the activity profile of macrophages in dependence of their complement component expression has not yet been investigated. Methods In our in vitro study, macrophages differentiated from bone marrows of properdin deficient and wildtype mice were stimulated with conditioned medium of a syngeneic tumour cell line, B16F10, a mouse melanoma subline. Results In comparison with macrophages from wildtype mice, those from congenic properdin deficient mice showed skewing towards M2 profile, encompassing mRNA expression for genes involved in arginine metabolism, production of type 2 cytokines, and relatively lower surface expression of molecules needed for antigen presentation. Conclusions These data suggest that properdin insufficiency promotes a tumour environment that helps the tumour evade the immune response. PMID:28250926

  8. Systemic and Cardiac Depletion of M2 Macrophage through CSF-1R Signaling Inhibition Alters Cardiac Function Post Myocardial Infarction

    PubMed Central

    Martin, Kenneth; Turner, Elizebeth C.; Kumar, Arun H.; Browne, Tara; Caplice, Noel M.

    2015-01-01

    The heart hosts tissue resident macrophages which are capable of modulating cardiac inflammation and function by multiple mechanisms. At present, the consequences of phenotypic diversity in macrophages in the heart are incompletely understood. The contribution of cardiac M2-polarized macrophages to the resolution of inflammation and repair response following myocardial infarction remains to be fully defined. In this study, the role of M2 macrophages was investigated utilising a specific CSF-1 receptor signalling inhibition strategy to achieve their depletion. In mice, oral administration of GW2580, a CSF-1R kinase inhibitor, induced significant decreases in Gr1lo and F4/80hi monocyte populations in the circulation and the spleen. GW2580 administration also induced a significant depletion of M2 macrophages in the heart after 1 week treatment as well as a reduction of cardiac arginase1 and CD206 gene expression indicative of M2 macrophage activity. In a murine myocardial infarction model, reduced M2 macrophage content was associated with increased M1-related gene expression (IL-6 and IL-1β), and decreased M2-related gene expression (Arginase1 and CD206) in the heart of GW2580-treated animals versus vehicle-treated controls. M2 depletion was also associated with a loss in left ventricular contractile function, infarct enlargement, decreased collagen staining and increased inflammatory cell infiltration into the infarct zone, specifically neutrophils and M1 macrophages. Taken together, these data indicate that CSF-1R signalling is critical for maintaining cardiac tissue resident M2-polarized macrophage population, which is required for the resolution of inflammation post myocardial infarction and, in turn, for preservation of ventricular function. PMID:26407006

  9. Systemic and Cardiac Depletion of M2 Macrophage through CSF-1R Signaling Inhibition Alters Cardiac Function Post Myocardial Infarction.

    PubMed

    Leblond, Anne-Laure; Klinkert, Kerstin; Martin, Kenneth; Turner, Elizebeth C; Kumar, Arun H; Browne, Tara; Caplice, Noel M

    2015-01-01

    The heart hosts tissue resident macrophages which are capable of modulating cardiac inflammation and function by multiple mechanisms. At present, the consequences of phenotypic diversity in macrophages in the heart are incompletely understood. The contribution of cardiac M2-polarized macrophages to the resolution of inflammation and repair response following myocardial infarction remains to be fully defined. In this study, the role of M2 macrophages was investigated utilising a specific CSF-1 receptor signalling inhibition strategy to achieve their depletion. In mice, oral administration of GW2580, a CSF-1R kinase inhibitor, induced significant decreases in Gr1lo and F4/80hi monocyte populations in the circulation and the spleen. GW2580 administration also induced a significant depletion of M2 macrophages in the heart after 1 week treatment as well as a reduction of cardiac arginase1 and CD206 gene expression indicative of M2 macrophage activity. In a murine myocardial infarction model, reduced M2 macrophage content was associated with increased M1-related gene expression (IL-6 and IL-1β), and decreased M2-related gene expression (Arginase1 and CD206) in the heart of GW2580-treated animals versus vehicle-treated controls. M2 depletion was also associated with a loss in left ventricular contractile function, infarct enlargement, decreased collagen staining and increased inflammatory cell infiltration into the infarct zone, specifically neutrophils and M1 macrophages. Taken together, these data indicate that CSF-1R signalling is critical for maintaining cardiac tissue resident M2-polarized macrophage population, which is required for the resolution of inflammation post myocardial infarction and, in turn, for preservation of ventricular function.

  10. IRF5 regulates lung macrophages M2 polarization during severe acute pancreatitis in vitro

    PubMed Central

    Sun, Kang; He, Song-Bing; Qu, Jian-Guo; Dang, Sheng-Chun; Chen, Ji-Xiang; Gong, Ai-Hua; Xie, Rong; Zhang, Jian-Xin

    2016-01-01

    AIM To investigate the role of interferon regulatory factor 5 (IRF5) in reversing polarization of lung macrophages during severe acute pancreatitis (SAP) in vitro. METHODS A mouse SAP model was established by intraperitoneal (ip) injections of 20 μg/kg body weight caerulein. Pathological changes in the lung were observed by hematoxylin and eosin staining. Lung macrophages were isolated from bronchoalveolar lavage fluid. The quantity and purity of lung macrophages were detected by fluorescence-activated cell sorting and evaluated by real-time polymerase chain reaction (RT-PCR). They were treated with IL-4/IRF5 specific siRNA (IRF5 siRNA) to reverse their polarization and were evaluated by detecting markers expression of M1/M2 using RT-PCR. RESULTS SAP associated acute lung injury (ALI) was induced successfully by ip injections of caerulein, which was confirmed by histopathology. Lung macrophages expressed high levels of IRF5 as M1 phenotype during the early acute pancreatitis stages. Reduction of IRF5 expression by IRF5 siRNA reversed the action of macrophages from M1 to M2 phenotype in vitro. The expressions of M1 markers, including IRF5 (S + IRF5 siRNA vs S + PBS, 0.013 ± 0.01 vs 0.054 ± 0.047, P < 0.01), TNF-α (S + IRF5 siRNA vs S + PBS, 0.0003 ± 0.0002 vs 0.019 ± 0.018, P < 0.001), iNOS (S + IRF5 siRNA vs S + PBS, 0.0003 ± 0.0002 vs 0.026 ± 0.018, P < 0.001) and IL-12 (S + IRF5 siRNA vs S + PBS, 0.000005 ± 0.00004 vs 0.024 ± 0.016, P < 0.001), were decreased. In contrast, the expressions of M2 markers, including IL-10 (S + IRF5 siRNA vs S + PBS, 0.060 ± 0.055 vs 0.0230 ± 0.018, P < 0.01) and Arg-1 (S + IRF5 siRNA vs S + PBS, 0.910 ± 0.788 vs 0.0036 ± 0.0025, P < 0.001), were increased. IRF5 siRNA could reverse the lung macrophage polarization more effectively than IL-4. CONCLUSION Treatment with IRF5 siRNA can reverse the pancreatitis-induced activation of lung macrophages from M1 phenotype to M2 phenotype in SAP associated with ALI. PMID:27895424

  11. Predictive screening of M1 and M2 macrophages reveals the immunomodulatory effectiveness of post spinal cord injury azithromycin treatment

    PubMed Central

    Gensel, John C.; Kopper, Timothy J.; Zhang, Bei; Orr, Michael B.; Bailey, William M.

    2017-01-01

    Spinal cord injury (SCI) triggers a heterogeneous macrophage response that when experimentally polarized toward alternative forms of activation (M2 macrophages) promotes tissue and functional recovery. There are limited pharmacological therapies that can drive this reparative inflammatory state. In the current study, we used in vitro systems to comprehensively defined markers of macrophages with known pathological (M1) and reparative (M2) properties in SCI. We then used these markers to objectively define the macrophage activation states after SCI in response to delayed azithromycin treatment. Mice were subjected to moderate-severe thoracic contusion SCI. Azithromycin or vehicle was administered beginning 30 minutes post-SCI and then daily for 3 or 7 days post injury (dpi). We detected a dose-dependent polarization toward purportedly protective M2 macrophages with daily AZM treatment. Specifically, AZM doses of 10, 40, or 160 mg/kg decreased M1 macrophage gene expression at 3 dpi while the lowest (10 mg/kg) and highest (160 mg/kg) doses increased M2 macrophage gene expression at 7 dpi. Azithromycin has documented immunomodulatory properties and is commonly prescribed to treat infections in SCI individuals. This work demonstrates the utility of objective, comprehensive macrophage gene profiling for evaluating immunomodulatory SCI therapies and highlights azithromycin as a promising agent for SCI treatment. PMID:28057928

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

    PubMed Central

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

    2016-01-01

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

  13. M2 macrophages coexist with a Th1-driven profile in periapical cysts.

    PubMed

    Ribeiro, C M; de Carli, M L; Nonogaki, S; Nogueira, D A; Pereira, A A C; Sperandio, F F; Hanemann, J A C

    2017-08-30

    To evaluate the participation of both Th1 and Th2 responses in periapical cysts by assessing the presence of M2 macrophages, as well as acute IL-1 β, TNF-α and IL-6 cytokines. Twenty-four cases of periapical cysts were selected. Immuno-expressions of IL-1 β, IL-6, TNF-α and CD163 were analysed in the cystic capsules in both superficial and deeper regions. Data were analysed with paired Wilcoxon test and Spearman correlation coefficient (P ≤ 0.05). There was a higher expression of IL-1β, IL-6, TNF-α and M2 macrophages in the superficial region (P < 0.001) of cystic capsules. All acute cytokines had significant positive correlations amongst them regardless of the cystic capsule region. Regarding CD163, positive correlations occurred only with TNF-α (P = 0.007; r = 0.537) and IL-6 (P = 0.018; r = 0.478) in the superficial regions of the cystic capsule. M2 macrophages participated actively in the inflammatory response of periapical cysts and correlated with the expression of certain acute Th1-related cytokines. This illustrates the coexistence of an acute and chronic Th2-driven immune response in these lesions. Although M2 macrophages favour the healing process, their presence is not sufficient for periapical cyst regression, once an acute active response has occurred due to an infectious stimuli. © 2017 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  14. Paraoxonase 2 Induces a Phenotypic Switch in Macrophage Polarization Favoring an M2 Anti-Inflammatory State.

    PubMed

    Koren-Gluzer, Marie; Rosenblat, Mira; Hayek, Tony

    2015-01-01

    Inflammatory processes are involved in atherosclerosis development. Macrophages play a major role in the early atherogenesis, and they are present in the atherosclerotic lesion in two phenotypes: proinflammatory (M1) or anti-inflammatory (M2). Paraoxonase 2 (PON2) is expressed in macrophages, and it was shown to protect against atherosclerosis. Thus, the aim of our study was to analyze the direct effect of PON2 on macrophage inflammatory phenotypes. Ex vivo studies were performed with murine peritoneal macrophages (MPM) harvested from control C57BL/6 and PON2-deficient (PON2KO) mice. PON2KO MPM showed an enhanced proinflammatory phenotype compared to the control, both in the basal state and following M1 activation by IFNγ and lipopolysaccharide (LPS). In parallel, PON2KO MPM also showed reduced anti-inflammatory responses in the basal state and also following M2 activation by IL-4. Moreover, the PON2-null MPM demonstrated enhanced phagocytosis and reactive oxygen species (ROS) production in the basal state and following M1 activation. The direct effect of PON2 was shown by transfecting human PON2 (hPON2) into PON2KO MPM. PON2 transfection attenuated the macrophages' response to M1 activation and enhanced M2 response. These PON2 effects were associated with attenuation of macrophages' abilities to phagocyte and to generate ROS. We conclude that PON2 promotes an M1 to M2 switch in macrophage phenotypes.

  15. Paraoxonase 2 Induces a Phenotypic Switch in Macrophage Polarization Favoring an M2 Anti-Inflammatory State

    PubMed Central

    Koren-Gluzer, Marie; Rosenblat, Mira; Hayek, Tony

    2015-01-01

    Inflammatory processes are involved in atherosclerosis development. Macrophages play a major role in the early atherogenesis, and they are present in the atherosclerotic lesion in two phenotypes: proinflammatory (M1) or anti-inflammatory (M2). Paraoxonase 2 (PON2) is expressed in macrophages, and it was shown to protect against atherosclerosis. Thus, the aim of our study was to analyze the direct effect of PON2 on macrophage inflammatory phenotypes. Ex vivo studies were performed with murine peritoneal macrophages (MPM) harvested from control C57BL/6 and PON2-deficient (PON2KO) mice. PON2KO MPM showed an enhanced proinflammatory phenotype compared to the control, both in the basal state and following M1 activation by IFNγ and lipopolysaccharide (LPS). In parallel, PON2KO MPM also showed reduced anti-inflammatory responses in the basal state and also following M2 activation by IL-4. Moreover, the PON2-null MPM demonstrated enhanced phagocytosis and reactive oxygen species (ROS) production in the basal state and following M1 activation. The direct effect of PON2 was shown by transfecting human PON2 (hPON2) into PON2KO MPM. PON2 transfection attenuated the macrophages' response to M1 activation and enhanced M2 response. These PON2 effects were associated with attenuation of macrophages' abilities to phagocyte and to generate ROS. We conclude that PON2 promotes an M1 to M2 switch in macrophage phenotypes. PMID:26779262

  16. Role of the tumor suppressor ARF in macrophage polarization: Enhancement of the M2 phenotype in ARF-deficient mice.

    PubMed

    Herranz, Sandra; Través, Paqui G; Luque, Alfonso; Hortelano, Sonsoles

    2012-11-01

    The ARF locus is frequently inactivated in human cancer. The oncosuppressor ARF has indeed been described as a general sensor for different situation of cellular stress. We have previously demonstrated that ARF deficiency severely impairs inflammatory responses in vitro and in vivo, establishing a role for ARF in the regulation of innate immunity. The aim of the present work was to get further insights into the immune functions of ARF and to evaluate its possible contribution to the polarization of macrophages toward the M1 or M2 phenotype. Our results demonstrate that resting Arf(-/-) macrophages express high levels of Ym1 and Fizz-1, two typical markers of alternatively-activated macrophages (M2). Additionally, Arf(-/-) peritoneal macrophages showed an impaired response to lipopolysaccharide (a classical inducer of M1 polaryzation) and a reduced production of pro-inflammatory cytokines/chemokines. Moreover, upon stimulation with interleukin-4 (IL-4), an inducer of the M2 phenotype, well established M2 markers such as Fizz-1, Ym1 and arginase-1 were upregulated in Arf(-/-) as compared with wild type macrophages. Accordingly, the cytokine and chemokine profile associated with the M2 phenotype was significantly overexpressed in Arf(-/-) macrophages responding to IL-4. In addition, multiple pro-angiogenic factors such as VEGF and MMP-9 were also increased. In summary, these results indicate that ARF contributes to the polarization and functional plasticity of macrophages.

  17. Effect of Cocoa Polyphenolic Extract on Macrophage Polarization from Proinflammatory M1 to Anti-Inflammatory M2 State

    PubMed Central

    Dugo, Laura; Belluomo, Maria Giovanna; Fanali, Chiara; Russo, Marina; Cacciola, Francesco

    2017-01-01

    Polyphenols-rich cocoa has many beneficial effects on human health, such as anti-inflammatory effects. Macrophages function as control switches of the immune system, maintaining the balance between pro- and anti-inflammatory activities. We investigated the hypothesis that cocoa polyphenol extract may affect macrophage proinflammatory phenotype M1 by favoring an alternative M2 anti-inflammatory state on macrophages deriving from THP-1 cells. Chemical composition, total phenolic content, and antioxidant capacity of cocoa polyphenols extracted from roasted cocoa beans were determined. THP-1 cells were activated with both lipopolysaccharides and interferon-γ for M1 or with IL-4 for M2 switch, and specific cytokines were quantified. Cellular metabolism, through mitochondrial oxygen consumption, and ATP levels were evaluated. Here, we will show that cocoa polyphenolic extract attenuated in vitro inflammation decreasing M1 macrophage response as demonstrated by a significantly lowered secretion of proinflammatory cytokines. Moreover, treatment of M1 macrophages with cocoa polyphenols influences macrophage metabolism by promoting oxidative pathways, thus leading to a significant increase in O2 consumption by mitochondrial complexes as well as a higher production of ATP through oxidative phosphorylation. In conclusion, cocoa polyphenolic extract suppresses inflammation mediated by M1 phenotype and influences macrophage metabolism by promoting oxidative pathways and M2 polarization of active macrophages. PMID:28744339

  18. Effect of Cocoa Polyphenolic Extract on Macrophage Polarization from Proinflammatory M1 to Anti-Inflammatory M2 State.

    PubMed

    Dugo, Laura; Belluomo, Maria Giovanna; Fanali, Chiara; Russo, Marina; Cacciola, Francesco; Maccarrone, Mauro; Sardanelli, Anna Maria

    2017-01-01

    Polyphenols-rich cocoa has many beneficial effects on human health, such as anti-inflammatory effects. Macrophages function as control switches of the immune system, maintaining the balance between pro- and anti-inflammatory activities. We investigated the hypothesis that cocoa polyphenol extract may affect macrophage proinflammatory phenotype M1 by favoring an alternative M2 anti-inflammatory state on macrophages deriving from THP-1 cells. Chemical composition, total phenolic content, and antioxidant capacity of cocoa polyphenols extracted from roasted cocoa beans were determined. THP-1 cells were activated with both lipopolysaccharides and interferon-γ for M1 or with IL-4 for M2 switch, and specific cytokines were quantified. Cellular metabolism, through mitochondrial oxygen consumption, and ATP levels were evaluated. Here, we will show that cocoa polyphenolic extract attenuated in vitro inflammation decreasing M1 macrophage response as demonstrated by a significantly lowered secretion of proinflammatory cytokines. Moreover, treatment of M1 macrophages with cocoa polyphenols influences macrophage metabolism by promoting oxidative pathways, thus leading to a significant increase in O2 consumption by mitochondrial complexes as well as a higher production of ATP through oxidative phosphorylation. In conclusion, cocoa polyphenolic extract suppresses inflammation mediated by M1 phenotype and influences macrophage metabolism by promoting oxidative pathways and M2 polarization of active macrophages.

  19. Inhibition of Notch Signaling Attenuates Schistosomiasis Hepatic Fibrosis via Blocking Macrophage M2 Polarization

    PubMed Central

    Chen, Yixiong; Zheng, Shaojiang; Zheng, Liping; Weng, Zhihong

    2016-01-01

    Macrophages play a key role in the pathogenesis of liver granuloma and fibrosis in schistosomiasis. However, the underlying mechanisms have not been fully characterized. This study revealed that the macrophages infiltrating the liver tissues in a murine model of Schistosoma japonica infection exhibited M2 functional polarization, and Notch1/Jagged1 signaling was significantly upregulated in the M2 polarized macrophages in vivo and in vitro. Furthermore, the blockade of Notch signaling pathway by a γ–secretase inhibitor could reverse macrophage M2 polarization in vitro and alleviate liver granuloma and fibrosis in the murine model of schistosomiasis. These results implied that the Notch1/Jagged1 signaling-dependent M2 polarization of macrophages might play an important role in liver granuloma and fibrosis in schistosomiasis, and the inhibition of Notch1/Jagged1 signaling might provide a novel therapeutic approach to administrate patients with schistosomiasis. PMID:27875565

  20. Critical role of Trib1 in differentiation of tissue-resident M2-like macrophages.

    PubMed

    Satoh, Takashi; Kidoya, Hiroyasu; Naito, Hisamichi; Yamamoto, Masahiro; Takemura, Naoki; Nakagawa, Katsuhiro; Yoshioka, Yoshichika; Morii, Eiichi; Takakura, Nobuyuki; Takeuchi, Osamu; Akira, Shizuo

    2013-03-28

    Macrophages consist of at least two subgroups, M1 and M2 (refs 1-3). Whereas M1 macrophages are proinflammatory and have a central role in host defence against bacterial and viral infections, M2 macrophages are associated with responses to anti-inflammatory reactions, helminth infection, tissue remodelling, fibrosis and tumour progression. Trib1 is an adaptor protein involved in protein degradation by interacting with COP1 ubiquitin ligase. Genome-wide association studies in humans have implicated TRIB1 in lipid metabolism. Here we show that Trib1 is critical for the differentiation of F4/80(+)MR(+) tissue-resident macrophages--that share characteristics with M2 macrophages (which we term M2-like macrophages)--and eosinophils but not for the differentiation of M1 myeloid cells. Trib1 deficiency results in a severe reduction of M2-like macrophages in various organs, including bone marrow, spleen, lung and adipose tissues. Aberrant expression of C/EBPα in Trib1-deficient bone marrow cells is responsible for the defects in macrophage differentiation. Unexpectedly, mice lacking Trib1 in haematopoietic cells show diminished adipose tissue mass accompanied by evidence of increased lipolysis, even when fed a normal diet. Supplementation of M2-like macrophages rescues the pathophysiology, indicating that a lack of these macrophages is the cause of lipolysis. In response to a high-fat diet, mice lacking Trib1 in haematopoietic cells develop hypertriglyceridaemia and insulin resistance, together with increased proinflammatory cytokine gene induction. Collectively, these results demonstrate that Trib1 is critical for adipose tissue maintenance and suppression of metabolic disorders by controlling the differentiation of tissue-resident M2-like macrophages.

  1. Novel feedback loop between M2 macrophages/microglia and regulatory B cells in estrogen-protected EAE mice.

    PubMed

    Benedek, Gil; Zhang, Jun; Nguyen, Ha; Kent, Gail; Seifert, Hilary; Vandenbark, Arthur A; Offner, Halina

    2017-04-15

    Immunoregulatory sex hormones, including estrogen and estriol, may prevent relapses in multiple sclerosis during pregnancy. Our previous studies have demonstrated that regulatory B cells are crucial for estrogen-mediated protection against experimental autoimmune encephalomyelitis (EAE). Herein, we demonstrate an estrogen-dependent induction of alternatively activated (M2) macrophages/microglia that results in an increased frequency of regulatory B cells in the spinal cord of estrogen treated mice with EAE. We further demonstrate that cultured M2-polarized microglia promote the induction of regulatory B cells. Our study suggests that estrogen neuroprotection induces a regulatory feedback loop between M2 macrophages/microglia and regulatory B cells.

  2. M2 polarized macrophages induced by CSE promote proliferation, migration, and invasion of alveolar basal epithelial cells.

    PubMed

    Fu, Xiao; Shi, Hengfei; Qi, Yue; Zhang, Weiyun; Dong, Ping

    2015-09-01

    Cigarette smoking plays an important role in the genesis of lung cancer, and tumor-associated macrophages (TAMs) are believed to accelerate the process. We therefore sought to clarify the relationship between cigarette smoking, TAMs and tumorigenesis. We treated macrophages (THP-1) with cigarette smoke extract (CSE) and found that the mRNA levels of IL-6, IL-10, IL-12 and TNF-α decreased, while TGF-β mRNA levels increased. CSE significantly inhibited the phagocytic ability of macrophages, as assessed by flow cytometric analysis of FITC-dextran internalization. JAK2/STAT3 was significantly activated by CSE, as determined by Western blot analysis. When the scavenger receptor CD163, a specific marker of M2 macrophages, was analyzed by flow cytometry, its expression was significantly increased. After inducing M2 polarization of THP-1 cells, we co-cultured macrophages and alveolar basal epithelial cells (A549). The proliferation of A549 cells was detected by the MTT assay and cell cycle analysis, while their migration and invasion were detected by scratch wound assay and transwell assay. The results showed that the proliferation, migration and invasion of A549 cells were significantly promoted by M2 macrophages but were slightly inhibited by CSE. In conclusion, we demonstrated that macrophage M2 polarization induced by CSE promotes proliferation, migration, and invasion of alveolar basal epithelial cells.

  3. M2-like macrophages are responsible for collagen degradation through a mannose receptor–mediated pathway

    PubMed Central

    Madsen, Daniel H.; Leonard, Daniel; Masedunskas, Andrius; Moyer, Amanda; Jürgensen, Henrik Jessen; Peters, Diane E.; Amornphimoltham, Panomwat; Selvaraj, Arul; Yamada, Susan S.; Brenner, David A.; Burgdorf, Sven; Engelholm, Lars H.; Behrendt, Niels; Holmbeck, Kenn; Weigert, Roberto

    2013-01-01

    Tissue remodeling processes critically depend on the timely removal and remodeling of preexisting collagen scaffolds. Nevertheless, many aspects related to the turnover of this abundant extracellular matrix component in vivo are still incompletely understood. We therefore took advantage of recent advances in optical imaging to develop an assay to visualize collagen turnover in situ and identify cell types and molecules involved in this process. Collagen introduced into the dermis of mice underwent cellular endocytosis in a partially matrix metalloproteinase–dependent manner and was subsequently routed to lysosomes for complete degradation. Collagen uptake was predominantly executed by a quantitatively minor population of M2-like macrophages, whereas more abundant Col1a1-expressing fibroblasts and Cx3cr1-expressing macrophages internalized collagen at lower levels. Genetic ablation of the collagen receptors mannose receptor (Mrc1) and urokinase plasminogen activator receptor–associated protein (Endo180 and Mrc2) impaired this intracellular collagen degradation pathway. This study demonstrates the importance of receptor-mediated cellular uptake to collagen turnover in vivo and identifies a key role of M2-like macrophages in this process. PMID:24019537

  4. M2-like macrophages are responsible for collagen degradation through a mannose receptor-mediated pathway.

    PubMed

    Madsen, Daniel H; Leonard, Daniel; Masedunskas, Andrius; Moyer, Amanda; Jürgensen, Henrik Jessen; Peters, Diane E; Amornphimoltham, Panomwat; Selvaraj, Arul; Yamada, Susan S; Brenner, David A; Burgdorf, Sven; Engelholm, Lars H; Behrendt, Niels; Holmbeck, Kenn; Weigert, Roberto; Bugge, Thomas H

    2013-09-16

    Tissue remodeling processes critically depend on the timely removal and remodeling of preexisting collagen scaffolds. Nevertheless, many aspects related to the turnover of this abundant extracellular matrix component in vivo are still incompletely understood. We therefore took advantage of recent advances in optical imaging to develop an assay to visualize collagen turnover in situ and identify cell types and molecules involved in this process. Collagen introduced into the dermis of mice underwent cellular endocytosis in a partially matrix metalloproteinase-dependent manner and was subsequently routed to lysosomes for complete degradation. Collagen uptake was predominantly executed by a quantitatively minor population of M2-like macrophages, whereas more abundant Col1a1-expressing fibroblasts and Cx3cr1-expressing macrophages internalized collagen at lower levels. Genetic ablation of the collagen receptors mannose receptor (Mrc1) and urokinase plasminogen activator receptor-associated protein (Endo180 and Mrc2) impaired this intracellular collagen degradation pathway. This study demonstrates the importance of receptor-mediated cellular uptake to collagen turnover in vivo and identifies a key role of M2-like macrophages in this process.

  5. M2 macrophages or IL-33 treatment attenuate ongoing Mycobacterium tuberculosis infection

    PubMed Central

    Piñeros, A. R.; Campos, L. W.; Fonseca, D. M.; Bertolini, T. B.; Gembre, A. F.; Prado, R. Q.; Alves-Filho, J. C.; Ramos, S. G.; Russo, M.; Bonato, V. L. D.

    2017-01-01

    The protective effects of mycobacterial infections on lung allergy are well documented. However, the inverse relationship between tuberculosis and type 2 immunity is still elusive. Although type 1 immunity is essential to protection against Mycobacterium tuberculosis it might be also detrimental to the host due to the induction of extensive tissue damage. Here, we determined whether lung type 2 immunity induced by allergen sensitization and challenge could affect the outcome of M. tuberculosis infection. We used two different protocols in which sensitization and allergen challenge were performed before or after M. tuberculosis infection. We found an increased resistance to M. tuberculosis only when allergen exposure was given after, but not before infection. Infected mice exposed to allergen exhibited lower bacterial load and cellular infiltrates in the lungs. Enhanced resistance to infection after allergen challenge was associated with increased gene expression of alternatively activated macrophages (M2 macrophages) and IL-33 levels. Accordingly, either adoptive transfer of M2 macrophages or systemic IL-33 treatment was effective in attenuating M. tuberculosis infection. Notably, the enhanced resistance induced by allergen exposure was dependent on IL-33 receptor ST2. Our work indicates that IL-33 might be an alternative therapeutic treatment for severe tuberculosis. PMID:28128217

  6. Pomegranate juice polyphenols induce a phenotypic switch in macrophage polarization favoring a M2 anti-inflammatory state.

    PubMed

    Aharoni, Saar; Lati, Yoni; Aviram, Michael; Fuhrman, Bianca

    2015-01-01

    It was documented that pomegranate has anti-inflammatory effects. In this study, we investigated a direct effect of pomegranate juice (PJ) and its polyphenols on macrophage inflammatory phenotype. In vitro, PJ and its major polyphenols dose-dependently attenuated macrophage response to M1 proinflammatory activation in J774.A1 macrophage-like cell line. This was evidenced by a significant decrease in TNFα and IL-6 secretion in response to stimulation by IFNγ and Lipopolysaccharide. In addition, PJ and punicalagin dose-dependently promoted the macrophages toward a M2 anti-inflammatory phenotype, as determined by a significant increase in the spontaneous secretion of IL-10. In mice, supplementation with dietary PJ substantially inhibited the M2 to M1 macrophage phenotypic shift associated with age, toward a favorable anti-inflammatory M2 phenotype. This effect was also reflected in the mice atherosclerotic plaques, as evaluated by the distinct expression of arginase isoforms. PJ consumption inhibited the increment of arginase II (Arg II, M1) mRNA expression during aging, and maintained the levels of Arg I (M2) expression similar to those in young mice aorta. This study demonstrates, for the first time, that pomegranate polyphenols directly suppress macrophage inflammatory responses and promote M1 to M2 switch in macrophage phenotype. Furthermore, this study indicates that PJ consumption may inhibit the progressive proinflammatory state in the aorta along atherosclerosis development with aging, due to a switch in macrophage phenotype from proinflammatory M1 to anti-inflammatory M2.

  7. Metabolism Supports Macrophage Activation

    PubMed Central

    Langston, P. Kent; Shibata, Munehiko; Horng, Tiffany

    2017-01-01

    Macrophages are found in most tissues of the body, where they have tissue- and context-dependent roles in maintaining homeostasis as well as coordinating adaptive responses to various stresses. Their capacity for specialized functions is controlled by polarizing signals, which activate macrophages by upregulating transcriptional programs that encode distinct effector functions. An important conceptual advance in the field of macrophage biology, emerging from recent studies, is that macrophage activation is critically supported by metabolic shifts. Metabolic shifts fuel multiple aspects of macrophage activation, and preventing these shifts impairs appropriate activation. These findings raise the exciting possibility that macrophage functions in various contexts could be regulated by manipulating their metabolism. Here, we review the rapidly evolving field of macrophage metabolism, discussing how polarizing signals trigger metabolic shifts and how these shifts enable appropriate activation and sustain effector activities. We also discuss recent studies indicating that the mitochondria are central hubs in inflammatory macrophage activation. PMID:28197151

  8. Embryonic stem cell-derived M2-like macrophages delay cutaneous wound healing.

    PubMed

    Dreymueller, Daniela; Denecke, Bernd; Ludwig, Andreas; Jahnen-Dechent, Willi

    2013-01-01

    In adults, repair of deeply injured skin wounds results in the formation of scar tissue, whereas in embryos wounds heal almost scar-free. Macrophages are important mediators of wound healing and secrete cytokines and tissue remodeling enzymes. In contrast to host defense mediated by inflammatory M1 macrophages, wound healing and tissue repair involve regulatory M2/M2-like macrophages. Embryonic/fetal macrophages are M2-like, and this may promote scar-free wound healing. In the present study, we asked whether atopical application of ex vivo generated, embryonic stem cell-derived macrophages (ESDM) improve wound healing in mice. ESDM were tested side by side with bone marrow-derived macrophages (BMDM). Compared to BMDM, ESDM resembled a less inflammatory and more M2-like macrophage subtype as indicated by their reduced responsiveness to lipopolysaccharide, reduced expression of Toll-like receptors, and reduced bacterial phagocytosis. Despite this anti-inflammatory phenotype in cell culture, ESDM prolonged the healing of deep skin wounds even more than BMDM. Healed wounds had more scar formation compared to wounds receiving BMDM or cell-free treatment. Our data indicate that atopical application of ex vivo generated macrophages is not a suitable cell therapy of dermal wounds. © 2012 by the Wound Healing Society.

  9. Differences in forward angular light scattering distributions between M1 and M2 macrophages

    NASA Astrophysics Data System (ADS)

    Halaney, David L.; Zahedivash, Aydin; Phipps, Jennifer E.; Wang, Tianyi; Dwelle, Jordan; Saux, Claude Jourdan Le; Asmis, Reto; Milner, Thomas E.; Feldman, Marc D.

    2015-11-01

    The ability to distinguish macrophage subtypes noninvasively could have diagnostic potential in cancer, atherosclerosis, and diabetes, where polarized M1 and M2 macrophages play critical and often opposing roles. Current methods to distinguish macrophage subtypes rely on tissue biopsy. Optical imaging techniques based on light scattering are of interest as they can be translated into biopsy-free strategies. Because mitochondria are relatively strong subcellular light scattering centers, and M2 macrophages are known to have enhanced mitochondrial biogenesis compared to M1, we hypothesized that M1 and M2 macrophages may have different angular light scattering profiles. To test this, we developed an in vitro angle-resolved forward light scattering measurement system. We found that M1 and M2 macrophage monolayers scatter relatively unequal amounts of light in the forward direction between 1.6 deg and 3.2 deg with M2 forward scattering significantly more light than M1 at increasing angles. The ratio of forward scattering can be used to identify the polarization state of macrophage populations in culture.

  10. Differences in forward angular light scattering distributions between M1 and M2 macrophages

    PubMed Central

    Halaney, David L.; Zahedivash, Aydin; Phipps, Jennifer E.; Wang, Tianyi; Dwelle, Jordan; Saux, Claude Jourdan Le; Asmis, Reto; Milner, Thomas E.; Feldman, Marc D.

    2015-01-01

    Abstract. The ability to distinguish macrophage subtypes noninvasively could have diagnostic potential in cancer, atherosclerosis, and diabetes, where polarized M1 and M2 macrophages play critical and often opposing roles. Current methods to distinguish macrophage subtypes rely on tissue biopsy. Optical imaging techniques based on light scattering are of interest as they can be translated into biopsy-free strategies. Because mitochondria are relatively strong subcellular light scattering centers, and M2 macrophages are known to have enhanced mitochondrial biogenesis compared to M1, we hypothesized that M1 and M2 macrophages may have different angular light scattering profiles. To test this, we developed an in vitro angle-resolved forward light scattering measurement system. We found that M1 and M2 macrophage monolayers scatter relatively unequal amounts of light in the forward direction between 1.6 deg and 3.2 deg with M2 forward scattering significantly more light than M1 at increasing angles. The ratio of forward scattering can be used to identify the polarization state of macrophage populations in culture. PMID:26538329

  11. Differences in forward angular light scattering distributions between M1 and M2 macrophages.

    PubMed

    Halaney, David L; Zahedivash, Aydin; Phipps, Jennifer E; Wang, Tianyi; Dwelle, Jordan; Saux, Claude Jourdan Le; Asmis, Reto; Milner, Thomas E; Feldman, Marc D

    2015-11-01

    The ability to distinguish macrophage subtypes noninvasively could have diagnostic potential in cancer, atherosclerosis, and diabetes, where polarized M1 and M2 macrophages play critical and often opposing roles. Current methods to distinguish macrophage subtypes rely on tissue biopsy. Optical imaging techniques based on light scattering are of interest as they can be translated into biopsy-free strategies. Because mitochondria are relatively strong subcellular light scattering centers, and M2 macrophages are known to have enhanced mitochondrial biogenesis compared to M1, we hypothesized that M1 and M2 macrophages may have different angular light scattering profiles. To test this, we developed an in vitro angle-resolved forward light scattering measurement system. We found that M1 and M2 macrophage monolayers scatter relatively unequal amounts of light in the forward direction between 1.6 deg and 3.2 deg with M2 forward scattering significantly more light than M1 at increasing angles. The ratio of forward scattering can be used to identify the polarization state of macrophage populations in culture.

  12. Myeloid PTEN deficiency protects livers from ischemia reperfusion injury by facilitating M2 macrophage differentiation.

    PubMed

    Yue, Shi; Rao, Jianhua; Zhu, Jianjun; Busuttil, Ronald W; Kupiec-Weglinski, Jerzy W; Lu, Ling; Wang, Xuehao; Zhai, Yuan

    2014-06-01

    Although the role of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in regulating cell proliferation is well established, its function in immune responses remains to be fully appreciated. In the current study, we analyzed myeloid-specific PTEN function in regulating tissue inflammatory immune response in a murine liver partial warm ischemia model. Myeloid-specific PTEN knockout (KO) resulted in liver protection from ischemia reperfusion injury (IRI) by deviating the local innate immune response against ischemia reperfusion toward the regulatory type: expression of proinflammatory genes was selectively decreased and anti-inflammatory IL-10 was simultaneously increased in ischemia reperfusion livers of PTEN KO mice compared with those of wild-type (WT) mice. PI3K inhibitor and IL-10-neutralizing Abs, but not exogenous LPS, recreated liver IRI in these KO mice. At the cellular level, Kupffer cells and peritoneal macrophages isolated from KO mice expressed higher levels of M2 markers and produced lower TNF-α and higher IL-10 in response to TLR ligands than did their WT counterparts. They had enhanced Stat3- and Stat6-signaling pathway activation, but diminished Stat1-signaling pathway activation, in response to TLR4 stimulation. Inactivation of Kupffer cells by gadolinium chloride enhanced proinflammatory immune activation and increased IRI in livers of myeloid PTEN KO mice. Thus, myeloid PTEN deficiency protects livers from IRI by facilitating M2 macrophage differentiation. Copyright © 2014 by The American Association of Immunologists, Inc.

  13. Adoptive transfer of M2 macrophages reduces neuropathic pain via opioid peptides.

    PubMed

    Pannell, Maria; Labuz, Dominika; Celik, Melih Ö; Keye, Jacqueline; Batra, Arvind; Siegmund, Britta; Machelska, Halina

    2016-10-07

    During the inflammation which occurs following nerve damage, macrophages are recruited to the site of injury. Phenotypic diversity is a hallmark of the macrophage lineage and includes pro-inflammatory M1 and anti-inflammatory M2 populations. Our aim in this study was to investigate the ability of polarized M0, M1, and M2 macrophages to secrete opioid peptides and to examine their relative contribution to the modulation of neuropathic pain. Mouse bone marrow-derived cells were cultured as unstimulated M0 macrophages or were stimulated into an M1 phenotype using lipopolysaccharide and interferon-γ or into an M2 phenotype using interleukin-4. The macrophage phenotypes were verified using flow cytometry for surface marker analysis and cytokine bead array for cytokine profile assessment. Opioid peptide levels were measured by radioimmunoassay and enzyme immunoassay. As a model of neuropathic pain, a chronic constriction injury (CCI) of the sciatic nerve was employed. Polarized M0, M1, and M2 macrophages (5 × 10(5) cells) were injected perineurally twice, on days 14 and 15 following CCI or sham surgery. Mechanical and heat sensitivity were measured using the von Frey and Hargreaves tests, respectively. To track the injected macrophages, we also transferred fluorescently stained polarized cells and analyzed the surface marker profile of endogenous and injected cells in the nerves ex vivo. Compared to M0 and M1 cells, M2 macrophages contained and released higher amounts of opioid peptides, including Met-enkephalin, dynorphin A (1-17), and β-endorphin. M2 cells transferred perineurally at the nerve injury site reduced mechanical, but not heat hypersensitivity following the second injection. The analgesic effect was reversed by the perineurally applied opioid receptor antagonist naloxone methiodide. M2 cells did not affect sensitivity following sham surgery. Neither M0 nor M1 cells altered mechanical and heat sensitivity in CCI or sham-operated animals. Tracing the

  14. Selective M2 Macrophage Depletion Leads to Prolonged Inflammation in Surgical Wounds.

    PubMed

    Klinkert, Kerstin; Whelan, Derek; Clover, Anthony J P; Leblond, Anne-Laure; Kumar, Arun H S; Caplice, Noel M

    2017-01-01

    A prolonged inflammatory phase is seen in aberrant wound healing and in chronic wounds. Macrophages are central to wound healing. Distinct macrophage subtypes have differing roles both in initial inflammation and in later tissue repair. Broadly, these cells can be divided into M1 and M2 macrophages. M2 macrophage proliferation and differentiation is regulated by colony-stimulating factor 1 (CSF-1) signalling and can be blocked by GW2580, a competitive cFMS kinase inhibitor, thereby allowing for analysis of the effect of M2 blockade on progression of surgical wounds. Macrophage Fas-induced apoptosis (MaFIA) transgenic mice with a macrophage-specific promoter used to express green fluorescent protein (GFP) were used to allow for cell tracking. The animals were treated by oral gavage with GW2580. Surgical wounds were created and harvested after 2 weeks for analysis. GW2580-treated mice had significantly more GFP+ cells in the surgical scar than vehicle-treated animals (GW2580, 68.0 ± 3.1%; vehicle, 42.8 ± 1.7%; p < 0.001), and GW2580 treatment depleted CD206+ M2 macrophages in the scar (GW2580, 1.4%; vehicle, 19.3%; p < 0.001). Treated animals showed significantly higher numbers of neutrophils (vehicle, 18.0%; GW2580, 51.3%; p < 0.01) and M1 macrophages (vehicle, 3.8%; GW2580, 12.8%; p < 0.01) in the scar compared to vehicle-treated animals. The total collagen content in the area of the scar was decreased in animals treated with GW2580 as compared to those treated with vehicle alone (GW2580, 67.1%; vehicle, 79.9%; p < 0.005). Depletion of M2 macrophages in surgical wounds via CSF-1 signalling blockade leads to persistent inflammation, with an increase in neutrophils and M1 macrophages and attenuated collagen deposition. © 2017 S. Karger AG, Basel.

  15. Dietary oleic acid increases m2 macrophages in the mesenteric adipose tissue.

    PubMed

    Camell, Christina; Smith, C Wayne

    2013-01-01

    Several studies have implicated fatty-acids as inflammatory regulators, suggesting that there may be a direct role for common dietary fatty-acids in regulating innate immune cells. In humans, a single high-fat meal increases systemic cytokines and leukocytes. In mice, short term high-fat feeding increases adipose tissue (AT) leukocytes and alters the inflammatory profile of AT macrophages. We have seen that short term high fat feeding to C57BL/6J male mice increases palmitic and oleic acid within AT depots, but oleic acid increase is highest in the mesenteric AT (MAT). In vitro, oleic acid increases M2 macrophage markers (CD206, MGL1, and ARG1) in a murine macrophage cell line, while addition of palmitic acid is able to inhibit that increase. Three day supplementation of a chow diet, with oleic acid, induced an increase in M2 macrophage markers in the MAT, but not in the epididymal AT. We tested whether increases in M2 macrophages occur during short term ad lib feeding of a high fat diet, containing oleic acid. Experiments revealed two distinct populations of macrophages were altered by a three day high milk-fat diet. One population, phenotypically intermediate for F4/80, showed diet-induced increases in CD206, an anti-inflammatory marker characteristic of M2 macrophages intrinsic to the AT. Evidence for a second population, phenotypically F4/80(HI)CD11b(HI) macrophages, showed increased association with the MAT following short term feeding that is dependent on the adhesion molecule, ICAM-1. Collectively, we have shown that short term feeding of a high-fat diet changes two population of macrophages, and that dietary oleic acid is responsible for increases in M2 macrophage polarization.

  16. Myeloid PTEN Deficiency Protects Livers from Ischemia Reperfusion Injury by Facilitating M2 Macrophage Differentiation

    PubMed Central

    Zhu, Jianjun; Busuttil, Ronald W.; Kupiec-Weglinski, Jerzy W.; Lu, Ling; Wang, Xuehao; Zhai, Yuan

    2014-01-01

    Although roles of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in regulating cell proliferation are well established, its function in immune responses remains to be fully appreciated. In the current study, we analyzed myeloid-specific PTEN function in regulating tissue inflammatory immune response in a murine liver partial warm ischemia model. Myeloid-specific PTEN knock-out (KO) resulted in liver protections from ischemia reperfusion injury (IRI) by deviating local innate immune response against IR toward the regulatory type: expressions of pro-inflammatory genes were selectively decreased and anti-inflammatory IL-10 was simultaneously increased in IR livers of PTEN KO mice, as compared with those of wild-type (WT) mice. PI3 kinase inhibitor and IL-10 neutralizing Abs, but not exogenous LPS, recreated liver IRI in these KO mice. At the cellular level, Kupffer cells, as well as peritoneal macrophages, isolated from KO mice expressed higher levels of M2 markers, and produced lower TNF-α and higher IL-10 in response to TLR ligands, than their WT counterparts. They had enhanced Stat3 and Stat6, but diminished Stat1 signaling pathway activations in response to TLR4 stimulation. Inactivation of KCs by gadolinium chloride enhanced pro-inflammatory immune activation and increased IRI in livers of myeloid PTEN KO mice. Thus, myeloid PTEN deficiency protects livers from IRI by facilitating M2 macrophage differentiation. PMID:24771857

  17. Modulators of the Balance between M1 and M2 Macrophages during Pregnancy

    PubMed Central

    Zhang, Yong-Hong; He, Ming; Wang, Yan; Liao, Ai-Hua

    2017-01-01

    Macrophages are a subset of mononuclear phagocytes of the innate immune system with high plasticity and heterogeneity. At the maternal–fetal interface, macrophages are present in all stages of pregnancy and involved in a variety of activities, including regulation of immune cell activities, decidualization, placental cell invasion, angiogenesis, parturition, and postpartum uterine involution. The activation state and function of uterine–placental macrophages are largely dependent on the local tissue microenvironment. However, disruption of the uterine microenvironment can have profound effects on macrophage activity and subsequently impact pregnancy outcome. Thus, appropriately and timely regulated macrophage polarization has been considered a key determinant of successful pregnancy. Targeting macrophage polarization might be an efficient strategy for maintaining maternal–fetal immune homeostasis and a normal pregnancy. Here, we will review the latest findings regarding the modulators regulating macrophage polarization in healthy pregnancies and pregnancy complications, which might provide a basis for macrophage-centered therapeutic strategies. PMID:28232836

  18. Melanoma-initiating cells exploit M2 macrophage TGFβ and arginase pathway for survival and proliferation

    PubMed Central

    Tham, Muly; Tan, Kar Wai; Keeble, Jo; Wang, Xiaojie; Hubert, Sandra; Barron, Luke; Tan, Nguan Soon; Kato, Masashi; Prevost-Blondel, Armelle; Angeli, Veronique; Abastado, Jean-Pierre

    2014-01-01

    M2 macrophages promote tumor growth and metastasis, but their interactions with specific tumor cell populations are poorly characterized. Using a mouse model of spontaneous melanoma, we showed that CD34− but not CD34+ tumor-initiating cells (TICs) depend on M2 macrophages for survival and proliferation. Tumor-associated macrophages (TAMs) and macrophage-conditioned media protected CD34− TICs from chemotherapy in vitro. In vivo, while inhibition of CD115 suppressed the macrophage-dependent CD34− TIC population, chemotherapy accelerated its development. The ability of TICs to respond to TAMs was acquired during melanoma progression and immediately preceded a surge in metastatic outgrowth. TAM-derived transforming growth factor-β (TGFβ) and polyamines produced via the Arginase pathway were critical for stimulation of TICs and synergized to promote their growth. PMID:25294815

  19. Leptin promotes migration and invasion of breast cancer cells by stimulating IL-8 production in M2 macrophages

    PubMed Central

    Wang, Lin; Wang, Hong; Pang, Xueli; Li, Kuangfa; Dang, Weiqi; Tang, Hao; Wei, Lan; Su, Min; Tang, Cuiping; Chen, Tingmei

    2016-01-01

    This study aims to investigate the mechanisms underlying leptin-mediated crosstalk between tumor-associated macrophages (M2 macrophages) and breast cancer cells. THP1 human leukemic monocytes were induced to differentiate into M2 macrophages by PMA (100 nM) and IL-4 (20 ng/mL). Quantitative RT-PCR and Western blot revealed that leptin (100 nM) significantly increased the expression of leptin receptor (ObR) in the M2 macrophages (P < 0.01) and stimulated interleukin (IL)-8 expression in the M2 macrophages, mouse macrophage cells RAW264.7, and primary mouse peritoneal macrophages in a dose- and time-dependent manner. Leptin-induced IL-8 production was sensitive to the ERK inhibitor PD980590 (10 μmol/L), p38 MAPK inhibitor SB203580 (20 μmol/L), and anti-ObR neutralizing antibody (4 μg/mL). Leptin (100 ng/mL) substantially increased the phosphorylation of p38 and ERK1/2. Thus, leptin may induce IL-8 production in M2 macrophages by interacting with ObR to activate the p38 and ERK signaling pathways. Scratch and transwell chamber assay showed that both recombinant IL-8 and leptin-induced M2 macrophage-derived IL-8 promoted the migration and invasion of human breast cancer cells MCF7 and MDA-MB-231 (All P < 0.01). In a nude mice xenograft model of breast cancer (n = 5 per group), injection of leptin (0.1 μg/g) dramatically increased tumor volume and mass, reduced survival, exacerbated pulmonary metastasis, and elevated IL-8 and Ki67 expression in the tumor tissue (All P < 0.05) compared with PBS injection. Depletion of mouse macrophage by Clophosome®-clodronate liposome and injection of anti-mouse IL-8 neutralizing antibodies in the xenograft tumor significantly attenuated those leptin-mediated stimulations (All P < 0.05). These findings indicate that leptin may promote tumor growth and metastasis by stimulating IL-8 production in tumor-associated macrophage. PMID:27588409

  20. M2b macrophage polarization accompanied with reduction of long noncoding RNA GAS5.

    PubMed

    Ito, Ichiaki; Asai, Akira; Suzuki, Sumihiro; Kobayashi, Makiko; Suzuki, Fujio

    2017-11-04

    Macrophages (Mϕ) are highly plastic and change their functional phenotypes depending on microenvironmental signals. Recent studies have shown that microRNAs are involved in the polarization of Mϕ. In this study, we demonstrated that the phenotype of M2bMϕ [CCL1(+) IL-10(+) LIGHT(+)] switches to other phenotypes with interchangeability attained through the increased expression of growth arrest-specific 5 RNA (GAS5 RNA), a long noncoding RNA. GAS5 RNA has been described as a silencer of the CCL1 gene. Various phenotypes of Mϕ were prepared from bone marrow-derived Mϕ (BMDMϕ) after stimulation with IFNγ [M(IFNγ)/M1Mϕ], IL-4 [M(IL-4)/M2aMϕ], LPS and immobilized IgG [M(LPS + IC)/M2bMϕ], and IL-10 [M(IL-10)/M2cMϕ]. BMDMϕ cultured with medium [M(no)/quiescent Mϕ] were used as a control. As compared to Μ(no), M(IFNγ), M(IL-4) and M(IL-10), the reduced level of GAS5 RNA was shown in M(LPS + IC). CCL1 and LIGHT mRNAs (typical biomarkers of M2bMϕ) were not expressed by M(LPS + IC) transduced with a GAS5 gene using lentiviral vector. The reduction of GAS5 RNA in M(LPS + IC) was mediated by the activation of nonsense-mediated RNA decay (NMD) pathway. BMDMϕ overexpressed with GAS5 RNA after GAS5 gene transduction did not polarize to M2bMϕ even though they were stimulated with LPS and IC in combination. These results indicate that the reduction of GAS5 RNA influenced by the NMD pathway activation leads to the Mϕ polarization stimulated with LPS and IC in combination. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Clinical experience with autologous M2 macrophages in children with severe cerebral palsy.

    PubMed

    Chernykh, Elena R; Kafanova, Marina Yu; Shevela, Ekaterina Ya; Sirota, Sergei I; Adonina, Elena I; Sakhno, Lyudmila V; Ostanin, Alexander A; Kozlov, Vladimir V

    2014-01-01

    Stem cell-based therapy is considered to be a new approach for the treatment of cerebral palsy (CP). Given the potent anti-inflammatory activity and high regenerative potential of M2 macrophages, these cells may be an alternative source for cell transplantation. To evaluate the safety and efficacy of autologous M2 macrophages, we conducted a pilot clinical trial in 21 children with severe CP. The primary outcome measure was safety, which included assessment of mortality of any cause, immediate adverse reactions, and serious adverse effects and comorbidities during 5-year follow-up. The secondary outcome measure was functional improvement in Gross Motor Function Measure (66-item GMFM) test, Peabody Developmental Motor Scale-Fine Motor (PDMS-FM) test, Ashworth scale, MRC scale, and an easy-to-understand questionnaire for evaluation of cognitive functions in our modification. Intradural injection of M2 cells (in mean dose of 0.8 × 10(6)/kg) into the lumbar spinal area did not induce any serious adverse events. No cases of mortality, psychomotor worsening, exacerbation of seizures, and long-term comorbidities, including tumors, were observed during a 5-year follow-up. After 3 months, GMFM score increased from 13.7 ± 7.8 to 58.6 ± 14.6, PDMS-FM score improved from 0.76 ± 0.42 to 5.05 ± 0.97, and the Ashworth score decreased from 3.8 ± 0.21 to 3.3 ± 0.24. Along with gross and fine motor function enhancement, an improvement of cognitive activity (from 1.62 ± 0.41 to 4.05 ± 0.64, according to questionnaire assessment) and reduction of seizure syndrome were registered as well. The neurological improvements did not diminish during the 5-year follow-up period. The data obtained suggest that cell therapy based on M2 macrophages is safe, does not induce early adverse effects and long-term comorbidities, and is accompanied with a significant improvement of motor and cognitive activities in severe CP patients. This manuscript is

  2. The pentacyclic triterpene Lupeol switches M1 macrophages to M2 and ameliorates experimental inflammatory bowel disease.

    PubMed

    Zhu, Yeshan; Li, Xueqing; Chen, Jianquan; Chen, Tongjun; Shi, Zhimin; Lei, Miaona; Zhang, Yanjun; Bai, Pengfei; Li, Yifang; Fei, Xuan

    2016-01-01

    Inflammatory bowel disease (IBD), including ulcerative colitis and Crohn's disease, is a chronic inflammatory disease in the lower gastrointestinal tract. Mounting evidence suggests that the predominance of the classically activated (M1) macrophages versus the alternatively activated (M2) macrophages plays a role in the progression of IBD. Thus, agents able to shift pro-inflammatory M1 macrophages to anti-inflammatory M2 macrophages may be beneficial to IBD. The pentacyclic triterpene Lup-20(29)-en-3β-ol (Lupeol), a potent anti-inflammatory natural product, has been shown to inhibit pro-inflammatory cytokine production, suggesting it is potentially able to modulate macrophage polarization, thereby beneficial to IBD. CD4(+) monocytes were differentiated to M1 or M2 macrophages, which were cocultured with epithelial cell lines, T84 and Caco-2, in the absence or presence of Lupeol (10μM). Experimental colitis was induced with dextran sodium sulfate (DSS), with or without oral administration of Lupeol (50mg/kg, q.d.). Cytokines were measured with Luminex kits. M1/M2 genes were measured with real-time polymerase chain reaction. Macrophage phenotypes were defined by measuring M1 and M2 markers with confocal microscopy. Proteins were measured with Western blotting, while cell surface markers were measured with confocal microscopy or flow cytometry. Histology was evaluated with H&E staining. Treatment of M1 macrophages with Lupeol resulted in a marked decrease in the production of pro-inflammatory cytokines, including IL-12, IL6, IL-1β and TNFα, and a marked increase in the production of IL-10, an anti-inflammatory cytokine. This was associated with a down-regulation of CD86, a typical marker of M1 macrophages, and an up-regulation of CD206, a typical M2 macrophage marker. IRF5, a transcription factor that is critically involved in M1 polarization, was down-regulated in M1 macrophages after being incubated with Lupeol, associated with a marked decrease in the

  3. Impact of Detachment Methods on M2 Macrophage Phenotype and Function.

    PubMed

    Chen, Shaodong; So, Edward C; Strome, Scott E; Zhang, Xiaoyu

    2015-11-01

    The methods of cell detachment influence phenotype and function of human macrophages cultured in vitro. However, comparative studies defining the influence of cell detachment techniques on secondary characterization of M1 or M2 polarized macrophages are largely absent from the literature. In this study we evaluated the impact of trypsin, accutase, EDTA, PBS, and cell scraping on: A. cell recovery, B. phenotype and C. function of in vitro polarized macrophages. Our data demonstrate that while exposure to trypsin or accutase yields highly efficient recovery of viable cells, such chemical cleavage results in loss of select M2 cell surface markers with correlative changes in cell function. In contrast, phenotype and function are maintained following detachment by EDTA on ice. Our data suggest that seemingly "trivial" changes in methodologies for macrophage detachment induce both variable and profound changes on cell phenotype and function which can dramatically impact the results of polarization experiments. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. 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.

  5. Culture supernatants of cervical cancer cells induce an M2 phenotypic profile in THP-1 macrophages.

    PubMed

    Pedraza-Brindis, Eliza Julia; Sánchez-Reyes, Karina; Hernández-Flores, Georgina; Bravo-Cuellar, Alejandro; Jave-Suárez, Luis Felipe; Aguilar-Lemarroy, Adriana; Gómez-Lomelí, Paulina; López-López, Brenda Anahí; Ortiz-Lazareno, Pablo Cesar

    2016-12-01

    Patients with cervical cancer (CxCa) typically present an infiltrate of tumor-associated macrophages, which is associated with a poor prognosis. We found that CxCa cell lines (HeLa, SiHa, and C-33A) secreted factors involved in regulating tumor growth including IL-6, IL-4, PDGFAA, HGF, VEGF, ANG-2, and TGF-β3. We assessed the effects of culture supernatants from these cell lines on macrophages derived from the THP-1 cell line. Macrophages treated with culture supernatants from CxCa cells developed an M2-like phenotype with expression of CD163, low nitric oxide release, and high secretion of IL-6, PDGFAA, HGF, ANG-2, and VEGF. The macrophages continued to produce PDGFAA, PDGFBB, and VEGF 48h after the CxCa cell culture supernatants were removed. The induction of M2 macrophages in vivo favors tumor growth, angiogenesis, tissue remodeling, and metastasis. These results demonstrated that factors secreted by CxCa cells induced a stable M2 phenotype in THP-1 macrophages. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Neisseria gonorrhoeae Modulates Immunity by Polarizing Human Macrophages to a M2 Profile

    PubMed Central

    Ortiz, María Carolina; Lefimil, Claudia; Rodas, Paula I.; Vernal, Rolando; Lopez, Mercedes; Acuña-Castillo, Claudio; Imarai, Mónica; Escobar, Alejandro

    2015-01-01

    Current data suggest that Neisseria gonorrhoeae is able to suppress the protective immune response at different levels, such as B and T lymphocytes and antigen-presenting cells. The present report is focused on gonococcus evasion mechanism on macrophages (MФ) and its impact in the subsequent immune response. In response to various signals MФ may undergo classical-M1 (M1-MФ) or alternative-M2 (M2-MФ) activation. Until now there are no reports of the gonococcus effects on human MФ polarization. We assessed the phagocytic ability of monocyte-derived MФ (MDM) upon gonococcal infection by immunofluorescence and gentamicin protection experiments. Then, we evaluated cytokine profile and M1/M2 specific-surface markers on MФ challenged with N. gonorrhoeae and their proliferative effect on T cells. Our findings lead us to suggest N. gonorrhoeae stimulates a M2-MФ phenotype in which some of the M2b and none of the M1-MФ-associated markers are induced. Interestingly, N. gonorrhoeae exposure leads to upregulation of a Programmed Death Ligand 1 (PD-L1), widely known as an immunosuppressive molecule. Moreover, functional results showed that N. gonorrhoeae-treated MФ are unable to induce proliferation of human T-cells, suggesting a more likely regulatory phenotype. Taken together, our data show that N. gonorroheae interferes with MФ polarization. This study has important implications for understanding the mechanisms of clearance versus long-term persistence of N. gonorroheae infection and might be applicable for the development of new therapeutic strategies. PMID:26125939

  7. Ixmyelocel-T, an expanded multicellular therapy, contains a unique population of M2-like macrophages

    PubMed Central

    2013-01-01

    Introduction M2 macrophages promote tissue repair and regeneration through various mechanisms including immunomodulation and scavenging of tissue debris. Delivering increased numbers of these cells to ischemic tissues may limit tissue injury and promote repair. Ixmyelocel-T is an expanded, autologous multicellular therapy cultured from bone-marrow mononuclear cells (BMMNCs). The purpose of this study was to characterize further a unique expanded population of M2-like macrophages, generated in ixmyelocel-T therapy. Methods Approximately 50 ml of whole bone marrow was obtained from healthy donors and shipped overnight. BMMNCs were produced by using density-gradient separation and cultured for approximately 12 days to generate ixmyelocel-T. CD14+ cells were isolated from ixmyelocel-T with positive selection for analysis. Cell-surface phenotype was examined with flow cytometry and immunofluorescence, and expression of cytokines and chemokines was analyzed with enzyme-linked immunosorbent assay (ELISA). Quantitative real-time PCR was used to analyze expression of genes in BMMNCs, ixmyelocel-T, the CD14+ population from ixmyelocel-T, and M1 and M2 macrophages. Ixmyelocel-T was cultured with apoptotic BMMNCs, and then visualized under fluorescence microscopy to assess efferocytosis. Results Macrophages in ixmyelocel-T therapy expressed surface markers of M2 macrophages, CD206, and CD163. These cells were also found to express several M2 markers, and few to no M1 markers. After stimulation with lipopolysaccharide (LPS), they showed minimal secretion of the proinflammatory cytokines interleukin-12 (IL-12) and tumor necrosis factor alpha (TNF-α) compared with M1 and M2 macrophages. Ixmyelocel-T macrophages efficiently ingested apoptotic BMMNCs. Conclusions Ixmyelocel-T therapy contains a unique population of M2-like macrophages that are characterized by expression of M2 markers, decreased secretion of proinflammatory cytokines after inflammatory stimuli, and efficient

  8. M2-like macrophage polarization in high lactic acid-producing head and neck cancer.

    PubMed

    Ohashi, Toshimitsu; Aoki, Mitsuhiro; Tomita, Hiroyuki; Akazawa, Takashi; Sato, Katsuya; Kuze, Bunya; Mizuta, Keisuke; Hara, Akira; Nagaoka, Hitoshi; Inoue, Norimitsu; Ito, Yatsuji

    2017-06-01

    Reprogramming of glucose metabolism in tumor cells is referred to as the Warburg effect and results in increased lactic acid secretion into the tumor microenvironment. We have previously shown that lactic acid has important roles as a pro-inflammatory and immunosuppressive mediator and promotes tumor progression. In this study, we examined the relationship between the lactic acid concentration and expression of LDHA and GLUT1, which are related to the Warburg effect, in human head and neck squamous cell carcinoma (HNSCC). Tumors expressing lower levels of LDHA and GLUT1 had a higher concentration of lactic acid than those with higher LDHA and GLUT1 expression. Lactic acid also suppressed the expression of LDHA and GLUT1 in vitro. We previously reported that lactic acid enhances expression of an M2 macrophage marker, ARG1, in murine macrophages. Therefore, we investigated the relationship between the lactic acid concentration and polarization of M2 macrophages in HNSCC by measuring the expression of M2 macrophage markers, CSF1R and CD163, normalized using a pan-macrophage marker, CD68. Tumors with lower levels of CD68 showed a higher concentration of lactic acid, whereas those with higher levels of CSF1R showed a significantly higher concentration of lactic acid. A similar tendency was observed for CD163. These results suggest that tumor-secreted lactic acid is linked to the reduction of macrophages in tumors and promotes induction of M2-like macrophage polarization in human HNSCC. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  9. M2 macrophage polarisation is associated with alveolar formation during postnatal lung development

    PubMed Central

    2013-01-01

    increased number of Csf1r-EGFP+ macrophages and upregulated expression of Arginase 1 (Arg1), Mannose receptor 1 (Mrc1) and Chemokine C-C motif ligand 17 (Ccl17), indicative of an M2 or tissue remodelling macrophage phenotype. Administration of CSF-1 to neonatal mice increased trophic macrophages during development and was associated with increased expression of the M2-associated gene Found in inflammatory zone (Fizz)1 and the growth regulator Insulin-like growth factor (Igf)1. The effects of CSF-1 were identified as macrophage-mediated, as the CSF-1R was found to be exclusively expressed on interstitial myeloid cells. Conclusions This study identifies the presence of CSF-1R+ M2-polarised macrophages localising to sites of branching morphogenesis and increasing in number during the alveolarisation stage of normal lung development. Improved understanding of the role of macrophages in lung developmental regulation has clinical relevance for addressing neonatal inflammatory perturbation of development and highlights macrophage modulation as a potential intervention to promote lung development. PMID:23560845

  10. Unique Macrophages Different from M1/M2 Macrophages Inhibit T Cell Mitogenesis while Upregulating Th17 Polarization

    PubMed Central

    Tatano, Yutaka; Shimizu, Toshiaki; Tomioka, Haruaki

    2014-01-01

    Mycobacterial infection induces suppressor macrophages (MΦs), causing disease exacerbation. There are two major MΦ subsets (M1 and M2 MΦs) that are phenotypically and functionally different. Here, we examined which of the MΦ subsets the mycobacterial infection-induced suppressor MΦs (MIS-MΦs) belong to. MIS-MΦs down-regulated T cell production of Th1 and Th2 cytokines but markedly increased production of interleukin (IL)-17A and IL-22 through up-regulation of Th17 cell expansion. In this phenomenon, a novel MΦ population, which is functionally distinguishable from M1 and M2 MΦ subsets and possesses unique phenotypes (IL-12+, IL-1βhigh, IL-6+, tumor necrosis factor (TNF)-α+, nitric oxide synthase (NOS) 2+, CCR7high, IL-10high, arginase (Arg)-1−, mannose receptor (MR)low, Ym1high, Fizzlow, and CD163high), played central roles through the action of IL-6 and transforming growth factor (TGF)-β but not IL-21 and IL-23. This new type of MΦ population was induced in infected mice and actively supported the in vivo expansion of Th17 cells. PMID:24553452

  11. Macrophages Undergo M1-to-M2 Transition in Adipose Tissue Regeneration in a Rat Tissue Engineering Model.

    PubMed

    Li, Zhijin; Xu, Fangfang; Wang, Zhifa; Dai, Taiqiang; Ma, Chao; Liu, Bin; Liu, Yanpu

    2016-10-01

    Macrophages are involved in the full processes of tissue healing or regeneration and play an important role in the regeneration of a variety of tissues. Although recent evidence suggests the role of different macrophage phenotypes in adipose tissue expansion, metabolism, and remodeling, the spectrum of macrophage phenotype in the adipose tissue engineering field remains unknown. The present study established a rat model of adipose tissue regeneration using a tissue engineering chamber. Macrophage phenotypes were assessed during the regenerative process in the model. Neo-adipose tissue was generated 6 weeks after implantation. Macrophages were obvious in the chamber constructs 3 days after implantation, peaked at day 7, and significantly decreased thereafter. At day 3, macrophages were predominantly M1 macrophages (CCR7+), and there were few M2 macrophages (CD206+). At day 7, the percentage of M2 macrophages significantly increased and remained stable at day 14. M2 macrophages became the predominant macrophage population at 42 days. Enzyme-linked immunosorbent assay demonstrated transition of cytokines from pro-inflammatory to anti-inflammatory, which was consistent with the transition of macrophage phenotype from M1 to M2. These results showed distinct transition of macrophage phenotypes from a pro-inflammatory M1 phenotype to an anti-inflammatory M2 in adipose tissue regeneration in our tissue engineering model. This study provides new insight into macrophage phenotype transition in the regeneration of adipose tissue.

  12. Predominance of M2-polarized macrophages in bladder cancer affects angiogenesis, tumor grade and invasiveness.

    PubMed

    Takeuchi, Hisashi; Tanaka, Michio; Tanaka, Ayako; Tsunemi, Akisa; Yamamoto, Hidenobu

    2016-05-01

    Tumor-associated macrophages (TAMs) often assume an immunoregulatory M2 phenotype. Thus, the aim of the present study was to clarify the correlation of vascularity and TAMs, in particular the M2 phenotype in the stroma and tumor areas, with the clinical and pathological outcomes of patients with bladder cancer. The TAM counts and microvessel counts (MVCs) were determined immunohistochemically in 21 patients with bladder cancer. The number of infiltrating TAMs was measured using immunohistochemistry with anti-cluster of differentiation (CD)68 and anti-CD163 antibodies, to identify a macrophage lineage marker and an M2-polarized-specific cell surface receptor, respectively. CD68(+) and CD163(+) macrophages were evaluated in the stroma and tumor areas, and areas with a high density of infiltrating cell spots were counted. MVCs were determined using immunohistochemistry with anti-CD34 antibodies. The results revealed that the higher ratio of CD163(+)/CD68(+) macrophages in the stroma, tumor and total tumor tissues were correlated with a higher stage and grade (P<0.05). In addition, the low ratio of CD68(+)/CD34(+) microvessels was correlated with a higher stage (P<0.05). There was also a positive correlation between TAMs and MVC (r(2)=0.25; P<0.05). These results suggest that the TAM polarized M2 phenotype affects microvessels, pathological outcome, tumor grade and invasiveness.

  13. M1 and M2 macrophage proteolytic and angiogenic profile analysis in atherosclerotic patients reveals a distinctive profile in type 2 diabetes.

    PubMed

    Roma-Lavisse, Charlotte; Tagzirt, Madjid; Zawadzki, Christophe; Lorenzi, Rodrigo; Vincentelli, André; Haulon, Stephan; Juthier, Francis; Rauch, Antoine; Corseaux, Delphine; Staels, Bart; Jude, Brigitte; Van Belle, Eric; Susen, Sophie; Chinetti-Gbaguidi, Giulia; Dupont, Annabelle

    2015-07-01

    This study aimed to investigate atherosclerotic mediators' expression levels in M1 and M2 macrophages and to focus on the influence of diabetes on M1/M2 profiles. Macrophages from 36 atherosclerotic patients (19 diabetics and 17 non-diabetics) were cultured with interleukin-1β (IL-1β) or IL-4 to induce M1 or M2 phenotype, respectively. The atherosclerotic mediators' expression was evaluated by quantitative reverse transcription-polymerase chain reaction (RT-PCR). The results showed that M1 and M2 macrophages differentially expressed mediators involved in proteolysis and angiogenesis processes. The proteolytic balance (matrix metalloproteinase-9 (MMP-9)/tissue inhibitor of metalloproteinase-1 (TIMP-1), MMP-9/plasminogen activator inhibitor-1 (PAI-1) and MMP-9/tissue factor pathway inhibitor-2 (TFPI-2) ratios) was higher in M1 versus M2, whereas M2 macrophages presented higher angiogenesis properties (increased vascular endothelial growth factor/TFPI-2 and tissue factor/TFPI-2 ratios). Moreover, M1 macrophages from diabetics displayed more important proangiogenic and proteolytic activities than non-diabetics. This study reveals that M1 and M2 macrophages could differentially modulate major atherosclerosis-related pathological processes. Moreover, M1 macrophages from diabetics display a deleterious phenotype that could explain the higher plaque vulnerability observed in these subjects.

  14. Cell plasticity in wound healing: paracrine factors of M1/ M2 polarized macrophages influence the phenotypical state of dermal fibroblasts.

    PubMed

    Ploeger, Diana Ta; Hosper, Nynke A; Schipper, Martin; Koerts, Jasper A; de Rond, Saskia; Bank, Ruud A

    2013-04-19

    Macrophages and fibroblasts are two major players in tissue repair and fibrosis. Despite the relevance of macrophages and fibroblasts in tissue homeostasis, remarkably little is known whether macrophages are able to influence the properties of fibroblasts. Here we investigated the role of paracrine factors secreted by classically activated (M1) and alternatively activated (M2) human macrophages on human dermal fibroblasts (HDFs). HDFs stimulated with paracrine factors from M1 macrophages showed a 10 to > 100-fold increase in the expression of the inflammatory cytokines IL6, CCL2 and CCL7 and the matrix metalloproteinases MMP1 and MMP3. This indicates that factors produced by M1 macrophages induce a fibroblast phenotype with pro-inflammatory and extracellular matrix (ECM) degrading properties. HDFs stimulated with paracrine factors secreted by M2 macrophages displayed an increased proliferation rate. Interestingly, the M1-activated pro-inflammatory fibroblasts downregulated, after exposure to paracrine factors produced by M2 macrophages or non-conditioned media, the inflammatory markers as well as MMPs and upregulated their collagen production. Paracrine factors of M1 or M2 polarized macrophages induced different phenotypes of HDFs and the HDF phenotypes can in turn be reversed, pointing to a high dynamic plasticity of fibroblasts in the different phases of tissue repair.

  15. Cell plasticity in wound healing: paracrine factors of M1/ M2 polarized macrophages influence the phenotypical state of dermal fibroblasts

    PubMed Central

    2013-01-01

    Background Macrophages and fibroblasts are two major players in tissue repair and fibrosis. Despite the relevance of macrophages and fibroblasts in tissue homeostasis, remarkably little is known whether macrophages are able to influence the properties of fibroblasts. Here we investigated the role of paracrine factors secreted by classically activated (M1) and alternatively activated (M2) human macrophages on human dermal fibroblasts (HDFs). Results HDFs stimulated with paracrine factors from M1 macrophages showed a 10 to > 100-fold increase in the expression of the inflammatory cytokines IL6, CCL2 and CCL7 and the matrix metalloproteinases MMP1 and MMP3. This indicates that factors produced by M1 macrophages induce a fibroblast phenotype with pro-inflammatory and extracellular matrix (ECM) degrading properties. HDFs stimulated with paracrine factors secreted by M2 macrophages displayed an increased proliferation rate. Interestingly, the M1-activated pro-inflammatory fibroblasts downregulated, after exposure to paracrine factors produced by M2 macrophages or non-conditioned media, the inflammatory markers as well as MMPs and upregulated their collagen production. Conclusions Paracrine factors of M1 or M2 polarized macrophages induced different phenotypes of HDFs and the HDF phenotypes can in turn be reversed, pointing to a high dynamic plasticity of fibroblasts in the different phases of tissue repair. PMID:23601247

  16. Ex vivo and in vitro effect of serum amyloid A in the induction of macrophage M2 markers and efferocytosis of apoptotic neutrophils1

    PubMed Central

    Sun, Lei; Zhou, Huibin; Zhu, Ziyan; Yan, Qian; Wang, Lili; Liang, Qing; Ye, Richard D.

    2015-01-01

    Macrophages affect the magnitude and duration of inflammatory response in a functionally heterogeneous manner. The phenotype of macrophages is maintained through a reversible homeostatic mechanism. A number of determinants that modulate macrophage plasticity have been identified, although the precise mechanisms are not fully understood. Here we report that stimulation of isolated human blood monocytes and mouse bone marrow-derived macrophages with human serum amyloid A (SAA), a major acute-phase protein, leads to induced expression of macrophage M2 markers including IL-10, Ym1, Fizz-1, MRC1, IL-1Rn and CCL17. The same effect was observed with macrophages exposed to SAA in peritoneal cavity. SAA also increases arginase 1 activity and enhances macrophage efferocytosis of apoptotic neutrophils in mouse macrophages. The induction of M2 markers requires MyD88 and the activation of multiple signaling pathways, but is independent of Stat6. SAA induces IRF4 expression and increases its DNA-binding activity. Silencing IRF4 by siRNA abrogates SAA-induced expression of the M2 markers. These results suggest a potential role for SAA to alter macrophage phenotype and modulate macrophage functions through a MyD88-dependent mechanism that involves IRF4-mediated transcription. PMID:25870242

  17. IL-37 Expression is Upregulated in Patients with Tuberculosis and Induces Macrophages Towards an M2-like Phenotype.

    PubMed

    Huang, Z; Gao, C; Chi, X; Hu, Y W; Zheng, L; Zeng, T; Wang, Q

    2015-10-01

    Interleukin-37 (IL-37), a member of the IL-1 family, primarily functions as an anti-inflammatory cytokine, reducing inflammation and suppressing the immune response. However, the expression and role of IL-37 in tuberculosis (TB) remains unknown. We aimed to measure serum levels of IL-37 and several important cytokines in 25 patients with active TB and to analyse their association with disease activity. We found that IL-37 levels decreased in patients with TB and recovered after treatment. IL-37 levels negatively correlated with the serum concentration of IFN-γ and IL-12 but positively correlated with IL-10 and TGF-β levels. After IL-37, secretion was blocked in peripheral blood mononuclear cells from active patients with TB, IFN-γ and IL-10 production was significantly upregulated; this was not observed in healthy donors or patients after treatment. IL-37 knockdown significantly enhanced the phagocytic activity of THP1-derived macrophages towards Mycobacterium tuberculosis (M. tb). M1/M2 polarization-associated markers were detected simultaneously, and IL-37 induced a phenotypic shift in THP1-derived macrophages towards a high CD206(+) and low CD86(+) macrophage subtype. Furthermore, this phenotypic shift was accompanied by upregulated mRNA levels of arginase 1, TGF-β and IL-10, which are characteristic hallmarks of M2 macrophages. In conclusion, our results suggest that increased levels of IL-37 in patients with TB are associated with IFN-γ, IL-12, IL-10 and TGF-β levels and that IL-37 plays a pathological role in TB infection by inhibiting the production of pro-inflammatory cytokines and inducing macrophages towards an M2-like phenotype. Thus, IL-37 may be a novel research target to understand the pathogenesis of TB infection. © 2015 The Foundation for the Scandinavian Journal of Immunology.

  18. The adenosine-dependent angiogenic switch of macrophages to an M2-like phenotype is independent of interleukin-4 receptor alpha (IL-4Rα) signaling.

    PubMed

    Ferrante, Christopher James; Pinhal-Enfield, Grace; Elson, Genie; Cronstein, Bruce Neil; Hasko, Gyorgy; Outram, Shalini; Leibovich, Samuel Joseph

    2013-08-01

    Murine macrophages are activated by interferon-γ (IFN-γ) and/or Toll-like receptor (TLR) agonists such as bacterial endotoxin (lipopolysaccharide [LPS]) to express an inflammatory (M1) phenotype characterized by the expression of nitric oxide synthase-2 (iNOS) and inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin (IL)-12. In contrast, Th2 cytokines IL-4 and IL-13 activate macrophages by inducing the expression of arginase-1 and the anti-inflammatory cytokine IL-10 in an IL-4 receptor-α (IL-4Rα)-dependent manner. Macrophages activated in this way are designated as "alternatively activated" (M2a) macrophages. We have shown previously that adenosine A2A receptor (A(2A)R) agonists act synergistically with TLR2, TLR4, TLR7, and TLR9 agonists to switch macrophages into an "M2-like" phenotype that we have termed "M2d." Adenosine signaling suppresses the TLR-dependent expression of TNF-α, IL-12, IFN-γ, and several other inflammatory cytokines by macrophages and induces the expression of vascular endothelial growth factor (VEGF) and IL-10. We show here using mice lacking a functional IL-4Rα gene (IL-4Rα(-/-) mice) that this adenosine-mediated switch does not require IL-4Rα-dependent signaling. M2d macrophages express high levels of VEGF, IL-10, and iNOS, low levels of TNF-α and IL-12, and mildly elevated levels of arginase-1. In contrast, M2d macrophages do not express Ym1, Fizz1 (RELM-α), or CD206 at levels greater than those induced by LPS, and dectin-1 expression is suppressed. The use of these markers in vivo to identify "M2" macrophages thus provides an incomplete picture of macrophage functional status and should be viewed with caution.

  19. Dopamine induces growth inhibition and vascular normalization through reprogramming M2-polarized macrophages in rat C6 glioma

    SciTech Connect

    Qin, Tian; Wang, Chenlong; Chen, Xuewei; Duan, Chenfan; Zhang, Xiaoyan; Zhang, Jing; Chai, Hongyan; Tang, Tian; Chen, Honglei; Yue, Jiang; Li, Ying; Yang, Jing

    2015-07-15

    Dopamine (DA), a monoamine catecholamine neurotransmitter with antiangiogenic activity, stabilizes tumor vessels in colon, prostate and ovarian cancers, thus increases chemotherapeutic efficacy. Here, in the rat C6 glioma models, we investigated the vascular normalization effects of DA and its mechanisms of action. DA (25, 50 mg/kg) inhibited tumor growth, while a precursor of DA (levodopa) prolonged the survival time of rats bearing orthotopic C6 glioma. DA improved tumor perfusion, with significant effects from day 3, and a higher level at days 5 to 7. In addition, DA decreased microvessel density and hypoxia-inducible factor-1α expression in tumor tissues, while increasing the coverage of pericyte. Conversely, an antagonist of dopamine receptor 2 (DR2) (eticlopride) but not DR1 (butaclamol) abrogated DA-induced tumor regression and vascular normalization. Furthermore, DA improved the delivery and efficacy of temozolomide therapy. Importantly, DA increased representative M1 markers (iNOS, CXCL9, etc.), while decreasing M2 markers (CD206, arginase-1, etc.). Depletion of macrophages by clodronate or zoledronic acid attenuated the effects of DA. Notably, DA treatment induced M2-to-M1 polarization in RAW264.7 cells and mouse peritoneal macrophages, and enhanced the migration of pericyte-like cells (10T1/2), which was reversed by eticlopride or DR2-siRNA. Such changes were accompanied by the downregulation of VEGF/VEGFR2 signaling. In summary, DA induces growth inhibition and vascular normalization through reprogramming M2-polarized macrophages. Thus, targeting the tumor microvasculature by DA represents a promising strategy for human glioma therapy. - Highlights: • Dopamine induces tumor growth inhibition and vascular normalization in rat C6 glioma. • Dopamine switches macrophage phenotype from M2 to M1. • Dopamine-induced vascular normalization is mediated by macrophage polarization. • Dopamine is a promising agent targeting the microvasculature in tumor

  20. Metabolic Characterization of Polarized M1 and M2 Bone Marrow-derived Macrophages Using Real-time Extracellular Flux Analysis.

    PubMed

    Van den Bossche, Jan; Baardman, Jeroen; de Winther, Menno P J

    2015-11-28

    Specific metabolic pathways are increasingly being recognized as critical hallmarks of macrophage subsets. While LPS-induced classically activated M1 or M(LPS) macrophages are pro-inflammatory, IL-4 induces alternative macrophage activation and these so-called M2 or M(IL-4) support resolution of inflammation and wound healing. Recent evidence shows the crucial role of metabolic reprogramming in the regulation of M1 and M2 macrophage polarization. In this manuscript, an extracellular flux analyzer is applied to assess the metabolic characteristics of naive, M1 and M2 polarized mouse bone marrow-derived macrophages. This instrument uses pH and oxygen sensors to measure the extracellular acidification rate (ECAR) and oxygen consumption rate (OCR), which can be related to glycolytic and mitochondrial oxidative metabolism. As such, both glycolysis and mitochondrial oxidative metabolism can be measured in real-time in one single assay. Using this technique, we demonstrate here that inflammatory M1 macrophages display enhanced glycolytic metabolism and reduced mitochondrial activity. Conversely, anti-inflammatory M2 macrophages show high mitochondrial oxidative phosphorylation (OXPHOS) and are characterized by an enhanced spare respiratory capacity (SRC). The presented functional assay serves as a framework to investigate how particular cytokines, pharmacological compounds, gene knock outs or other interventions affect the macrophage's metabolic phenotype and inflammatory status.

  1. BCL6 mediates the effects of Gastrodin on promoting M2-like macrophage polarization and protecting against oxidative stress-induced apoptosis and cell death in macrophages.

    PubMed

    Jia, Jing; Shi, Xiaojie; Jing, Xiaoqian; Li, Jianguo; Gao, Jie; Liu, Mengya; Lin, Chi-Iou; Guo, Xinzhi; Hua, Qian

    2017-03-16

    Cerebral palsy (CP) is the most common childhood disability worldwide, yet biomarkers for predicting CP are lacking. By subjecting peripheral blood samples from 62 CP patients and 30 healthy controls to Affymetrix GeneChip(®) PrimeView™ HumanGene Expression Microarray analysis, we identified the novel biomarker B-cell lymphoma 6 (BCL6) as the most upregulated gene in the CP samples. Gastrodin is a traditional Chinese medicine and bioactive compound that promotes adductor angle release, as well as gross and fine motor performance by increasing Gross Motor Function Measure-66 and Fine Motor Function Measure-45 scores. Gastrodin upregulates the mRNA expression of Mgl2 and Mrc1, M2 macrophage markers, and arginase activity, an M2 polarization indicator, in murine RAW264.7 macrophages. Moreover, these effects were blocked by BCL6 siRNA, which also abrogated the protective effects of Gastrodin against hydrogen peroxide-induced apoptosis and death in RAW264.7 cells. Our work identified BCL6 as a novel biomarker for early prediction of CP. Moreover, we demonstrated that Gastrodin not only stimulated polarization toward M2-like macrophages, which promote tissue repair, but also rescued macrophages from oxidative stress, apoptosis and death by inducing BCL6 expression. BCL6-targeted therapeutic strategies have promise for improving motor performance in CP patients.

  2. Complement protein C1q promotes macrophage anti-inflammatory M2-like polarization during the clearance of atherogenic lipoproteins

    PubMed Central

    Spivia, Weston; Magno, Patrick S.; Le, Patrick; Fraser, Deborah A.

    2016-01-01

    Objective Innate immune protein C1q plays a dual role in the chronic inflammatory disease of atherosclerosis. Complement activation via C1q exacerbates pathology in the atherosclerotic lesion in later stages of the disease. However, in early stages of disease C1q is protective. We hypothesize that complement-independent activities of C1q are involved in reprogramming macrophage inflammatory polarization. Methods The influence of C1q on macrophage inflammatory responses during clearance of oxLDL was examined. Changes in cytokines at the gene and protein level were measured by quantitative PCR and ELISA assay. Results C1q modulated cytokine expression in Raw264.7 macrophages during ingestion of oxLDL. Levels of proinflammatory cytokines IL-1β and IL-6 were downregulated by C1q, whereas levels of the anti-inflammatory cytokine IL-10 were increased. In addition, data from an NFκB-luciferase gene reporter assay suggest that C1q suppresses activation of NFκB during lipoprotein clearance in macrophages, providing one mechanism by which C1q downregulates pro-inflammatory cytokine production. Conclusions C1q-polarization of macrophages toward an anti-inflammatory (M2-like) phenotype may be important in dampening inflammation in the early atherosclerotic lesion. Further investigation of molecular pathways targeted by C1q may provide novel therapeutic targets for this disease. PMID:25091012

  3. All-Trans Retinoic Acid Prevents Osteosarcoma Metastasis by Inhibiting M2 Polarization of Tumor-Associated Macrophages.

    PubMed

    Zhou, Qian; Xian, Miao; Xiang, Senfeng; Xiang, Danyan; Shao, Xuejing; Wang, Jincheng; Cao, Ji; Yang, Xiaochun; Yang, Bo; Ying, Meidan; He, Qiaojun

    2017-07-01

    M2-polarized tumor-associated macrophages (TAM) play a critical role in cancer invasion and metastasis. Here, we report that M2 macrophages enhanced metastasis of K7M2 WT osteosarcoma cells to the lungs in mice, thus establishing M2 TAMs as a therapeutic target for blocking osteosarcoma metastasis. We found that all-trans retinoic acid (ATRA) inhibited osteosarcoma metastasis via inhibiting the M2 polarization of TAMs. ATRA suppressed IL13- or IL4-induced M2-type macrophages, and then inhibited migration of osteosarcoma cells as promoted by M2-type macrophages in vitro ATRA reduced the number of pulmonary metastatic nodes of osteosarcoma and decreased expression of M2-type macrophages in metastatic nodes both in intravenous injection and orthotopic transplantation models. ATRA's effect was independent of conventional STAT3/6 or C/EBPβ signaling, which regulate M2-like polarization of macrophages. Quantitative genomic and functional analyses revealed that MMP12, a macrophage-secreted elastase, was elevated in IL13-skewed TAM polarization, whereas ATRA treatment downregulated IL13-induced secretion of MMP12. This downregulation correlates with the antimetastasis effect of ATRA. Our results show the role of TAM polarization in osteosarcoma metastasis, identify a therapeutic opportunity for antimetastasis treatment, and indicate ATRA treatment as an approach for preventing osteosarcoma metastasis via M2-type polarization intervention. Cancer Immunol Res; 5(7); 547-59. ©2017 AACR. ©2017 American Association for Cancer Research.

  4. IL-33 improves wound healing through enhanced M2 macrophage polarization in diabetic mice.

    PubMed

    He, Rongguo; Yin, Hui; Yuan, Baohong; Liu, Tao; Luo, Li; Huang, Ping; Dai, Liangcheng; Zeng, Kang

    2017-10-01

    IL-33 is a newly discovered member of the IL-1 family and has been identified as a potent inducer of Th2 type immunity. Emerging evidence imply that IL-33 may also act as an alarm to alert the immune system when released by epithelial barrier tissues during trauma or infection. In this study, we further investigate the potential efficacy of IL-33 on dermal wound healing in streptozotocin-induced diabetic mice. A full-thickness skin wound was generated on the back of diabetic mice and treated with IL-33 or vehicle topically. Our data showed that IL-33 delivery contributed to diabetic wound closure with wounds gaping narrower and exhibiting elevated re-epithelialization. IL-33 promoted the new extracellular matrix (ECM) deposition and angiogenesis formation, which indicates an important role of IL-33 on matrix synthesis and neovascularization. Meanwhile, IL-33 accelerated the development of M2 macrophages in wound sites in vivo, and amplified IL-13-induced polarization of bone marrow-derived macrophages toward a M2 phenotype in vitro. Furthermore, IL-33-amplified M2 macrophages augmented the proliferation of fibroblasts and ECM deposition. All together, these results strongly suggest manipulation of IL-33-mediated signal might be a potential therapeutic approach for diabetic skin wounds. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

    PubMed Central

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

    2016-01-01

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

  6. CXCR3 deficiency enhances tumor progression by promoting macrophage M2 polarization in a murine breast cancer model.

    PubMed

    Oghumu, Steve; Varikuti, Sanjay; Terrazas, Cesar; Kotov, Dmitri; Nasser, Mohd W; Powell, Catherine A; Ganju, Ramesh K; Satoskar, Abhay R

    2014-09-01

    Tumor associated macrophages play a vital role in determining the outcome of breast cancer. We investigated the contribution of the chemokine receptor CXCR3 to antitumor immune responses using a cxcr3 deficient mouse orthotopically injected with a PyMT breast cancer cell line. We observed that cxcr3 deficient mice displayed increased IL-4 production and M2 polarization in the tumors and spleens compared to WT mice injected with PyMT cells. This was accompanied by larger tumor development in cxcr3(-/-) than in WT mice. Further, tumor-promoting myeloid derived immune cell populations accumulated in higher proportions in the spleens of cxcr3 deficient mice. Interestingly, cxcr3(-/-) macrophages displayed a deficiency in up-regulating inducible nitric oxide synthase after stimulation by either IFN-γ or PyMT supernatants. Stimulation of bone marrow derived macrophages by PyMT supernatants also resulted in greater induction of arginase-1 in cxcr3(-/-) than WT mice. Further, cxcr3(-/-) T cells activated with CD3/CD28 in vitro produced greater amounts of IL-4 and IL-10 than T cells from WT mice. Our data suggests that a greater predisposition of cxcr3 deficient macrophages towards M2 polarization contributes to an enhanced tumor promoting environment in cxcr3 deficient mice. Although CXCR3 is known to be expressed on some macrophages, this is the first report that demonstrates a role for CXCR3 in macrophage polarization and subsequent breast tumor outcomes. Targeting CXCR3 could be a potential therapeutic approach in the management of breast cancer tumors.

  7. Arctigenin ameliorates inflammation in vitro and in vivo by inhibiting the PI3K/AKT pathway and polarizing M1 macrophages to M2-like macrophages.

    PubMed

    Hyam, Supriya R; Lee, In-Ah; Gu, Wan; Kim, Kyung-Ah; Jeong, Jin-Ju; Jang, Se-Eun; Han, Myung Joo; Kim, Dong-Hyun

    2013-05-15

    Seeds of Arctium lappa, containing arctigenin and its glycoside arctiin as main constituents, have been used as a diuretic, anti-inflammatory and detoxifying agent in Chinese traditional medicine. In our preliminary study, arctigenin inhibited IKKβ and NF-κB activation in peptidoglycan (PGN)- or lipopolysaccharide (LPS)-induced peritoneal macrophages. To understand the anti-inflammatory effect of arctigenin, we investigated its anti-inflammatory effect in LPS-stimulated peritoneal macrophages and on LPS-induced systemic inflammation as well as 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis in mice. Arctigenin inhibited LPS-increased IL-1β, IL-6 and TNF-α expression in LPS-stimulated peritoneal macrophages, but increased LPS-reduced IL-10 and CD204 expression. Arctigenin inhibited LPS-induced PI3K, AKT and IKKβ phosphorylation, but did not suppress LPS-induced IRAK-1 phosphorylation. However, arctigenin did not inhibit NF-κB activation in LPS-stimulated PI3K siRNA-treated peritoneal macrophages. Arctigenin suppressed the binding of p-PI3K antibody and the nucleus translocation of NF-κB p65 in LPS-stimulated peritoneal macrophages. Arctigenin suppressed blood IL-1β and TNF-α level in mice systemically inflamed by intraperitoneal injection of LPS. Arctigenin also inhibited colon shortening, macroscopic scores and myeloperoxidase activity in TNBS-induced colitic mice. Arctigenin inhibited TNBS-induced IL-1β, TNF-α and IL-6 expression, as well as PI3K, AKT and IKKβ phosphorylation and NF-κB activation in mice, but increased IL-10 and CD204 expression. However, it did not affect IRAK-1 phosphorylation. Based on these findings, arctigenin may ameliorate inflammatory diseases, such as colitis, by inhibiting PI3K and polarizing M1 macrophages to M2-like macrophages. Copyright © 2013 Elsevier B.V. All rights reserved.

  8. IL-10 enhances the phenotype of M2 macrophages induced by IL-4 and confers the ability to increase eosinophil migration.

    PubMed

    Makita, Naoyuki; Hizukuri, Yoshiyuki; Yamashiro, Kyoko; Murakawa, Masao; Hayashi, Yasuhiro

    2015-03-01

    M2 macrophages have been subdivided into subtypes such as IL-4-induced M2a and IL-10-induced M2c in vitro. Although it was reported that IL-10 stimulation leads to an increase in IL-4Rα, the effect of IL-4 and IL-10 in combination with macrophage subtype differentiation remains unclear. Thus, we sought to clarify whether IL-10 enhanced the M2 phenotype induced by IL-4. In this study, we showed that IL-10 enhanced IL-4Rα expression in M-CSF-induced bone marrow-derived macrophages (BMDMs). Global gene expression analysis of M2 macrophages induced by IL-4, IL-10 or IL-4 + IL-10 showed that IL-10 enhanced gene expression of M2a markers induced by IL-4 in M-CSF-induced BMDMs. Moreover, IL-4 and IL-10 synergistically induced CCL24 (Eotaxin-2) production. Enhanced CCL24 expression was also observed in GM-CSF-induced BMDMs and zymosan-elicited, thioglycolate-elicited and naive peritoneal macrophages. CCL24 is a CCR3 agonist and an eosinophil chemoattractant. In vitro, IL-4 + IL-10-stimulated macrophages produced a large amount of CCL24 and increased eosinophil migration, which was inhibited by anti-CCL24 antibody. We also showed that IL-4 + IL-10-stimulated (but not IL-4 or IL-10 alone) macrophages transferred into the peritoneum of C57BL/6J mice increased eosinophil infiltration into the peritoneal cavity. These results demonstrate that IL-4 + IL-10-simulated macrophages have enhanced M2a macrophage-related gene expression, CCL24 production and eosinophil infiltration-inducing activity, thereby suggesting their contribution to eosinophil-related diseases.

  9. Tolerance and M2 (alternative) macrophage polarization are related processes orchestrated by p50 nuclear factor κB

    PubMed Central

    Porta, Chiara; Rimoldi, Monica; Raes, Geert; Brys, Lea; Ghezzi, Pietro; Di Liberto, Diana; Dieli, Francesco; Ghisletti, Serena; Natoli, Gioacchino; De Baetselier, Patrick; Mantovani, Alberto; Sica, Antonio

    2009-01-01

    Cells of the monocyte–macrophage lineage play a central role in the orchestration and resolution of inflammation. Plasticity is a hallmark of mononuclear phagocytes, and in response to environmental signals these cells undergo different forms of polarized activation, the extremes of which are called classic or M1 and alternative or M2. NF-κB is a key regulator of inflammation and resolution, and its activation is subject to multiple levels of regulation, including inhibitory, which finely tune macrophage functions. Here we identify the p50 subunit of NF-κB as a key regulator of M2-driven inflammatory reactions in vitro and in vivo. p50 NF-κB inhibits NF-κB–driven, M1-polarizing, IFN-β production. Accordingly, p50-deficient mice show exacerbated M1-driven inflammation and defective capacity to mount allergy and helminth-driven M2-polarized inflammatory reactions. Thus, NF-κB p50 is a key component in the orchestration of M2-driven inflammatory reactions. PMID:19706447

  10. Feedback mechanisms between M2 macrophages and Th17 cells in colorectal cancer patients.

    PubMed

    Mao, Hui; Pan, Fei; Guo, Hongxia; Bu, Fangfang; Xin, Tong; Chen, Shukun; Guo, Yajun

    2016-09-01

    IL-17 and IL-22 are linked to the development of intestinal inflammation and colorectal cancer (CRC). However, the maintenance of IL-17 and IL-22 production, as well as the cell type (Th17) that mediates these cytokines in CRC patients, remains unknown. To examine this, untreated CRC patients and healthy controls were recruited in this study. We first observed that CRC patients contained significantly elevated levels of IL-17- and IL-22-producing CD4(+) T cells. The vast majority of IL-22-expressing CD4(+) T cells also expressed IL-17. We then found that the production of both IL-17 and IL-22 required support from autologous monocytes, since the depletion of monocytes significantly downregulated IL-17 and IL-22 secretion. Naive T cells from CRC patients did not secrete IL-17 or IL-22 initially, but long-term coculture with autologous monocytes significantly upregulated IL-17 and IL-22 production in an IL-6-dependent manner. Blockade of IL-6 significantly reduced the levels of both IL-17 and IL-22. We then observed that CD163(+) M2 macrophages were the main contributor of IL-6. Interestingly, incubation of monocytes with CCR4(+)CCR6(+) Th17 cells resulted in significantly higher levels of CD163(+) macrophages as well as higher IL-6 secretion, than incubation with non-Th17 CD4(+) T cells. Together, our study discovered a positive feedback mechanism between Th17 and M2 macrophages in CRC patients.

  11. Substance P Induces HO-1 Expression in RAW 264.7 Cells Promoting Switch towards M2-Like Macrophages

    PubMed Central

    Montana, Giovanna

    2016-01-01

    Substance P (SP) is a neuropeptide that mediates many physiological as well as inflammatory responses. Recently, SP has been implicated in the resolution of inflammation through induction of M2 macrophages phenotype. The shift between M1-like and M2-like, allowing the resolution of inflammatory processes, also takes place by means of hemeoxygenase-1 (HO-1). HO-1 is induced in response to oxidative stress and inflammatory stimuli and modulates the immune response through macrophages polarisation. SP induces HO-1 expression in human periodontal ligament (PDL), the latter potentially plays a role in cytoprotection. We demonstrated that SP promotes M2-like phenotype from resting as well as from M1 macrophages. Indeed, SP triggers the production of interleukine-10 (IL-10), interleukine-4 (IL-4) and arginase-1 (Arg1) without nitric oxide (NO) generation. In addition, SP increases HO-1 expression in a dose- and time-dependent manner. Here we report that SP, without affecting cell viability, significantly reduces the production of pro-inflammatory cytokines and enzymes, such as tumor necrosis factor-alpha (TNF-α), interleukine-6 (IL-6), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and ameliorates migration and phagocytic properties in LPS-stimulated RAW 264.7 cells. M2-like conversion required retention of NF-κB p65 into the cytoplasm and HO-1 induced expression. Silencing of the HO-1 mRNA expression reversed the induction of pro-inflammatory cytokines in RAW 264.7 stimulated by LPS and down-regulated anti-inflammatory hallmarks of M2 phenotype. In conclusion, our data show that SP treatment might be associated with anti-inflammatory effects in LPS-stimulated RAW 264.7 cells by suppressing NF-κB activation and inducing HO-1 expression. PMID:27907187

  12. Pentraxin-3 Attenuates Renal Damage in Diabetic Nephropathy by Promoting M2 Macrophage Differentiation.

    PubMed

    Sun, Huaibin; Tian, Jun; Xian, Wanhua; Xie, Tingting; Yang, Xiangdong

    2015-10-01

    As one of the most important long-term complications of diabetes, diabetic nephropathy (DN) is the major cause of end-stage renal disease and high mortality in diabetic patients. The long pentraxin 3 (Ptx3) is a member of a superfamily of conserved proteins characterized by a cyclic multimeric structure and a conserved C-terminal domain. Several clinical investigations have demonstrated that elevated plasma Ptx3 levels are associated with cardiovascular and chronic kidney diseases (CKD). However, the therapeutic effect of Ptx3 on DN has never been investigated. In our current study, we showed a crucial role for Ptx3 in attenuating renal damage in DN. In our mouse hyperglycemia-induced nephropathy model, Ptx3 treatment showed significantly increased expression of nephrin, acetylated nephrin, and Wilm's tumor-1 protein (WT-1) when compared with control. The number of CD4(+) T cells, CD8(+) T cells, Ly6G(+) neutrophils, and CD11b(+) macrophages were all significantly lower in the Ptx3-treated group than that in the control group in DN. The IL-4 and IL-13 levels in the Ptx3-treated group were markedly higher than that in the control group in DN. Correspondingly, the Ptx3-treated group showed increased numbers of Arg1- or CD206-expressing macrophages compared with the control group. Furthermore, inhibition of Ptx3-treated macrophages abrogated the alleviated renal damage induced by Ptx3 treatment. In conclusion, Ptx3 attenuates renal damage in DN by promoting M2 macrophage differentiation.

  13. Carboxyl- and amino-functionalized polystyrene nanoparticles differentially affect the polarization profile of M1 and M2 macrophage subsets.

    PubMed

    Fuchs, Ann-Kathrin; Syrovets, Tatiana; Haas, Karina A; Loos, Cornelia; Musyanovych, Anna; Mailänder, Volker; Landfester, Katharina; Simmet, Thomas

    2016-04-01

    Macrophages are key regulators of innate and adaptive immune responses. Exposure to microenvironmental stimuli determines their polarization into proinflammatory M1 and anti-inflammatory M2 macrophages. M1 exhibit high expression of proinflammatory TNF-α and IL-1β, and M2 promote tissue repair, but likewise support tumor growth and cause immune suppression by expressing IL-10. Thus, the M1/M2 balance critically determines tissue homeostasis. By using carboxyl- (PS-COOH) and amino-functionalized (PS-NH2) polystyrene nanoparticles, the effects of surface decoration on the polarization of human macrophages were investigated. The nanoparticles did not compromise macrophage viability nor did they affect the expression of the M1 markers CD86, NOS2, TNF-α, and IL-1β. By contrast, in M2, both nanoparticles impaired expression of scavenger receptor CD163 and CD200R, and the release of IL-10. PS-NH2 also inhibited phagocytosis of Escherichia coli by both, M1 and M2. PS-COOH did not impair phagocytosis by M2, but increased protein mass in M1 and M2, TGF-β1 release by M1, and ATP levels in M2. Thus, nanoparticles skew the M2 macrophage polarization without affecting M1 markers. Given the critical role of the M1 and M2 polarization for the immunological balance in patients with cancer or chronic inflammation, functionalized nanoparticles might serve as tools for reprogramming the M1/M2 polarization.

  14. Isoliquiritigenin, a flavonoid from licorice, blocks M2 macrophage polarization in colitis-associated tumorigenesis through downregulating PGE{sub 2} and IL-6

    SciTech Connect

    Zhao, Haixia; Zhang, Xinhua; Chen, Xuewei; Li, Ying; Ke, Zunqiong; Tang, Tian; Chai, Hongyan; Guo, Austin M.; Chen, Honglei; Yang, Jing

    2014-09-15

    M2 macrophage polarization is implicated in colorectal cancer development. Isoliquiritigenin (ISL), a flavonoid from licorice, has been reported to prevent azoxymethane (AOM) induced colon carcinogenesis in animal models. Here, in a mouse model of colitis-associated tumorigenesis induced by AOM/dextran sodium sulfate (DSS), we investigated the chemopreventive effects of ISL and its mechanisms of action. Mice were treated with AOM/DSS and randomized to receive either vehicle or ISL (3, 15 and 75 mg/kg). Tumor load, histology, immunohistochemistry, and gene and protein expressions were determined. Intragastric administration of ISL for 12 weeks significantly decreased colon cancer incidence, multiplicity and tumor size by 60%, 55.4% and 42.6%, respectively. Moreover, ISL inhibited M2 macrophage polarization. Such changes were accompanied by downregulation of PGE{sub 2} and IL-6 signaling. Importantly, depletion of macrophages by clodronate (Clod) or zoledronic acid (ZA) reversed the effects of ISL. In parallel, in vitro studies also demonstrated that ISL limited the M2 polarization of RAW264.7 cells and mouse peritoneal macrophages with concomitant inactivation of PGE{sub 2}/PPARδ and IL-6/STAT3 signaling. Conversely, exogenous addition of PGE{sub 2} or IL-6, or overexpression of constitutively active STAT3 reversed ISL-mediated inhibition of M2 macrophage polarization. In summary, dietary flavonoid ISL effectively inhibits colitis-associated tumorigenesis through hampering M2 macrophage polarization mediated by the interplay between PGE{sub 2} and IL-6. Thus, inhibition of M2 macrophage polarization is likely to represent a promising strategy for chemoprevention of colorectal cancer. - Highlights: • Isoliquiritigenin (ISL) prevents colitis-associated tumorigenesis. • ISL inhibits M2 macrophage polarization in vivo and in vitro. • ISL inhibits PGE{sub 2} and IL-6 signaling in colitis-associated tumorigenesis. • ISL may be an attractive candidate agent for

  15. M1/M2-macrophage phenotypes regulate renal calcium oxalate crystal development

    PubMed Central

    Taguchi, Kazumi; Okada, Atsushi; Hamamoto, Shuzo; Unno, Rei; Moritoki, Yoshinobu; Ando, Ryosuke; Mizuno, Kentaro; Tozawa, Keiichi; Kohri, Kenjiro; Yasui, Takahiro

    2016-01-01

    In our previous report, M2-macrophage (Mφs) deficient mice showed increased renal calcium oxalate (CaOx) crystal formation; however, the role of Mφs-related-cytokines and chemokines that affect kidney stone formation remains unknown. Here, we investigated the role of M1/M2s in crystal development by using in vitro and in vivo approaches. The crystal phagocytic rate of bone marrow-derived M2Mφs was higher than that of bone marrow-derived Mφs and M1Mφs and increased on co-culture with renal tubular cells (RTCs). However, the amount of crystal attachment on RTCs reduced on co-culture with M2Mφs. In six hyperoxaluric C57BL/6J mice, M1Mφ transfusion and induction by LPS and IFN-γ facilitated renal crystal formation, whereas M2Mφ transfusion and induction by IL-4 and IL-13 suppressed renal crystal formation compared with the control. These M2Mφ treatments reduced the expression of crystal-related genes, such as osteopontin and CD44, whereas M1Mφ treatment increased the expression of pro-inflammatory and adhesion-related genes such as IL-6, inducible NOS, TNF-α, C3, and VCAM-1. The expression of M2Mφ-related genes was lower whereas that of M1Mφ-related genes was higher in papillary tissue of CaOx stone formers. Overall, our results suggest that renal crystal development is facilitated by M1Mφs, but suppressed by M2Mφs. PMID:27731368

  16. Role of C/EBP homologous protein and endoplasmic reticulum stress in asthma exacerbation by regulating the IL-4/signal transducer and activator of transcription 6/transcription factor EC/IL-4 receptor α positive feedback loop in M2 macrophages.

    PubMed

    Wang, Yi; Zhu, Jianghui; Zhang, Lei; Zhang, Zhijun; He, Long; Mou, Yong; Deng, Yanhan; Cao, Yong; Yang, Ping; Su, Yunchao; Zhao, Jianping; Zhang, Shu; Yu, Qilin; Hu, Jifa; Chen, Zhishui; Ning, Qin; Xiang, Xudong; Xu, Yongjian; Wang, Cong-Yi; Xiong, Weining

    2017-02-24

    C/EBP homologous protein (Chop), a marker of endoplasmic reticulum (ER) stress, exhibits aberrant expression patterns during asthma development. However, its exact role in asthma pathogenesis is not fully understood. We aimed to determine the function and mechanism of Chop in the pathogenesis of allergic asthma in patients and animals. Studies were conducted in asthmatic patients and Chop(-/-) mice to dissect the role of Chop and ER stress in asthma pathogenesis. An ovalbumin (OVA)-induced allergic airway inflammation model was used to address the effect of Chop deficiency on asthma development. Next, the effect of Chop deficiency on macrophage polarization and related signaling pathways was investigated to demonstrate the underlying mechanisms. Asthmatic patients and mice after OVA induction exhibited aberrant Chop expression along with ER stress. Specifically, Chop was noted to be specifically overexpressed in macrophages, and mice deficient in Chop were protected from OVA-induced allergic airway inflammation, as manifested by attenuated airway inflammation, remodeling, and hyperresponsiveness. Chop was found to exacerbate allergic airway inflammation by enhancing M2 programming in macrophages. Mechanistic studies characterized an IL-4/signal transducer and activator of transcription 6/transcription factor EC (Tfec)/IL-4 receptor α positive feedback regulatory loop, in which IL-4 induces Chop expression, which then promotes signal transducer and activator of transcription 6 signaling to transcribe Tfec expression. Finally, Tfec transcribes IL-4 receptor α expression to promote M2 programming in macrophages. Chop and ER stress are implicated in asthma pathogenesis, which involves regulation of M2 programming in macrophages. Copyright © 2017 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  17. The M2 macrophages induce autophagic vascular disorder and promote mouse sensitivity to urethane-related lung carcinogenesis.

    PubMed

    Li, G-G; Guo, Z-Z; Ma, X-F; Cao, N; Geng, S-N; Zheng, Y-Q; Meng, M-J; Lin, H-H; Han, G; Du, G-J

    2016-06-01

    Tumor vessels are known to be abnormal, with typically aberrant, leaky and disordered vessels. Here, we investigated whether polarized macrophage phenotypes are involved in tumor abnormal angiogenesis and what is its mechanism. We found that there was no difference in chemotaxis of polarized M1 and M2 macrophages to lewis lung carcinoma (LLC) cells and that either M1 or M2 macrophage-conditioned media had no effect on LLC cell proliferation. Unexpectedly, the M2 but not M1 macrophage-conditioned media promoted the proliferation of human umbilical vein endothelial cells (HUVECs) and simultaneously increased endothelial cell permeability in vitro and angiogenic index in the chick embryo chorioallantoic membrane (CAM). The treatment with M2 but not M1 macrophage-conditioned media increased autophagosomes as well as microtubule-associated protein light chain 3B (LC3-B) expression (a robust marker of autophagosomes) but decreased p62 protein expression (a selective autophagy substrate) in HUVECs, the treatment with chloroquine that blocked autophagy abrogated the abnormal angiogenic efficacy of M2 macrophage-conditioned media. These results were confirmed in urethane-induced lung carcinogenic progression. Urethane-induced lung carcinogenesis led to more M2 macrophage phenotype and increased abnormal angiogenesis concomitant with the upregulation of LC3-B and the downregulation of p62. Clodronate liposome-induced macrophage depletion, chloroquine-induced autophagic prevention or salvianolic acid B-induced vascular protection decreased abnormal angiogenesis and lung carcinogenesis. In addition, we found that the tendency of age-related M2 macrophage polarization also promoted vascular permeability and carcinogenesis in urethane carcinogenic progression. These findings indicate that the M2 macrophages induce autophagic vascular disorder to promote lung cancer progression, and the autophagy improvement represents an efficacious strategy for abnormal angiogenesis and cancer

  18. Anti-CD47 Treatment Stimulates Phagocytosis of Glioblastoma by M1 and M2 Polarized Macrophages and Promotes M1 Polarized Macrophages In Vivo.

    PubMed

    Zhang, Michael; Hutter, Gregor; Kahn, Suzana A; Azad, Tej D; Gholamin, Sharareh; Xu, Chelsea Y; Liu, Jie; Achrol, Achal S; Richard, Chase; Sommerkamp, Pia; Schoen, Matthew Kenneth; McCracken, Melissa N; Majeti, Ravi; Weissman, Irving; Mitra, Siddhartha S; Cheshier, Samuel H

    2016-01-01

    Tumor-associated macrophages (TAMs) represent an important cellular subset within the glioblastoma (WHO grade IV) microenvironment and are a potential therapeutic target. TAMs display a continuum of different polarization states between antitumorigenic M1 and protumorigenic M2 phenotypes, with a lower M1/M2 ratio correlating with worse prognosis. Here, we investigated the effect of macrophage polarization on anti-CD47 antibody-mediated phagocytosis of human glioblastoma cells in vitro, as well as the effect of anti-CD47 on the distribution of M1 versus M2 macrophages within human glioblastoma cells grown in mouse xenografts. Bone marrow-derived mouse macrophages and peripheral blood-derived human macrophages were polarized in vitro toward M1 or M2 phenotypes and verified by flow cytometry. Primary human glioblastoma cell lines were offered as targets to mouse and human M1 or M2 polarized macrophages in vitro. The addition of an anti-CD47 monoclonal antibody led to enhanced tumor-cell phagocytosis by mouse and human M1 and M2 macrophages. In both cases, the anti-CD47-induced phagocytosis by M1 was more prominent than that for M2. Dissected tumors from human glioblastoma xenografted within NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice and treated with anti-CD47 showed a significant increase of M1 macrophages within the tumor. These data show that anti-CD47 treatment leads to enhanced tumor cell phagocytosis by both M1 and M2 macrophage subtypes with a higher phagocytosis rate by M1 macrophages. Furthermore, these data demonstrate that anti-CD47 treatment alone can shift the phenotype of macrophages toward the M1 subtype in vivo.

  19. Anti-CD47 Treatment Stimulates Phagocytosis of Glioblastoma by M1 and M2 Polarized Macrophages and Promotes M1 Polarized Macrophages In Vivo

    PubMed Central

    Kahn, Suzana A.; Azad, Tej D.; Gholamin, Sharareh; Xu, Chelsea Y.; Liu, Jie; Achrol, Achal S.; Richard, Chase; Sommerkamp, Pia; Schoen, Matthew Kenneth; McCracken, Melissa N.; Majeti, Ravi; Weissman, Irving; Mitra, Siddhartha S.; Cheshier, Samuel H.

    2016-01-01

    Tumor-associated macrophages (TAMs) represent an important cellular subset within the glioblastoma (WHO grade IV) microenvironment and are a potential therapeutic target. TAMs display a continuum of different polarization states between antitumorigenic M1 and protumorigenic M2 phenotypes, with a lower M1/M2 ratio correlating with worse prognosis. Here, we investigated the effect of macrophage polarization on anti-CD47 antibody-mediated phagocytosis of human glioblastoma cells in vitro, as well as the effect of anti-CD47 on the distribution of M1 versus M2 macrophages within human glioblastoma cells grown in mouse xenografts. Bone marrow-derived mouse macrophages and peripheral blood-derived human macrophages were polarized in vitro toward M1 or M2 phenotypes and verified by flow cytometry. Primary human glioblastoma cell lines were offered as targets to mouse and human M1 or M2 polarized macrophages in vitro. The addition of an anti-CD47 monoclonal antibody led to enhanced tumor-cell phagocytosis by mouse and human M1 and M2 macrophages. In both cases, the anti-CD47-induced phagocytosis by M1 was more prominent than that for M2. Dissected tumors from human glioblastoma xenografted within NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice and treated with anti-CD47 showed a significant increase of M1 macrophages within the tumor. These data show that anti-CD47 treatment leads to enhanced tumor cell phagocytosis by both M1 and M2 macrophage subtypes with a higher phagocytosis rate by M1 macrophages. Furthermore, these data demonstrate that anti-CD47 treatment alone can shift the phenotype of macrophages toward the M1 subtype in vivo. PMID:27092773

  20. Ethanol Extract of Mylabris phalerata Inhibits M2 Polarization Induced by Recombinant IL-4 and IL-13 in Murine Macrophages

    PubMed Central

    Chung, Hwan-Suck; Lee, Bong-Seon

    2017-01-01

    Mylabris phalerata (MP) is an insect used in oriental herbal treatments for tumor, tinea infections, and stroke. Recent studies have shown that tumor-associated macrophages (TAM) have detrimental roles such as tumor progression, angiogenesis, and metastasis. Although TAM has phenotypes and characteristics in common with M2-polarized macrophages, M1 macrophages have tumor suppression and immune stimulation effects. Medicines polarizing macrophages to M1 have been suggested to have anticancer effects via the modulation of the tumor microenvironment. In this line, we screened oriental medicines to find M1 polarizing medicines in M2-polarized macrophages. Among approximately 400 types of oriental medicine, the ethanol extract of M. phalerata (EMP) was the most proficient in increasing TNF-α secretion in M2-polarized macrophages and TAM. Although EMP enhanced the levels of an M1 cytokine (TNF-α) and a marker (CD86), it significantly reduced the levels of an M2 marker (arginase-1) in M2-polarized macrophages. In addition, EMP-treated macrophages increased the levels of M1 markers (Inos and Tnf-α) and reduced those of the enhanced M2 markers (Fizz-1, Ym-1, and arginase-1). EMP-treated macrophages significantly reduced Lewis lung carcinoma cell migration in a transwell migration assay and inhibited EL4-luc2 lymphoma proliferation. In our mechanism study, EMP was found to inhibit STAT3 phosphorylation in M2-polarized macrophages. These results suggest that EMP is effective in treating TAM-mediated tumor progression and metastasis. PMID:28811825

  1. Macrophage Ablation Reduces M2-Like Populations and Jeopardizes Tumor Growth in a MAFIA-Based Glioma Model.

    PubMed

    Gabrusiewicz, Konrad; Hossain, Mohammad B; Cortes-Santiago, Nahir; Fan, Xuejun; Kaminska, Bozena; Marini, Frank C; Fueyo, Juan; Gomez-Manzano, Candelaria

    2015-04-01

    Monocytes/macrophages are an influential component of the glioma microenvironment. However, understanding their diversity and plasticity constitute one of the most challenging areas of research due to the paucity of models to study these cells' inherent complexity. Herein, we analyzed the role of monocytes/macrophages in glioma growth by using a transgenic model that allows for conditional ablation of this cell population. We modeled glioma using intracranial GL261-bearing CSF-1R-GFP(+) macrophage Fas-induced apoptosis (MAFIA) transgenic mice. Conditional macrophage ablation was achieved by exposure to the dimerizer AP20187. Double immunofluorescence was used to characterize M1- and M2-like monocytes/macrophages during tumor growth and after conditional ablation. During glioma growth, the monocyte/macrophage population consisted predominantly of M2 macrophages. Conditional temporal depletion of macrophages reduced the number of GFP(+) cells, targeting mainly the repopulation of M2-polarized cells, and altered the appearance of M1-like monocytes/macrophages, which suggested a shift in the M1/M2 macrophage balance. Of interest, compared with control-treated mice, macrophage-depleted mice had a lower tumor mitotic index, microvascular density, and reduced tumor growth. These results demonstrated the possibility of studying in vivo the role and phenotype of macrophages in gliomas and suggested that transitory depletion of CSF-1R(+) population influences the reconstitutive phenotypic pool of these cells, ultimately suppressing tumor growth. The MAFIA model provides a much needed advance in defining the role of macrophages in gliomas. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  2. Macrophage Ablation Reduces M2-Like Populations and Jeopardizes Tumor Growth in a MAFIA-Based Glioma Model12

    PubMed Central

    Gabrusiewicz, Konrad; Hossain, Mohammad B.; Cortes-Santiago, Nahir; Fan, Xuejun; Kaminska, Bozena; Marini, Frank C.; Fueyo, Juan; Gomez-Manzano, Candelaria

    2015-01-01

    Monocytes/macrophages are an influential component of the glioma microenvironment. However, understanding their diversity and plasticity constitute one of the most challenging areas of research due to the paucity of models to study these cells' inherent complexity. Herein, we analyzed the role of monocytes/macrophages in glioma growth by using a transgenic model that allows for conditional ablation of this cell population. We modeled glioma using intracranial GL261-bearing CSF-1R–GFP+ macrophage Fas-induced apoptosis (MAFIA) transgenic mice. Conditional macrophage ablation was achieved by exposure to the dimerizer AP20187. Double immunofluorescence was used to characterize M1- and M2-like monocytes/macrophages during tumor growth and after conditional ablation. During glioma growth, the monocyte/macrophage population consisted predominantly of M2 macrophages. Conditional temporal depletion of macrophages reduced the number of GFP+ cells, targeting mainly the repopulation of M2-polarized cells, and altered the appearance of M1-like monocytes/macrophages, which suggested a shift in the M1/M2 macrophage balance. Of interest, compared with control-treated mice, macrophage-depleted mice had a lower tumor mitotic index, microvascular density, and reduced tumor growth. These results demonstrated the possibility of studying in vivo the role and phenotype of macrophages in gliomas and suggested that transitory depletion of CSF-1R+ population influences the reconstitutive phenotypic pool of these cells, ultimately suppressing tumor growth. The MAFIA model provides a much needed advance in defining the role of macrophages in gliomas. PMID:25925380

  3. M2/M1 Ratio of Tumor Associated Macrophages and PPAR-gamma Expression in Uveal Melanomas with Class 1 and Class 2 Molecular Profiles

    PubMed Central

    Herwig, Martina C.; Bergstrom, Chris; Wells, Jill R.; Höller, Tobias; Grossniklaus, Hans E.

    2012-01-01

    Macrophages have been found to be negative predictors of outcome in patients with uveal melanoma. In particular, recent studies point towards a disease-progressing role of proangiogenic M2 macrophages in melanomas with monosomy 3. Although most studies implicate a protective effect of PPAR-gamma activation in tumors, PPAR-gamma has also been shown to promote the polarization of M1 macrophages towards the M2 phenotype. The purpose of this investigation was first, to characterize the phenotype of tumor infiltrating macrophages and second, to study PPAR-gamma expression in uveal melanomas with molecular gene expression profile as prognostic predictors for patients’ outcome. Twenty specimens from patients with uveal melanoma were analyzed for clinical and histologic tumor characteristics. The molecular RNA profile (class 1 or class 2) was commercially determined. Using immunohistochemical techniques, the specimens were dual labeled for CD68 and CD163. CD68+CD163− M1 macrophages and CD68+CD163+ M2 macrophages were analyzed in ten high power fields sparing macrophage-poor areas and a mean value was calculated for each tumor. The tumors were immunostained for von Willebrand factor and the mean vascular density (MVD) was analyzed according to Foss. To assess the proliferative rate of each tumor, Ki67 expression was evaluated in ten high power fields followed by calculation of a mean value. Expression of PPAR-gamma was evaluated using a score from 0 (no staining) to 3 (tumor entirely stained). Statistical analysis and a respective correlation was made between histologic characteristics, molecular profile, type of tumor infiltrating macrophages (M1 versus M2), MVD, proliferative rate, and PPAR-gamma expression. Our results showed a correlation between the ratio of M2/M1 macrophages and the molecular profile with a ratio of approximately 1 corresponding to molecular class 1 and a ratio of approximately 2 corresponding to molecular class 2 (p=0.01). The ratio of M2/M1

  4. M2-polarized macrophages contribute to neovasculogenesis, leading to relapse of oral cancer following radiation

    PubMed Central

    Okubo, Makiko; Kioi, Mitomu; Nakashima, Hideyuki; Sugiura, Kei; Mitsudo, Kenji; Aoki, Ichiro; Taniguchi, Hideki; Tohnai, Iwai

    2016-01-01

    Despite the fact that radiation is one of the standard therapies in the treatment of patients with oral cancer, tumours can recur even in the early stages of the disease, negatively impacting prognosis and quality of life. We previously found that CD11b+ bone marrow-derived cells (BMDCs) were recruited into human glioblastoma multiforme (GBM), leading to re-organization of the vasculature and tumour regrowth. However, it is not yet known how these cells contribute to tumour vascularization. In the present study, we investigated the role of infiltrating CD11b+ myeloid cells in the vascularization and recurrence of oral squamous cell carcinoma (OSCC). In a xenograft mouse model, local irradiation caused vascular damage and hypoxia in the tumour and increased infiltration of CD11b+ myeloid cells. These infiltrating cells showed characteristics of M2 macrophages (M2Mφs) and are associated with the promotion of vascularization. M2Mφs promoted tumour progression in recurrence after irradiation compared to non-irradiated tumours. In addition, we found that CD11b+ myeloid cells, as well as CD206+ M2Mφs, are increased during recurrence after radiotherapy in human OSCC specimens. Our findings may lead to the development of potential clinical biomarkers or treatment targets in irradiated OSCC patients. PMID:27271009

  5. Macrophage M1/M2 polarization dynamically adapts to changes in cytokine microenvironments in Cryptococcus neoformans infection.

    PubMed

    Davis, Michael J; Tsang, Tiffany M; Qiu, Yafeng; Dayrit, Jeremy K; Freij, Joudeh B; Huffnagle, Gary B; Olszewski, Michal A

    2013-06-18

    The outcome of cryptococcal pneumonia correlates with local macrophage polarization status, as M1 and M2 polarization marks protective and nonprotective responses, respectively. Overall, pulmonary macrophage polarization status changes over time during a cryptococcal infection. This could have been caused by repolarization of individual macrophages or by a replacement of M2-polarized cells by new M1-polarized cells. To explore the ability of macrophages to change between polarization states, we conducted a series of experiments using in vitro macrophages. Coculture of macrophages with Cryptococcus neoformans resulted in development of a weak M1-like phenotype, with modestly increased inducible nitric oxide synthase (iNOS) but lacking interleukin 6 (IL-6) induction. The C. neoformans-induced M1-like polarization state was plastic, as macrophages stimulated first with C. neoformans and then with gamma interferon (IFN-γ) or IL-4 expressed mRNA polarization patterns similar to those stimulated with cytokines alone. To further evaluate macrophage polarization plasticity, cytokine stimulatory conditions were established which fully polarized macrophages. IFN-γ and IL-4 stimulation differentially induced complete M1 and M2 polarization, defined by differential expression of marker mRNA panels, surface marker expression, and tumor necrosis factor alpha (TNF-α) protein production. Switching IFN-γ- to IL-4-stimulating conditions, and vice versa, resulted in uniform changes in profiles of polarization marker genes consistent with the most recent cytokine environment. Furthermore, the ability of sequentially stimulated macrophages to inhibit C. neoformans reflected the most recent polarizing condition, independent of previous polarization. Collectively, these data indicate that M1/M2 macrophage polarization phenotypes are highly plastic to external signals, and interventions which therapeutically repolarize macrophages could be beneficial for treatment of cryptococcosis.

  6. Zoledronic acid impairs stromal reactivity by inhibiting M2-macrophages polarization and prostate cancer-associated fibroblasts

    PubMed Central

    Comito, Giuseppina; Segura, Coral Pons; Taddei, Maria Letizia; Lanciotti, Michele; Serni, Sergio; Morandi, Andrea; Chiarugi, Paola; Giannoni, Elisa

    2017-01-01

    Zoledronic acid (ZA) is a biphosphonate used for osteoporosis treatment and also proved to be effective to reduce the pain induced by bone metastases when used as adjuvant therapy in solid cancers. However, it has been recently proposed that ZA could have direct anti-tumour effects, although the molecular mechanism is unknown. We herein unravel a novel anti-tumour activity of ZA in prostate cancer (PCa), by targeting the pro-tumorigenic properties of both stromal and immune cells. Particularly, we demonstrate that ZA impairs PCa-induced M2-macrophages polarization, reducing their pro-invasive effect on tumour cells and their pro-angiogenic features. Crucially, ZA administration reverts cancer associated fibroblasts (CAFs) activation by targeting the mevalonate pathway and RhoA geranyl-geranylation, thereby impairing smooth muscle actin-α fibers organization, a prerequisite of fibroblast activation. Moreover, ZA prevents the M2 macrophages-mediated activation of normal fibroblast, highlighting the broad efficacy of this drug on tumour microenvironment. These results are confirmed in a metastatic xenograft PCa mouse model in which ZA-induced stromal normalization impairs cancer-stromal cells crosstalk, resulting in a significant reduction of primary tumour growth and metastases. Overall these findings reinforce the efficacy of ZA as a potential therapeutic approach to reduce cancer aggressiveness, by abrogating the supportive role of tumour microenvironment. PMID:27223431

  7. IL-4/IL-13-mediated polarization of renal macrophages/dendritic cells to an M2a phenotype is essential for recovery from acute kidney injury.

    PubMed

    Zhang, Ming-Zhi; Wang, Xin; Wang, Yinqiu; Niu, Aolei; Wang, Suwan; Zou, Chenhang; Harris, Raymond C

    2017-02-01

    Cytokines IL-4 and IL-13 play important roles in polarization of macrophages/dendritic cells to an M2 phenotype, which is important for recovery from acute kidney injury. Both IL-4 and IL-13 activate JAK3/STAT6 signaling. In mice with diphtheria toxin receptor expression in proximal tubules (selective injury model), a relatively selective JAK3 inhibitor, tofacitinib, led to more severe kidney injury, delayed recovery from acute kidney injury, increased inflammatory M1 phenotype markers and decreased reparative M2 phenotype markers of macrophages/dendritic cells, and development of more severe renal fibrosis after diphtheria toxin administration. Similarly, there was delayed recovery and increased tubulointerstitial fibrosis in these diphtheria toxin-treated mice following tamoxifen-induced deletion of both IL-4 and IL-13, with increased levels of M1 and decreased levels of M2 markers in the macrophages/dendritic cells. Furthermore, deletion of IL-4 and IL-13 led to a decrease of tissue reparative M2a phenotype markers but had no effect on anti-inflammatory M2c phenotype markers. Deletion of IL-4 and IL-13 also inhibited recovery from ischemia-reperfusion injury in association with increased M1 and decreased M2 markers and promoted subsequent tubulointerstitial fibrosis. Thus, IL-4 and IL-13 are required to effectively polarize macrophages/dendritic cells to an M2a phenotype and to promote recovery from acute kidney injury.

  8. Monocyte Differentiation towards Protumor Activity Does Not Correlate with M1 or M2 Phenotypes

    PubMed Central

    Chimal-Ramírez, G. Karina; Espinoza-Sánchez, Nancy Adriana; Chávez-Sánchez, Luis; Arriaga-Pizano, Lourdes

    2016-01-01

    Macrophages facilitate breast cancer progression. Macrophages were initially classified as M1 or M2 based on their distinct metabolic programs and then expanded to include antitumoral (M1) and protumoral (M2) activities. However, it is still uncertain what markers define the pro- and antitumoral phenotypes and what conditions lead to their formation. In this study, monocytic cell lines and primary monocytes were subjected to commonly reported protocols of M1/M2 polarization and conditions known to engage monocytes into protumoral functions. The results showed that only IDO enzyme and CD86 M1 markers were upregulated correlating with M1 polarization. TNF-α, CCR7, IL-10, arginase I, CD36, and CD163 were expressed indistinguishably from M1 or M2 polarization. Similarly, protumoral engaging resulted in upregulation of both M1 and M2 markers, with conditioned media from the most aggressive breast cancer cell line promoting the greatest changes. In spite of the mixed phenotype, M1-polarized macrophages exhibited the highest expression/secretion of inflammatory mediators, many of which have previously been associated with breast cancer aggressiveness. These data argue that although the existence of protumoral macrophages is unquestionable, their associated phenotypes and the precise conditions driving their formation are still unclear, and those conditions may need both M1 and M2 stimuli. PMID:27376091

  9. Alcohol-Induced miR-27a Regulates Differentiation and M2 Macrophage Polarization of Normal Human Monocytes

    PubMed Central

    Saha, Banishree; Bruneau, Johanna C.; Kodys, Karen; Szabo, Gyongyi

    2015-01-01

    Alcohol abuse is a leading cause of liver disease characterized by liver inflammation, fatty liver, alcoholic hepatitis, or liver cirrhosis. Immunomodulatory effects of alcohol on monocytes and macrophages contribute to alcoholic liver disease. Alcohol use, an independent risk factor for progression of hepatitis C virus (HCV) infection–mediated liver disease, impairs host defense and alters cytokine production and monocyte/macrophage activation. We hypothesized that alcohol and HCV have synergistic effects on the phenotype and function of monocytes. Our data show that acute alcohol binge drinking in healthy volunteers results in increased frequency of CD16+ and CD68+ and M2-type (CD206+, dendritic cell [DC]-SIGN+–expressing and IL-10–secreting) circulating CD14+ monocytes. Expression of HCV-induced CD68 and M2 markers (CD206 and DC-SIGN) in normal monocytes was further enhanced in the presence of alcohol. The levels of microRNA (miR)-27a was significantly upregulated in monocytes cultured in the presence of alcohol or alcohol and HCV as compared with HCV alone. The functional role of miR-27a in macrophage polarization was demonstrated by transfecting monocytes with an miR-27a inhibitor that resulted in reduced alcohol- and HCV- mediated monocyte activation (CD14 and CD68 expression), polarization (CD206 and DC-SIGN expression), and IL-10 secretion. Over-expression of miR-27a in monocytes enhanced IL-10 secretion via activation of the ERK signaling pathway. We found that miR-27a promoted ERK phosphorylation by downregulating the expression of ERK inhibitor sprouty2 in monocytes. Thus, we identified that sprouty2 is a target of miR-27a in human monocytes. In summary, our study demonstrates the regulatory role of miR-27a in alcohol-induced monocyte activation and polarization. PMID:25716995

  10. Polarization dictates iron handling by inflammatory and alternatively activated macrophages

    PubMed Central

    Corna, Gianfranca; Campana, Lara; Pignatti, Emanuele; Castiglioni, Alessandra; Tagliafico, Enrico; Bosurgi, Lidia; Campanella, Alessandro; Brunelli, Silvia; Manfredi, Angelo A.; Apostoli, Pietro; Silvestri, Laura; Camaschella, Clara; Rovere-Querini, Patrizia

    2010-01-01

    Background Macrophages play a key role in iron homeostasis. In peripheral tissues, they are known to polarize into classically activated (or M1) macrophages and alternatively activated (or M2) macrophages. Little is known on whether the polarization program influences the ability of macrophages to store or recycle iron and the molecular machinery involved in the processes. Design and Methods Inflammatory/M1 and alternatively activated/M2 macrophages were propagated in vitro from mouse bone-marrow precursors and polarized in the presence of recombinant interferon-γ or interleukin-4. We characterized and compared their ability to handle radioactive iron, the characteristics of the intracellular iron pools and the expression of molecules involved in internalization, storage and export of the metal. Moreover we verified the influence of iron on the relative ability of polarized macrophages to activate antigen-specific T cells. Results M1 macrophages have low iron regulatory protein 1 and 2 binding activity, express high levels of ferritin H, low levels of transferrin receptor 1 and internalize – albeit with low efficiency -iron only when its extracellular concentration is high. In contrast, M2 macrophages have high iron regulatory protein binding activity, express low levels of ferritin H and high levels of transferrin receptor 1. M2 macrophages have a larger intracellular labile iron pool, effectively take up and spontaneously release iron at low concentrations and have limited storage ability. Iron export correlates with the expression of ferroportin, which is higher in M2 macrophages. M1 and M2 cells activate antigen-specific, MHC class II-restricted T cells. In the absence of the metal, only M1 macrophages are effective. Conclusions Cytokines that drive macrophage polarization ultimately control iron handling, leading to the differentiation of macrophages into a subset which has a relatively sealed intracellular iron content (M1) or into a subset endowed with

  11. Chemical transfection of dye-conjugated microRNA precursors for microRNA functional analysis of M2 macrophages

    PubMed Central

    Ng, Yee Seng; Roca, Hernan; Fuller, David; Sud, Sudha

    2012-01-01

    Background MicroRNAs (miRNAs) are short non-coding ribonucleic acids known to affect gene expression at the translational level and there is mounting evidence that miRNAs play a role in the function of tumor-associated macrophages (TAMs). To aid the functional analyses of miRNAs in an in-vitro model of TAMs known as M2 macrophages, a transfection method to introduce artificial miRNA constructs or miRNA molecules into primary human monocytes is needed. Unlike differentiated macrophages or dendritic cells, undifferentiated primary human monocytes have been known to show resistance to lentiviral transduction. To circumvent this challenge, other techniques such as electroporation and chemical transfection have been used in other applications to deliver small gene constructs into human monocytes. To date, no studies have compared these two methods objectively to evaluate their suitability in the miRNA functional analysis of M2 macrophages. Results Of the methods tested, the electroporation of miRNA-construct containing plasmids and the chemical transfection of miRNA precursor molecules are the most efficient approaches. The use of a silencer siRNA labeling kit (Ambion) to conjugate Cy 3 fluorescence dyes to the precursor molecules allowed the isolation of successfully transfected cells with fluorescence-activated cell sorting. The chemical transfection of these dye-conjugated miRNA precursors yield an efficiency of 37.5 ± 0.6% and a cell viability of 74 ± 1%. RNA purified from the isolated cells demonstrated good quality, and was fit for subsequent mRNA expression qPCR analysis. While electroporation of plasmids containing miRNA constructs yield transfection efficiencies comparable to chemical transfection of miRNA precursors, these electroporated primary monocytes seemed to have lost their potential for differentiation. Conclusions Among the most common methods of transfection, the chemical transfection of dye-conjugated miRNA precursors was determined to be the best

  12. The Synthetic Melanocortin (CKPV)2 Exerts Anti-Fungal and Anti-Inflammatory Effects against Candida albicans Vaginitis via Inducing Macrophage M2 Polarization

    PubMed Central

    Jia, Zhi-rong; Li, Xian-jing; Wang, Zhuo; Li, Li; Li, Yong-wen; Liu, Gen-yan; Tong, Ming-Qing; Li, Xiao-yi; Zhang, Guo-hui; Dai, Xiang-rong; He, Ling; Li, Zhi-yu; Cao, Cong; Yang, Yong

    2013-01-01

    In this study, we examined anti-fungal and anti-inflammatory effects of the synthetic melanocortin peptide (Ac-Cys-Lys-Pro-Val-NH2)2 or (CKPV)2 against Candida albicans vaginitis. Our in vitro results showed that (CKPV)2 dose-dependently inhibited Candida albicans colonies formation. In a rat Candida albicans vaginitis model, (CKPV)2 significantly inhibited vaginal Candida albicans survival and macrophages sub-epithelial mucosa infiltration. For mechanisms study, we observed that (CKPV)2 inhibited macrophages phagocytosis of Candida albicans. Meanwhile, (CKPV)2 administration inhibited macrophage pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) release, while increasing the arginase activity and anti-inflammatory cytokine IL-10 production, suggesting macrophages M1 to M2 polarization. Cyclic AMP (cAMP) production was also induced by (CKPV)2 administration in macrophages. These above effects on macrophages by (CKPV)2 were almost reversed by melanocortin receptor-1(MC1R) siRNA knockdown, indicating the requirement of MC1R in the process. Altogether, our results suggest that (CKPV)2 exerted anti-fungal and anti-inflammatory activities against Candida albicans vaginitis probably through inducing macrophages M1 to M2 polarization and MC1R activation. PMID:23457491

  13. Tumor-associated macrophages of the M2 phenotype contribute to progression in gastric cancer with peritoneal dissemination.

    PubMed

    Yamaguchi, Takahisa; Fushida, Sachio; Yamamoto, Yasuhiko; Tsukada, Tomoya; Kinoshita, Jun; Oyama, Katsunobu; Miyashita, Tomoharu; Tajima, Hidehiro; Ninomiya, Itasu; Munesue, Seiichi; Harashima, Ai; Harada, Shinichi; Yamamoto, Hiroshi; Ohta, Tetsuo

    2016-10-01

    Tumor-associated macrophages (TAMs) of the M2 phenotype are known to promote tumor proliferation and to be associated with a poor prognosis in numerous cancers. Here, we investigated whether M2 macrophages participate in the development of peritoneal dissemination in gastric cancer. The characteristics of peritoneal macrophages in gastric cancer patients with or without peritoneal dissemination were examined by flow cytometry and the real-time quantitative polymerase chain reaction. The effects of M2 macrophages on phenotypic changes of the gastric cancer cell line MKN45 were assessed with a direct or indirect co-culture system in vitro and an in vivo mouse xenograft model. The number of peritoneal macrophages with the M2 phenotype (CD68(+)CD163(+) or CD68(+)CD204(+)) was significantly higher in gastric cancer patients with peritoneal dissemination than in those without peritoneal dissemination. Higher expression of the M2-related messenger RNAs (IL-10, vascular endothelial growth factor A, vascular endothelial growth factor C, matrix metalloproteinase 1, and amphiregulin) and lower expression of M1-related messenger RNAs (TNF-α, CD80, CD86, and IL-12p40) were also confirmed in the TAMs. Macrophage co-culture with gastric cancer cells converted M1 phenotype into M2 phenotype. Moreover, the coexistence of MKN45 cells with M2 macrophages resulted in cancer cell proliferation and an acceleration of tumor growth in the xenograft model. Intraperitoneal TAMs in gastric cancer patients with peritoneal dissemination were polarized to the M2 phenotype, and could contribute to tumor proliferation and progression. Therefore, intraperitoneal TAMs are expected to be a promising target in the treatment of peritoneal dissemination in gastric cancer.

  14. Cardiosphere-Derived Cells Facilitate Heart Repair by Modulating M1/M2 Macrophage Polarization and Neutrophil Recruitment

    PubMed Central

    Hasan, Al Shaimaa; Luo, Lan; Yan, Chen; Zhang, Tian-Xia; Urata, Yoshishige; Goto, Shinji; Mangoura, Safwat A.; Abdel-Raheem, Mahmoud H.; Zhang, Shouhua; Li, Tao-Sheng

    2016-01-01

    Cardiosphere-derived cells (CDCs), one of the promising stem cell sources for myocardial repair, have been tested in clinical trials and resulted in beneficial effects; however, the relevant mechanisms are not fully understood. In this study, we examined the hypothesis that CDCs favor heart repair by switching the macrophages from a pro-inflammatory phenotype (M1) into a regulatory anti-inflammatory phenotype (M2). Macrophages from mice were cultured with CDCs-conditioned medium or with fibroblasts-conditioned medium as a control. Immunostaining showed that CDCs-conditioned medium significantly enhanced the expression of CD206 (a marker for M2 macrophages), but decreased the expression of CD86 (a marker for M1 macrophages) 3 days after culture. For animal studies, we used an acute myocardial infarction model of mice. We injected CDCs, fibroblasts, or saline only into the border zone of infarction. Then we collected the heart tissues for histological analysis 5 and 14 days after treatment. Compared with control animals, CDCs treatment significantly decreased M1 macrophages and neutrophils but increased M2 macrophages in the infarcted heart. Furthermore, CDCs-treated mice had reduced infarct size and fewer apoptotic cells compared to the controls. Our data suggest that CDCs facilitate heart repair by modulating M1/M2 macrophage polarization and neutrophil recruitment, which may provide a new insight into the mechanisms of stem cell-based myocardial repair. PMID:27764217

  15. Phosphocholine‐containing ligands direct CRP induction of M2 macrophage polarization independent of T cell polarization: Implication for chronic inflammatory states

    PubMed Central

    Cieslik, Katarzyna A.; Entman, Mark L.

    2016-01-01

    Introduction We studied monocyte transendothelial migration and subsequent polarization into M1/M2 macrophages in response to C‐reactive protein (CRP) with two disease‐related ligands: (1) phosphocholine (PC) and (2) multilamellar liposomes containing both unoxidized and oxidized forms of the lipid, phosphatidylcholine. These ligands differ in biological origin: PC is present on bacterial cell walls while oxidized lipids are present in atherogenic lipids. Methods We used an in vitro model of human monocyte transendothelial migration and assessed the polarization of monocytes and T cells and signaling through Fcγ receptors in monocytes. Results CRP without ligands did not promote M2 macrophage differentiation over background levels. However, when paired with either ligand, it increased M2 numbers. M2 differentiation was dependent on IL‐13, and in the case of CRP with PC, was associated with a Th2 response. Paradoxically, while CRP with PC initiated a Th2 response, the combination of liposomes with CRP resulted in a Th1 response without any change in Th2 numbers despite association with M2 macrophage polarization. To resolve the conundrum of an anti‐inflammatory macrophage response coexisting with a proinflammatory T cell response, we investigated signaling of CRP and its ligands through Fcγ receptors, which leads to macrophage activation independent of T cell signaling. We found that CRP plus PC acted via FcγRI, whereas CRP with liposomes bound to FcγRII. Both were activating signals as evidenced by SYK phosphorylation. Conclusion We conclude that CRP with ligands can promote M2 macrophage differentiation to fibroblasts through FcγR activation, and this may result in an anti‐inflammatory influence despite a proinflammatory T cell environment caused by oxidized lipids. The potential relationship of this mechanism to chronic inflammatory disease is discussed. PMID:27621811

  16. γδ T Cells Are Required for M2 Macrophage Polarization and Resolution of Ozone-Induced Pulmonary Inflammation in Mice.

    PubMed

    Mathews, Joel A; Kasahara, David I; Ribeiro, Luiza; Wurmbrand, Allison P; Ninin, Fernanda M C; Shore, Stephanie A

    2015-01-01

    We examined the role of γδ T cells in the induction of alternatively activated M2 macrophages and the resolution of inflammation after ozone exposure. Wildtype (WT) mice and mice deficient in γδ T cells (TCRδ-/- mice) were exposed to air or to ozone (0.3 ppm for up to 72h) and euthanized immediately or 1, 3, or 5 days after cessation of exposure. In WT mice, M2 macrophages accumulated in the lungs over the course of ozone exposure. Pulmonary mRNA abundance of the M2 genes, Arg1, Retnla, and Clec10a, also increased after ozone. In contrast, no evidence of M2 polarization was observed in TCRδ-/- mice. WT but not TCRδ-/- mice expressed the M2c polarizing cytokine, IL-17A, after ozone exposure and WT mice treated with an IL-17A neutralizing antibody exhibited attenuated ozone-induced M2 gene expression. In WT mice, ozone-induced increases in bronchoalveolar lavage neutrophils and macrophages resolved quickly after cessation of ozone exposure returning to air exposed levels within 3 days. However, lack of M2 macrophages in TCRδ-/- mice was associated with delayed clearance of inflammatory cells after cessation of ozone and increased accumulation of apoptotic macrophages in the lungs. Delayed restoration of normal lung architecture was also observed in TCRδ-/- mice. In summary, our data indicate that γδ T cells are required for the resolution of ozone-induced inflammation, likely because γδ T cells, through their secretion of IL-17A, contribute to changes in macrophage polarization that promote clearance of apoptotic cells.

  17. Increased intermediate M1-M2 macrophage polarization and improved cognition in mild cognitive impairment patients on ω-3 supplementation.

    PubMed

    Famenini, Sam; Rigali, Elizabeth A; Olivera-Perez, Henry M; Dang, Johnny; Chang, Michael To; Halder, Ramesh; Rao, Rammohan V; Pellegrini, Matteo; Porter, Verna; Bredesen, Dale; Fiala, Milan

    2017-01-01

    Monocyte/macrophages of patients with mild cognitive impairment (MCI) and Alzheimer disease (AD) are defective in phagocytosis and degradation amyloid β1-42 (Aβ1-42), but are improved by ω-3 fatty acids (ω-3s). The hypothesis of this study was that active Aβ1-42 phagocytosis by macrophages prevents brain amyloidosis and thus maintains cognition. We studied the effects of self-supplementation with a drink with ω-3s, antioxidants, and resveratrol on Mini-Mental State Examination (MMSE) scores, macrophage M1M2 phenotype [the ratio of inflammatory cluster of differentiation (CD)54+CD80 and proresolution markers CD163+CD206], and Aβ1-42 phagocytosis in patients initially diagnosed as having MCI or subjective cognitive impairment (SCI). At baseline, the median MMSE score in patients in both the apolipoprotein E (ApoE) ε3/ε3 and ApoE ε3/ε4 groups was 26.0 and macrophage Aβ1-42 phagocytosis was defective. The MMSE rate of change increased in the ApoE ε3/ε3 group a median 2.2 points per year (P = 0.015 compared to 0) but did not change in the ApoE ε3/ε4 group (P = 0.014 between groups). In the ApoE ε3/ε3 group, all patients remained cognitively stable or improved; in the ApoE ε3/ε4 group, 1 recovered from dementia, but 3 lapsed into dementia. The macrophage phenotype polarized in patients bearing ApoE ε3/ε3 to an intermediate (green zone) M1-M2 type at the rate of 0.226 U/yr, whereas in patients bearing ApoE ε3/ε4, polarization was negative (P = 0.08 between groups). The baseline M1M2 type in the extreme M1 (red zone) or M2 (white zone) was unfavorable for cognitive outcome. Aβ1-42 phagocytosis increased in both ApoE groups (P = 0.03 in each groups). In vitro, the lipidic mediator resolvin D1 (RvD1) down regulated the M1 type in patients with ApoE ε3/ε3 but in some patients with ε3/ε4, paradoxically up-regulated the M1 type. Antioxidant/ω-3/resveratrol supplementation was associated with favorable immune and cognitive responses in ApoE ε3/ε3

  18. Increased intermediate M1-M2 macrophage polarization and improved cognition in mild cognitive impairment patients on ω-3 supplementation

    PubMed Central

    Famenini, Sam; Rigali, Elizabeth A.; Olivera-Perez, Henry M.; Dang, Johnny; Chang, Michael To; Halder, Ramesh; Rao, Rammohan V.; Pellegrini, Matteo; Porter, Verna; Bredesen, Dale; Fiala, Milan

    2017-01-01

    Monocyte/macrophages of patients with mild cognitive impairment (MCI) and Alzheimer disease (AD) are defective in phagocytosis and degradation amyloid β1–42 (Aβ1–42), but are improved by ω-3 fatty acids (ω-3s). The hypothesis of this study was that active Aβ1–42 phagocytosis by macrophages prevents brain amyloidosis and thus maintains cognition. We studied the effects of self-supplementation with a drink with ω-3s, antioxidants, and resveratrol on Mini-Mental State Examination (MMSE) scores, macrophage M1M2 phenotype [the ratio of inflammatory cluster of differentiation (CD)54+CD80 and proresolution markers CD163+CD206], and Aβ1–42 phagocytosis in patients initially diagnosed as having MCI or subjective cognitive impairment (SCI). At baseline, the median MMSE score in patients in both the apolipoprotein E (ApoE) ε3/ε3 and ApoE ε3/ε4 groups was 26.0 and macrophage Aβ1–42 phagocytosis was defective. The MMSE rate of change increased in the ApoE ε3/ε3 group a median 2.2 points per year (P = 0.015 compared to 0) but did not change in the ApoE ε3/ε4 group (P = 0.014 between groups). In the ApoE ε3/ε3 group, all patients remained cognitively stable or improved; in the ApoE ε3/ε4 group, 1 recovered from dementia, but 3 lapsed into dementia. The macrophage phenotype polarized in patients bearing ApoE ε3/ε3 to an intermediate (green zone) M1-M2 type at the rate of 0.226 U/yr, whereas in patients bearing ApoE ε3/ε4, polarization was negative (P = 0.08 between groups). The baseline M1M2 type in the extreme M1 (red zone) or M2 (white zone) was unfavorable for cognitive outcome. Aβ1–42 phagocytosis increased in both ApoE groups (P = 0.03 in each groups). In vitro, the lipidic mediator resolvin D1 (RvD1) down regulated the M1 type in patients with ApoE ε3/ε3 but in some patients with ε3/ε4, paradoxically up-regulated the M1 type. Antioxidant/ω-3/resveratrol supplementation was associated with favorable immune and cognitive responses in

  19. Efficient clearance of early apoptotic cells by human macrophages requires M2c polarization and MerTK induction.

    PubMed

    Zizzo, Gaetano; Hilliard, Brendan A; Monestier, Marc; Cohen, Philip L

    2012-10-01

    Mer tyrosine kinase (MerTK) is a major macrophage apoptotic cell (AC) receptor. Its functional impairment promotes autoimmunity and atherosclerosis, whereas overexpression correlates with poor prognosis in cancer. However, little is known about mechanisms regulating MerTK expression in humans. We found that MerTK expression is heterogenous among macrophage subsets, being mostly restricted to anti-inflammatory M2c (CD14(+)CD16(+)CD163(+)CD204(+)CD206(+)CD209(-)) cells, differentiated by M-CSF or glucocorticoids. Small numbers of MerTK(+) "M2c-like" cells are also detectable among circulating CD14(bright)CD16(+) monocytes. MerTK expression levels adapt to changing immunologic environment, being suppressed in M1 and M2a macrophages and in dendritic cells. Remarkably, although glucocorticoid-induced differentiation is IL-10 independent, M-CSF-driven M2c polarization and related MerTK upregulation require IL-10. However, neither IL-10 alone nor TGF-β are sufficient to fully differentiate M2c (CD16(+)CD163(+)MerTK(+)) macrophages. M-CSF and IL-10, both released by T lymphocytes, may thus be required together to promote regulatory T cell-mediated induction of anti-inflammatory monocytes-macrophages. MerTK enables M2c macrophages to clear early ACs more efficiently than other macrophage subsets, and it mediates AC clearance by CD14(bright)CD16(+) monocytes. Moreover, M2c cells release Gas6, which in turn amplifies IL-10 secretion via MerTK. IL-10-dependent induction of the Gas6/MerTK pathway may, therefore, constitute a positive loop for M2c macrophage homeostasis and a critical checkpoint for maintenance of anti-inflammatory conditions. Our findings give new insight into human macrophage polarization and favor a central role for MerTK in regulation of macrophage functions. Eliciting M2c polarization can have therapeutic utility for diseases such as lupus, in which a defective AC clearance contributes to initiate and perpetuate the pathological process.

  20. Fucoidan inhibits CCL22 production through NF-κB pathway in M2 macrophages: a potential therapeutic strategy for cancer

    PubMed Central

    Sun, Jia; Sun, Jintang; Song, Bingfeng; Zhang, Lin; Shao, Qianqian; Liu, Yanguo; Yuan, Daoying; Zhang, Yun; Qu, Xun

    2016-01-01

    In tumor microenvironment, macrophages as a polarized M2 population promote tumor progression via releasing multiple cytokines and chemokines. A brown seaweed fucose-rich polysaccharide, fucoidan has antitumor activity and immune modulation through affecting tumor cells and lymphocytes. Here, we focused on the effect of fucoidan on macrophages especially M2 subtype. Our results demonstrated that fucoidan down-regulated partial cytokines and chemokines, especially a M2-type chemokine CCL22. Furthermore, fucoidan inhibited tumor cells migration and CD4+ T lymphocytes, especially Treg cells, recruitment induced by M2 macrophages conditioned medium through suppression of CCL22. Mechanismly, fucoidan inhibited CCL22 via suppressing p65-NF-κB phosphorylation and nuclear translocation. In addition, p38-MAPK and PI3K-AKT also affected the expression of CCL22 through differential modulation of NF-κB transcriptional activity. Taken together, we reveal an interesting result that fucoidan can inhibit tumor cell migration and lymphocytes recruitment by suppressing CCL22 in M2 macrophages via NF-κB-dependent transcription, which may be a novel and promising mechanism for tumor immunotherapy. PMID:27775051

  1. Increases of M2a macrophages and fibrosis in aging muscle are influenced by bone marrow aging and negatively regulated by muscle-derived nitric oxide.

    PubMed

    Wang, Ying; Wehling-Henricks, Michelle; Samengo, Giuseppina; Tidball, James G

    2015-08-01

    Muscle aging is associated with changes in myeloid cell phenotype that may influence age-related changes in muscle structure. We tested whether preventing age-related reductions in muscle neuronal nitric oxide synthase (nNOS) would obviate age-related changes in myeloid cells in muscle. Our findings show that muscle aging is associated with elevations of anti-inflammatory M2a macrophages that can increase muscle fibrosis. Expression of a muscle-specific nNOS transgene in mice prevented age-related increases in M2a macrophages. Transgene expression also reduced expression of collagens and decreased muscle fibrosis. The nNOS transgene prevented age-related increases in arginase-1 but did not influence TGFβ expression, indicating that the transgene may prevent age-related muscle fibrosis by inhibiting the arginase-dependent profibrotic pathway. Although aged satellite cells or fibro-adipogenic precursor (FAPs) cells also promote fibrosis, transgene expression had no effect on the expression of key signaling molecules that regulate fibrogenic activity of those cells. Finally, we tested whether increases in M2a macrophages and the associated increase in fibrosis were attributable to aging of myeloid lineage cells. Young bone marrow cells (BMCs) were transplanted into young or old mice, and muscles were collected 8 months later. Muscles of young mice receiving young BMCs showed no effect on M2a macrophage number or collagen accumulation compared to age-matched, nontransplanted controls. However, muscles of old mice receiving young BMCs showed fewer M2a macrophages and less accumulation of collagen. Thus, the age-related increase in M2a macrophages in aging muscle and the associated muscle fibrosis are determined in part by the age of bone marrow cells.

  2. Cathepsin S-mediated autophagic flux in tumor-associated macrophages accelerate tumor development by promoting M2 polarization

    PubMed Central

    2014-01-01

    Background Tumor-associated macrophages (TAMs) are the major component of tumor-infiltrating leukocytes. TAMs are heterogeneous, with distinct phenotypes influenced by the microenvironment surrounding tumor tissues, but relatively little is known about the key molecular in these cells that contribute to malignant phenotypes. Autophagic activity is a critical factor in tumor development that contributes to enhancing cellular fitness and survival in the hostile tumor microenvironment. However, the molecular basis and relations between autophagy and TAMs polarization remain unclear. Methods Cathepsin S (Cat S) expression was analyzed in human colon carcinoma and normal colon tissues. In vivo effects were evaluated using PancO2 subcutaneous tumor model and SL4 hepatic metastasis model. Immunofluorescence staining, flow cytometry and real-time PCR were done to examine TAMs polarization. Western blotting assay, transmission electron microscopy, mCherry-GFP-LC3 transfection and DQ-BSA degradation assays were carried out to determine its role in regulating autophagy. Results In the present study, we showed that the enhanced expression of Cat S correlated with the severity of histologic grade as well as clinical stage, metastasis, and recurrence, which are known indicators of a relatively poor prognosis of human colon carcinoma. Cat S knockout led to decreased tumor growth and metastasis. Moreover, Cat S knockout inhibited M2 macrophage polarization during tumor development. We further demonstrated that Cat S was required for not only autophagic flux but also the fusion processes of autophagosomes and lysosomes in TAMs. Importantly, we found that Cat S contributed to tumor development by regulating the M2 phenotype of TAMs through the activation of autophagy. Conclusions These results indicated that Cat S-mediated autophagic flux is an important mechanism for inducing M2-type polarization of TAMs, which leads to tumor development. These data provide strong evidence for a

  3. Myeloperoxidase-Oxidized LDLs Enhance an Anti-Inflammatory M2 and Antioxidant Phenotype in Murine Macrophages

    PubMed Central

    Sauvage, Aude; Van Steenbrugge, Martine

    2016-01-01

    Macrophages and oxidized LDLs play a key role in atherogenesis but their heterogeneity has been neglected up to now. Macrophages are prone to polarization and subsets of polarized macrophages have been described in atheromas. LDLs can be oxidized not only chemically by copper (Ox-LDLs) but also enzymatically by myeloperoxidase (MpOx-LDLs) resulting in oxidized LDLs poor in lipid peroxides. The effects of physiologically relevant myeloperoxidase-oxidized LDLs on macrophage polarization or on polarized macrophages remain largely unknown. In this study, the effects of LDLs on macrophage polarization were investigated by monitoring the expression of M1 and M2 genes following stimulation with native LDLs, Ox-LDLs, or MpOx-LDLs in RAW 264.7 cells. Except for MRC1, which is induced only by Ox-LDLs, MpOx-LDLs induced an overexpression of most of the selected marker genes at the mRNA level. MpOx-LDLs also modulate marker gene expression in polarized macrophages favoring notably anti-inflammatory Arg1 expression in M2 cells and also in the other phenotypes. Noteworthy, MpOx-LDLs were the most efficient to accumulate lipids intracellularly in (un)polarized macrophages whatever the phenotype. These data were largely confirmed in murine bone marrow-derived macrophages. Our data suggest that MpOx-LDLs were the most efficient to accumulate within cells and to enhance an anti-inflammatory and antioxidant phenotype in M2 cells and also in the other macrophage phenotypes. PMID:27656049

  4. Bifidobacterium pseudocatenulatum CECT7765 induces an M2 anti-inflammatory transition in macrophages from patients with cirrhosis.

    PubMed

    Moratalla, Alba; Caparrós, Esther; Juanola, Oriol; Portune, Kevin; Puig-Kröger, Amaya; Estrada-Capetillo, Lizbeth; Bellot, Pablo; Gómez-Hurtado, Isabel; Piñero, Paula; Zapater, Pedro; González-Navajas, José M; Such, José; Sanz, Yolanda; Francés, Rubén

    2016-01-01

    Patients with cirrhosis show recurrent access of bacterial products into the bloodstream inducing a multi-altered immunological status leading to relevant complications. We aimed at evaluating Bifidobacterium pseudocatenulatum CECT7765 effect on the host's macrophage function. Patients with cirrhosis and ascites were included. Granulocyte-macrophage colony-stimulating factor (GM-CSF) monocyte-derived and ascitic fluid (AF) macrophages were cultured with M-CSF, lipopolysaccharide (LPS) and/or the bifidobacterial strain. Pellets and supernatants were evaluated for gene expression of M1 and M2-related genes and cytokine secretion. Cell surface expression molecules were evaluated by flow cytometry. Kupffer cells from bile duct ligated and CCl4 rats were also evaluated. Experiments were run on GM-CSF blood-derived and AF macrophages from 10 patients with cirrhosis and 10 healthy donors. Different macrophage morphology was observed by optical microscopy in cells stimulated with bifidobacteria vs. LPS. M2-like expression of CD206, CD163 and CD16 was significantly increased in macrophages after stimulation with the bifidobacterial strain vs. LPS. B. pseudocatenulatum CECT7765 was able to significantly change the cytokine secretion pattern of blood-derived and AF macrophages and Kupffer cells from bile duct ligated and CCl4 cirrhotic rats compared to that induced by LPS. B. pseudocatenulatum CECT7765 was also effective in inducing a phenotype transition and a functional change from an M1- to an M2-related gene expression and cytokine secretion pattern in AF macrophages even after LPS-pretreatment. B. pseudocatenulatum CECT7765 did not reduce AF macrophage bacterial killing capacity. B. pseudocatenulatum CECT7765 induces a morphologic, phenotypic and functional transition towards an anti-inflammatory profile in GM-CSF monocyte-derived and AF macrophages from patients with cirrhosis that may help in controlling sustained inflammation in decompensated cirrhosis. Copyright

  5. The human fetal placenta promotes tolerance against the semiallogeneic fetus by inducing regulatory T cells and homeostatic M2 macrophages.

    PubMed

    Svensson-Arvelund, Judit; Mehta, Ratnesh B; Lindau, Robert; Mirrasekhian, Elahe; Rodriguez-Martinez, Heriberto; Berg, Göran; Lash, Gendie E; Jenmalm, Maria C; Ernerudh, Jan

    2015-02-15

    A successful pregnancy requires that the maternal immune system is instructed to a state of tolerance to avoid rejection of the semiallogeneic fetal-placental unit. Although increasing evidence supports that decidual (uterine) macrophages and regulatory T cells (Tregs) are key regulators of fetal tolerance, it is not known how these tolerogenic leukocytes are induced. In this article, we show that the human fetal placenta itself, mainly through trophoblast cells, is able to induce homeostatic M2 macrophages and Tregs. Placental-derived M-CSF and IL-10 induced macrophages that shared the CD14(+)CD163(+)CD206(+)CD209(+) phenotype of decidual macrophages and produced IL-10 and CCL18 but not IL-12 or IL-23. Placental tissue also induced the expansion of CD25(high)CD127(low)Foxp3(+) Tregs in parallel with increased IL-10 production, whereas production of IFN-γ (Th1), IL-13 (Th2), and IL-17 (Th17) was not induced. Tregs expressed the suppressive markers CTLA-4 and CD39, were functionally suppressive, and were induced, in part, by IL-10, TGF-β, and TRAIL. Placental-derived factors also limited excessive Th cell activation, as shown by decreased HLA-DR expression and reduced secretion of Th1-, Th2-, and Th17-associated cytokines. Thus, our data indicate that the fetal placenta has a central role in promoting the homeostatic environment necessary for successful pregnancy. These findings have implications for immune-mediated pregnancy complications, as well as for our general understanding of tissue-induced tolerance.

  6. M2-like tumor-associated macrophages drive vasculogenic mimicry through amplification of IL-6 expression in glioma cells

    PubMed Central

    Zhang, Lin; Xu, Yangyang; Sun, Jintang; Chen, Weiliang; Zhao, Lei; Ma, Chao; Wang, Qingjie; Sun, Jia; Huang, Bin; Zhang, Yun; Li, Xingang; Qu, Xun

    2017-01-01

    Vasculogenic mimicry (VM) has offered a new horizon for understanding tumor angiogenesis, but the mechanisms of VM in glioma progression have not been studied explicitly until now. As a significant component of immune infiltration in tumor microenvironment, macrophages have been demonstrated to play an important role in tumor growth and angiogenesis. However, whether macrophages could play a potential key role in glioma VM is still poorly understood. Herein we reported that both VM and CD163+ cells were associated with WHO grade and reduced patient survival, and VM channel counting was correlated to the number of infiltrated CD163+ cells in glioma specimens. In vitro studies of glioma cell lines implicated that M2-like macrophages (M2) promoted glioma VM. We found that conditional medium derived from M2 amplified IL-6 expression in glioma cells. Furthermore, our data indicated that IL-6 could promote glioma VM, as blocking IL-6 with neutralizing antibodies abrogated M2-mediated VM enhancement. In addition, the potent PKC inhibitor bisindolylmaleimide I could prevent M2-induced IL-6 upregulation and further inhibited glioma VM facilitation. Taken together, our results suggested that M2-like macrophages drove glioma VM through amplifying IL-6 secretion in glioma cells via PKC pathway. PMID:27903982

  7. ABCG1 regulates mouse adipose tissue macrophage cholesterol levels and ratio of M1 to M2 cells in obesity and caloric restriction.

    PubMed

    Wei, Hao; Tarling, Elizabeth J; McMillen, Timothy S; Tang, Chongren; LeBoeuf, Renée C

    2015-12-01

    In addition to triacylglycerols, adipocytes contain a large reserve of unesterified cholesterol. During adipocyte lipolysis and cell death seen during severe obesity and weight loss, free fatty acids and cholesterol become available for uptake and processing by adipose tissue macrophages (ATMs). We hypothesize that ATMs become cholesterol enriched and participate in cholesterol clearance from adipose tissue. We previously showed that ABCG1 is robustly upregulated in ATMs taken from obese mice and further enhanced by caloric restriction. Here, we found that ATMs taken from obese and calorie-restricted mice derived from transplantation of WT or Abcg1-deficient bone marrow are cholesterol enriched. ABCG1 levels regulate the ratio of classically activated (M1) to alternatively activated (M2) ATMs and their cellular cholesterol content. Using WT and Abcg1(-/-) cultured macrophages, we found that Abcg1 is most highly expressed by M2 macrophages and that ABCG1 deficiency is sufficient to retard macrophage chemotaxis. However, changes in myeloid expression of Abcg1 did not protect mice from obesity or impaired glucose homeostasis. Overall, ABCG1 modulates ATM cholesterol content in obesity and weight loss regimes leading to an alteration in M1 to M2 ratio that we suggest is due to the extent of macrophage egress from adipose tissue.

  8. NMAAP1 Expressed in BCG-Activated Macrophage Promotes M1 Macrophage Polarization.

    PubMed

    Liu, Qihui; Tian, Yuan; Zhao, Xiangfeng; Jing, Haifeng; Xie, Qi; Li, Peng; Li, Dong; Yan, Dongmei; Zhu, Xun

    2015-10-01

    Macrophages are divided into two subpopulations: classically activated macrophages (M1) and alternatively activated macrophages (M2). BCG (Bacilli Calmette-GuC)rin) activates disabled naC/ve macrophages to M1 macrophages, which act as inflammatory, microbicidal and tumoricidal cells through cell-cell contact and/or the release of soluble factors. Various transcription factors and signaling pathways are involved in the regulation of macrophage activation and polarization. We discovered that BCG-activated macrophages (BAM) expressed a new molecule, and we named it Novel Macrophage Activated Associated Protein 1 (NMAAP1). The current study found that the overexpression of NMAAP1 in macrophages results in M1 polarization with increased expression levels of M1 genes, such as inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-N1), Interleukin 6 (IL-6), Interleukin 12 (IL-12), Monocyte chemoattractant protein-1 (MCP-1) and Interleukin-1 beta (IL-1N2), and decreased expression of some M2 genes, such as Kruppel-like factor 4 (KLF4) and suppressor of cytokine signaling 1 (SOCS1), but not other M2 genes, including arginase-1 (Arg-1), Interleukin (IL-10), transforming growth factor beta (TGF-N2) and found in inflammatory zone 1 (Fizz1). Moreover, NMAAP1 overexpression in the RAW264.7 cell line increased cytotoxicity against MCA207 tumor cells, which depends on increased inflammatory cytokines rather than cell-cell contact. NMAAP1 also substantially enhanced the phagocytic ability of macrophages, which implies that NMAAP1 promoted macrophage adhesive and clearance activities. Our results indicate that NMAAP1 is an essential molecule that modulates macrophages phenotype and plays an important role in macrophage tumoricidal functions.

  9. Alternatively activated macrophages in infection and autoimmunity

    PubMed Central

    Fairweather, DeLisa; Cihakova, Daniela

    2009-01-01

    Macrophages are innate immune cells that play an important role in activation of the immune response and wound healing. Pathogens that require T helper-type 2 (Th2) responses for effective clearance, such as parasitic worms, are strong inducers of alternatively activated or M2 macrophages. However, infections such as bacteria and viruses that require Th1-type responses may induce M2 as a strategy to evade the immune system. M2 are particularly efficient at scavenging self tissues following injury through receptors like the mannose receptor and scavenger receptor-A. Thus, M2 may increase autoimmune disease by presenting self tissue to T cells. M2 may also exacerbate immune complex (IC)-mediated pathology and fibrosis, a hallmark of autoimmune disease in women, due to the release of profibrotic factors such as interleukin (IL)-1β, transforming growth factor-β, fibronectin and matrix metalloproteinases. We have found that M2 comprise anywhere from 30% to 70% of the infiltrate during acute viral or experimental autoimmune myocarditis, and shifts in M2 populations correlate with increased IC-deposition, fibrosis and chronic autoimmune pathology. Thus, women may be at an increased risk of M2-mediated autoimmunity due to estrogen’s ability to increase Th2 responses. PMID:19819674

  10. PTEN inhibits macrophage polarization from M1 to M2 through CCL2 and VEGF-A reduction and NHERF-1 synergism

    PubMed Central

    Li, Ning; Qin, Junfang; Lan, Lan; Zhang, Hongyao; Liu, Fang; Wu, Zhaozhen; Ni, Hong; Wang, Yue

    2015-01-01

    PTEN has been studied in several tumor models as a tumor suppressor. In this study, we explored the role of PTEN in the inhibition state of polarized M2 subtype of macrophage in tumor microenvironment (TME) and the underlying mechanisms. To elucidate the potential effect in TME, RAW 264.7 macrophages and 4T1 mouse breast cancer cells were co-cultured to reconstruct tumor microenvironment. After PTEN was down-regulated with shRNA, the expression of CCL2 and VEGF-A, which are definited to promote the formation of M2 macrophages, have a dramatically increase on the level of both gene and protein in co-cultured RAW 264.7 macrophages. And at the same time, NHERF-1 (Na+/H+ exchanger regulating factor-1), another tumor suppressor has a similar tendency to PTEN. Q-PCR and WB results suggested that PTEN and NHERF-1 were consistent with one another no matter at mRNA or protein level when exposed to the same stimulus. Coimmunoprecipitation and immunofluorescence techniques confirmed that PTEN and NHERF-1 were coprecipitated, and NHERF-1 protein expression was properly reduced with rCCL2 effect. In addition, cell immunofluorescence images revealed a profound transferance, in co-cultured RAW 264.7 macrophages, an up-regulation of NHERF-1 could promote the PTEN marked expression on the cell membrane, and this form for the interaction was not negligible. These observations illustrate PTEN with a certain synergy of NHERF-1, as well as down-regulation of CCL2 suppressing M2 macrophage transformation pathway. The results suggest that the activation of PTEN and NHERF-1 may impede the evolution of macrophages beyond the M1 into M2 phenotype in tumor microenvironment. PMID:25756512

  11. Update on the role of alternatively activated macrophages in asthma.

    PubMed

    Jiang, Zhilong; Zhu, Lei

    2016-01-01

    Lung macrophages link innate and adaptive immune responses during allergic airway inflammatory responses. Alveolar macrophages (AMs) and interstitial macrophages are two different phenotypes that differentially exert immunological function under physiological and pathological conditions. Exposure to pathogen induces polarization of AM cells into classically activated macrophages (M1 cells) and alternatively activated macrophages (M2 cells). M1 cells dominantly express proinflammatory cytokines such as TNF-α and IL-1 β and induce lung inflammation and tissue damage. M2 cells are further divided into M2a and M2c subsets. M2a cells dominantly produce allergic cytokines IL-4 and IL-13, but M2c cells dominantly produce anti-inflammatory cytokine IL-10. M2a and M2c cells are differently involved in initiation, inflammation resolution, and tissue remodeling in the different stages of asthma. Microenvironment dynamically influences polarization of AM cells. Cytokines, chemokines, and immune-regulatory cells interplay and affect the balance between the polarization of M1 and M2 cells, subsequently influencing disease progression. Thus, modulation of AM phenotypes through molecular intervention has therapeutic potential in the treatment of asthma and other allergic inflammatory diseases. This review updated recent advances in polarization and functional specialization of these macrophage subtypes with emphasis on modulation of polarization of M2 cells in asthma of human subjects and animal models.

  12. Update on the role of alternatively activated macrophages in asthma

    PubMed Central

    Jiang, Zhilong; Zhu, Lei

    2016-01-01

    Lung macrophages link innate and adaptive immune responses during allergic airway inflammatory responses. Alveolar macrophages (AMs) and interstitial macrophages are two different phenotypes that differentially exert immunological function under physiological and pathological conditions. Exposure to pathogen induces polarization of AM cells into classically activated macrophages (M1 cells) and alternatively activated macrophages (M2 cells). M1 cells dominantly express proinflammatory cytokines such as TNF-α and IL-1 β and induce lung inflammation and tissue damage. M2 cells are further divided into M2a and M2c subsets. M2a cells dominantly produce allergic cytokines IL-4 and IL-13, but M2c cells dominantly produce anti-inflammatory cytokine IL-10. M2a and M2c cells are differently involved in initiation, inflammation resolution, and tissue remodeling in the different stages of asthma. Microenvironment dynamically influences polarization of AM cells. Cytokines, chemokines, and immune-regulatory cells interplay and affect the balance between the polarization of M1 and M2 cells, subsequently influencing disease progression. Thus, modulation of AM phenotypes through molecular intervention has therapeutic potential in the treatment of asthma and other allergic inflammatory diseases. This review updated recent advances in polarization and functional specialization of these macrophage subtypes with emphasis on modulation of polarization of M2 cells in asthma of human subjects and animal models. PMID:27350756

  13. M2-Polarized Macrophages Compose Lupus Vulgaris Arising from a Bacillus Calmette-Guérin Vaccination Site

    PubMed Central

    Sato, Yota; Fujimura, Taku; Furudate, Sadanori; Kakizaki, Aya; Iizawa, Osamu; Aiba, Setsuya

    2016-01-01

    Since bacillus Calmette-Guérin (BCG) is an attenuated strain of Mycobacterium bovis, lupus vulgaris (LV) is reported as one of the rare complications after BCG vaccination, correlating with immunosuppression in the lesional skin. In this report, we describe a case of LV arising from the BCG vaccination site 22 years after vaccination. Interestingly, in the present case, granuloma cells were composed of M2-polarized macrophages. Our case might explain the contribution of M2-polarized macrophages to the biology of LV arising from a BCG vaccination site. PMID:28101015

  14. M2-Polarized Macrophages Compose Lupus Vulgaris Arising from a Bacillus Calmette-Guérin Vaccination Site.

    PubMed

    Sato, Yota; Fujimura, Taku; Furudate, Sadanori; Kakizaki, Aya; Iizawa, Osamu; Aiba, Setsuya

    2016-01-01

    Since bacillus Calmette-Guérin (BCG) is an attenuated strain of Mycobacterium bovis, lupus vulgaris (LV) is reported as one of the rare complications after BCG vaccination, correlating with immunosuppression in the lesional skin. In this report, we describe a case of LV arising from the BCG vaccination site 22 years after vaccination. Interestingly, in the present case, granuloma cells were composed of M2-polarized macrophages. Our case might explain the contribution of M2-polarized macrophages to the biology of LV arising from a BCG vaccination site.

  15. Lipopolysaccharide preconditioning facilitates M2 activation of resident microglia after spinal cord injury.

    PubMed

    Hayakawa, Kentaro; Okazaki, Rentaro; Morioka, Kazuhito; Nakamura, Kozo; Tanaka, Sakae; Ogata, Toru

    2014-12-01

    The inflammatory response following spinal cord injury (SCI) has both harmful and beneficial effects; however, it can be modulated for therapeutic benefit. Endotoxin/lipopolysaccharide (LPS) preconditioning, a well-established method for modifying the immune reaction, has been shown to attenuate damage induced by stroke and brain trauma in rodent models. Although such effects likely are conveyed by tissue-repairing functions of the inflammatory response, the mechanisms that control the effects have not yet been elucidated. The present study preconditioned C57BL6/J mice with 0.05 mg/kg of LPS 48 hr before inducing contusion SCI to investigate the effect of LPS preconditioning on the activation of macrophages/microglia. We found that LPS preconditioning promotes the polarization of M1/M2 macrophages/microglia toward an M2 phenotype in the injured spinal cord on quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and immunohistochemical analyses. Flow cytometric analyses reveal that LPS preconditioning facilitates M2 activation in resident microglia but not in infiltrating macrophages. Augmented M2 activation was accompanied by vascularization around the injured lesion, resulting in improvement in both tissue reorganization and functional recovery. Furthermore, we found that M2 activation induced by LPS preconditioning is regulated by interleukin-10 gene expression, which was preceded by the transcriptional activation of interferon regulatory factor (IRF)-3, as demonstrated by Western blotting and an IRF-3 binding assay. Altogether, our findings demonstrate that LPS preconditioning has a therapeutic effect on SCI through the modulation of M1/M2 polarization of resident microglia. The present study suggests that controlling M1/M2 polarization through endotoxin signal transduction could become a promising therapeutic strategy for various central nervous system diseases. © 2014 Wiley Periodicals, Inc.

  16. The involvement of M2 macrophage polarization inhibition in fenretinide-mediated chemopreventive effects on colon cancer.

    PubMed

    Dong, Rong; Gong, Yanling; Meng, Wen; Yuan, Meng; Zhu, Hong; Ying, Meidan; He, Qiaojun; Cao, Ji; Yang, Bo

    2017-03-01

    Clinical studies have shown that fenretinide (4-HPR) is an attractive chemopreventive agent for cancer treatment. However, to date, few studies have demonstrated the mechanism of the preventive effect of 4-HPR. In our current study, we revealed that 4-HPR could significantly suppress IL-4/IL-13 induced M2-like polarization of macrophages, which was demonstrated by the reduced expression of M2 surface markers, the down-regulation of M2 marker genes, and the inhibition of M2-like macrophages promoted angiogenesis. Mechanistically, our study suggested that the inhibition of the phosphorylation of STAT6, rather than the generation of oxidative stress, is involved in the 4-HPR-driven inhibition of M2 polarization. More intriguingly, by utilizing adenomatous polyposis coli (APC(min/+)) transgenic mice, we demonstrated that the tumorigenesis was dramatically decreased by 4-HPR treatment accompanied with fewer M2-like macrophages in the tumor tissues, thereby profoundly blocking tumor angiogenesis. These findings, for the first time, reveal the involvement of M2 polarization inhibition in 4-HPR-mediated chemoprevention, which provides a new point of insight and indicates the potential mechanism underlying the chemopreventive effect of 4-HPR. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  17. The Many Alternative Faces of Macrophage Activation.

    PubMed

    Hume, David A

    2015-01-01

    Monocytes and macrophages provide the first line of defense against pathogens. They also initiate acquired immunity by processing and presenting antigens and provide the downstream effector functions. Analysis of large gene expression datasets from multiple cells and tissues reveals sets of genes that are co-regulated with the transcription factors that regulate them. In macrophages, the gene clusters include lineage-specific genes, interferon-responsive genes, early inflammatory genes, and genes required for endocytosis and lysosome function. Macrophages enter tissues and alter their function to deal with a wide range of challenges related to development and organogenesis, tissue injury, malignancy, sterile, or pathogenic inflammatory stimuli. These stimuli alter the gene expression to produce "activated macrophages" that are better equipped to eliminate the cause of their influx and to restore homeostasis. Activation or polarization states of macrophages have been classified as "classical" and "alternative" or M1 and M2. These proposed states of cells are not supported by large-scale transcriptomic data, including macrophage-associated signatures from large cancer tissue datasets, where the supposed markers do not correlate with other. Individual macrophage cells differ markedly from each other, and change their functions in response to doses and combinations of agonists and time. The most studied macrophage activation response is the transcriptional cascade initiated by the TLR4 agonist lipopolysaccharide. This response is reviewed herein. The network topology is conserved across species, but genes within the transcriptional network evolve rapidly and differ between mouse and human. There is also considerable divergence in the sets of target genes between mouse strains, between individuals, and in other species such as pigs. The deluge of complex information related to macrophage activation can be accessed with new analytical tools and new databases that provide

  18. Macrophage activation and polarization: nomenclature and experimental guidelines.

    PubMed

    Murray, Peter J; Allen, Judith E; Biswas, Subhra K; Fisher, Edward A; Gilroy, Derek W; Goerdt, Sergij; Gordon, Siamon; Hamilton, John A; Ivashkiv, Lionel B; Lawrence, Toby; Locati, Massimo; Mantovani, Alberto; Martinez, Fernando O; Mege, Jean-Louis; Mosser, David M; Natoli, Gioacchino; Saeij, Jeroen P; Schultze, Joachim L; Shirey, Kari Ann; Sica, Antonio; Suttles, Jill; Udalova, Irina; van Ginderachter, Jo A; Vogel, Stefanie N; Wynn, Thomas A

    2014-07-17

    Description of macrophage activation is currently contentious and confusing. Like the biblical Tower of Babel, macrophage activation encompasses a panoply of descriptors used in different ways. The lack of consensus on how to define macrophage activation in experiments in vitro and in vivo impedes progress in multiple ways, including the fact that many researchers still consider there to be only two types of activated macrophages, often termed M1 and M2. Here, we describe a set of standards encompassing three principles-the source of macrophages, definition of the activators, and a consensus collection of markers to describe macrophage activation-with the goal of unifying experimental standards for diverse experimental scenarios. Collectively, we propose a common framework for macrophage-activation nomenclature. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Hepatitis B Virus Infection and Immunopathogenesis in a Humanized Mouse Model: Induction of Human-Specific Liver Fibrosis and M2-Like Macrophages

    PubMed Central

    Bility, Moses T.; Cheng, Liang; Zhang, Zheng; Luan, Yan; Li, Feng; Chi, Liqun; Zhang, Liguo; Tu, Zhengkun; Gao, Yanhang; Fu, Yangxin; Niu, Junqi; Wang, Fusheng; Su, Lishan

    2014-01-01

    The mechanisms of chronic HBV infection and immunopathogenesis are poorly understood due to a lack of a robust small animal model. Here we report the development of a humanized mouse model with both human immune system and human liver cells by reconstituting the immunodeficient A2/NSG (NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ mice with human HLA-A2 transgene) with human hematopoietic stem cells and liver progenitor cells (A2/NSG-hu HSC/Hep mice). The A2/NSG-hu HSC/Hep mouse supported HBV infection and approximately 75% of HBV infected mice established persistent infection for at least 4 months. We detected human immune responses, albeit impaired in the liver, chronic liver inflammation and liver fibrosis in infected animals. An HBV neutralizing antibody efficiently inhibited HBV infection and associated liver diseases in humanized mice. In addition, we found that the HBV mediated liver disease was associated with high level of infiltrated human macrophages with M2-like activation phenotype. Importantly, similar M2-like macrophage accumulation was confirmed in chronic hepatitis B patients with liver diseases. Furthermore, gene expression analysis showed that induction of M2-like macrophage in the liver is associated with accelerated liver fibrosis and necrosis in patients with acute HBV-induced liver failure. Lastly, we demonstrate that HBV promotes M2-like activation in both M1 and M2 macrophages in cell culture studies. Our study demonstrates that the A2/NSG-hu HSC/Hep mouse model is valuable in studying HBV infection, human immune responses and associated liver diseases. Furthermore, results from this study suggest a critical role for macrophage polarization in hepatitis B virus-induced immune impairment and liver pathology. PMID:24651854

  20. Hepatitis B virus infection and immunopathogenesis in a humanized mouse model: induction of human-specific liver fibrosis and M2-like macrophages.

    PubMed

    Bility, Moses T; Cheng, Liang; Zhang, Zheng; Luan, Yan; Li, Feng; Chi, Liqun; Zhang, Liguo; Tu, Zhengkun; Gao, Yanhang; Fu, Yangxin; Niu, Junqi; Wang, Fusheng; Su, Lishan

    2014-03-01

    The mechanisms of chronic HBV infection and immunopathogenesis are poorly understood due to a lack of a robust small animal model. Here we report the development of a humanized mouse model with both human immune system and human liver cells by reconstituting the immunodeficient A2/NSG (NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl)/SzJ mice with human HLA-A2 transgene) with human hematopoietic stem cells and liver progenitor cells (A2/NSG-hu HSC/Hep mice). The A2/NSG-hu HSC/Hep mouse supported HBV infection and approximately 75% of HBV infected mice established persistent infection for at least 4 months. We detected human immune responses, albeit impaired in the liver, chronic liver inflammation and liver fibrosis in infected animals. An HBV neutralizing antibody efficiently inhibited HBV infection and associated liver diseases in humanized mice. In addition, we found that the HBV mediated liver disease was associated with high level of infiltrated human macrophages with M2-like activation phenotype. Importantly, similar M2-like macrophage accumulation was confirmed in chronic hepatitis B patients with liver diseases. Furthermore, gene expression analysis showed that induction of M2-like macrophage in the liver is associated with accelerated liver fibrosis and necrosis in patients with acute HBV-induced liver failure. Lastly, we demonstrate that HBV promotes M2-like activation in both M1 and M2 macrophages in cell culture studies. Our study demonstrates that the A2/NSG-hu HSC/Hep mouse model is valuable in studying HBV infection, human immune responses and associated liver diseases. Furthermore, results from this study suggest a critical role for macrophage polarization in hepatitis B virus-induced immune impairment and liver pathology.

  1. Modulation of macrophage activation and programming in immunity.

    PubMed

    Liu, Guangwei; Yang, Hui

    2013-03-01

    Macrophages are central mediators of the immune, contributing both to the initiation and the resolution of inflammation. The concept of macrophage activation and program has stimulated interest in its definition, and functional significance in homeostasis and diseases. It has been known that macrophages could be differently activated and programmed into different functional subtypes in response to different types of antigen stumuli or different kinds of cytokines present in the microenvironment and could thus profoundly influence immune responses, but little is known about the state and exact regulatory mechanism of macrophage activation and program from cell or molecular signaling level in immunity. In this review, we summarize the recent finding regarding the regulatory mechanism of macrophage activation and program toward M1 and M2, especially on M2 macrophages.

  2. The macrophage in HIV-1 infection: from activation to deactivation?

    PubMed

    Herbein, Georges; Varin, Audrey

    2010-04-09

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

  3. Modulation of the inflammatory response to chitosan through M2 macrophage polarization using pro-resolution mediators.

    PubMed

    Vasconcelos, Daniela P; Costa, Madalena; Amaral, Isabel F; Barbosa, Mário A; Águas, Artur P; Barbosa, Judite N

    2015-01-01

    Tissue engineering and regenerative medicine have created a demand for biomaterials with specific functions such as the ability to modify the host immune response. The objective of this study was to evaluate the effect of two different pro-resolution lipid mediators, lipoxin A4 (LxA4) and resolvin D1 (RvD1), in the modulation of the inflammatory response to biomaterials through M2 macrophage polarization. This was investigated in vivo using a mouse air-pouch model of inflammation. Our results demonstrated that both LxA4 and RvD1 are able to shift the macrophage response to implanted Ch scaffolds to an M2 reparative response. The injection of these pro-resolution mediators caused a decrease in inflammatory cells recruited to the implant site together with higher numbers of F4/80(+)/CD206(+) cells (M2 macrophages) and lower numbers of F4/80(+)/CCR7(+) cells (M1 macrophages); it also induced a general decrease in several pro-inflammatory cytokines; and caused a significant decrease in the thickness and area of the fibrous capsule formed around the implanted scaffolds. In conclusion, the use of either LxA4 or RvD1 allowed the in vivo control of macrophage phenotypic profile and thus may play a significant role in regenerative medicine applications, namely through modulation of the inflammatory response.

  4. Silencing MicroRNA-155 Attenuates Cardiac Injury and Dysfunction in Viral Myocarditis via Promotion of M2 Phenotype Polarization of Macrophages.

    PubMed

    Zhang, Yingying; Zhang, Mengying; Li, Xueqin; Tang, Zongsheng; Wang, Xiangmin; Zhong, Min; Suo, Qifeng; Zhang, Yao; Lv, Kun

    2016-03-02

    Macrophage infiltration is a hallmark feature of viral myocarditis. As studies have shown that microRNA-155 regulates the differentiation of macrophages, we aimed to investigate the role of microRNA-155 in VM. We report that silencing microRNA-155 protects mice from coxsackievirus B3 induced myocarditis. We found that microRNA-155 expression was upregulated and localized primarily in heart-infiltrating macrophages and CD4(+) T lymphocytes during acute myocarditis. In contrast with wildtype (WT) mice, microRNA-155(-/-) mice developed attenuated viral myocarditis, which was characterized by decreased cardiac inflammation and decreased intracardiac CD45(+) leukocytes. Hearts of microRNA-155(-/-) mice expressed decreased levels of the IFN-γ and increased levels of the cytokines IL-4 and IL-13. Although total CD4(+) and regulatory T cells were unchanged in miR-155(-/-) spleen proportionally, the activation of T cells and CD4(+) T cell proliferation in miR-155(-/-) mice were significantly decreased. Beyond the acute phase, microRNA-15(5-/-) mice had reduced mortality and improved cardiac function during 5 weeks of follow-up. Moreover, silencing microRNA-155 led to increased levels of alternatively-activated macrophages (M2) and decreased levels of classically-activated macrophages (M1) in the heart. Combined, our studies suggest that microRNA-155 confers susceptibility to viral myocarditis by affecting macrophage polarization, and thus may be a potential therapeutic target for viral myocarditis.

  5. Dendrosomal curcumin suppresses metastatic breast cancer in mice by changing m1/m2 macrophage balance in the tumor microenvironment.

    PubMed

    Shiri, Sadaf; Alizadeh, Ali Mohammad; Baradaran, Behzad; Farhanghi, Baharak; Shanehbandi, Dariush; Khodayari, Saeed; Khodayari, Hamid; Tavassoli, Abbas

    2015-01-01

    Curcumin, a lipid-soluble compound extracted from the plant Curcuma Longa, has been found to exert immunomodulatory effects via macrophages. However, most studies focus on the low bioavailability issue of curcumin by nano and microparticles, and thus the role of macrophages in the anticancer mechanism of curcumin has received little attention so far. We have previously shown the potential biocompatibility, biodegradability and anti-cancer effects of dendrosomal curcumin (DNC). In this study, twenty-seven BALB/c mice were equally divided into control as well as 40 and 80 mg/kg groups of DNC to investigate the involvement of macrophages in the antitumor effects of curcumin in a typical animal model of metastatic breast cancer. At the end of intervention, the tumor volume and weight were significantly reduced in DNC groups compared to control (P<0.05). Histopathological data showed the presence of macrophages in tumor and spleen tissues. Real-time PCR results showed that DNC increased the expression of STAT4 and IL-12 genes in tumor and spleen tissues in comparison with control (P<0.05), referring to the high levels of M1 macrophages. Furthermore treatment with DNC decreased STAT3, IL-10 and arginase I gene expression (P<0.05), indicating low levels of M2 macrophage. The results confirm the role of macrophages in the protective effects of dendrosomal curcumin against metastatic breast cancer in mice.

  6. The Many Alternative Faces of Macrophage Activation

    PubMed Central

    Hume, David A.

    2015-01-01

    Monocytes and macrophages provide the first line of defense against pathogens. They also initiate acquired immunity by processing and presenting antigens and provide the downstream effector functions. Analysis of large gene expression datasets from multiple cells and tissues reveals sets of genes that are co-regulated with the transcription factors that regulate them. In macrophages, the gene clusters include lineage-specific genes, interferon-responsive genes, early inflammatory genes, and genes required for endocytosis and lysosome function. Macrophages enter tissues and alter their function to deal with a wide range of challenges related to development and organogenesis, tissue injury, malignancy, sterile, or pathogenic inflammatory stimuli. These stimuli alter the gene expression to produce “activated macrophages” that are better equipped to eliminate the cause of their influx and to restore homeostasis. Activation or polarization states of macrophages have been classified as “classical” and “alternative” or M1 and M2. These proposed states of cells are not supported by large-scale transcriptomic data, including macrophage-associated signatures from large cancer tissue datasets, where the supposed markers do not correlate with other. Individual macrophage cells differ markedly from each other, and change their functions in response to doses and combinations of agonists and time. The most studied macrophage activation response is the transcriptional cascade initiated by the TLR4 agonist lipopolysaccharide. This response is reviewed herein. The network topology is conserved across species, but genes within the transcriptional network evolve rapidly and differ between mouse and human. There is also considerable divergence in the sets of target genes between mouse strains, between individuals, and in other species such as pigs. The deluge of complex information related to macrophage activation can be accessed with new analytical tools and new databases

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

    PubMed

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

    2017-08-01

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

  8. Multiple myeloma cells recruit tumor-supportive macrophages through the CXCR4/CXCL12 axis and promote their polarization toward the M2 phenotype.

    PubMed

    Beider, Katia; Bitner, Hanna; Leiba, Merav; Gutwein, Odit; Koren-Michowitz, Maya; Ostrovsky, Olga; Abraham, Michal; Wald, Hanna; Galun, Eithan; Peled, Amnon; Nagler, Arnon

    2014-11-30

    Multiple myeloma (MM) cells specifically attract peripheral-blood monocytes, while interaction of MM with bone marrow stromal cells (BMSCs) significantly increased monocyte recruitment (p<0.01). The CXCL12 chemokine, produced by both the MM and BMSCs, was found to be a critical regulator of monocyte migration. CXCL12 production was up-regulated under MM-BMSCs co-culture conditions, whereas blockage with anti-CXCR4 antibodies significantly abrogated monocyte recruitment toward a MM-derived conditioned medium (p<0.01). Furthermore, elevated levels of CXCL12 were detected in MM, but not in normal BM samples, whereas malignant MM cells often represented the source of increased CXCL12 in the BM. Blood-derived macrophages effectively supported MM cells proliferation and protected them from chemotherapy-induced apoptosis. Importantly, MM cells affected macrophage polarization, elevating the expression of M2-related scavenger receptor CD206 in macrophages and blocking LPS-induced TNFα secretion (a hallmark of M1 response). Of note, MM-educated macrophages suppressed T-cell proliferation and IFNγ production in response to activation. Finally, increased numbers of CXCR4-expressing CD163+CD206+ macrophages were detected in the BM of MM patients (n=25) in comparison to MGUS (n=11) and normal specimens (n=8). Taken together, these results identify macrophages as important players in MM tumorogenicity, and recognize the CXCR4/CXCL12 axis as a critical regulator of MM-stroma interactions and microenvironment formation.

  9. Multiple myeloma cells recruit tumor-supportive macrophages through the CXCR4/CXCL12 axis and promote their polarization toward the M2 phenotype

    PubMed Central

    Beider, Katia; Bitner, Hanna; Leiba, Merav; Gutwein, Odit; Koren-Michowitz, Maya; Ostrovsky, Olga; Abraham, Michal; Wald, Hanna; Galun, Eithan; Peled, Amnon; Nagler, Arnon

    2014-01-01

    Multiple myeloma (MM) cells specifically attract peripheral-blood monocytes, while interaction of MM with bone marrow stromal cells (BMSCs) significantly increased monocyte recruitment (p<0.01). The CXCL12 chemokine, produced by both the MM and BMSCs, was found to be a critical regulator of monocyte migration. CXCL12 production was up-regulated under MM-BMSCs co-culture conditions, whereas blockage with anti-CXCR4 antibodies significantly abrogated monocyte recruitment toward a MM-derived conditioned medium (p<0.01). Furthermore, elevated levels of CXCL12 were detected in MM, but not in normal BM samples, whereas malignant MM cells often represented the source of increased CXCL12 in the BM. Blood-derived macrophages effectively supported MM cells proliferation and protected them from chemotherapy-induced apoptosis. Importantly, MM cells affected macrophage polarization, elevating the expression of M2-related scavenger receptor CD206 in macrophages and blocking LPS-induced TNFα secretion (a hallmark of M1 response). Of note, MM-educated macrophages suppressed T-cell proliferation and IFNγ production in response to activation. Finally, increased numbers of CXCR4-expressing CD163+CD206+ macrophages were detected in the BM of MM patients (n=25) in comparison to MGUS (n=11) and normal specimens (n=8). Taken together, these results identify macrophages as important players in MM tumorogenicity, and recognize the CXCR4/CXCL12 axis as a critical regulator of MM-stroma interactions and microenvironment formation. PMID:25526031

  10. The Protectin PCTR1 Is Produced by Human M2 Macrophages and Enhances Resolution of Infectious Inflammation

    PubMed Central

    Ramon, Sesquile; Dalli, Jesmond; Sanger, Julia M.; Winkler, Jeremy W.; Aursnes, Marius; Tungen, Jørn E.; Hansen, Trond V.; Serhan, Charles N.

    2017-01-01

    Inflammation and its natural resolution are host-protective responses triggered by infection or injury. The resolution phase of inflammation is regulated by enzymatically produced specialized pro-resolving mediators. We recently identified a new class of peptide-conjugated specialized pro-resolving mediators that carry potent tissue regenerative actions that belong to the protectin family and are coined protectin conjugates in tissue regeneration (PCTR). Herein, with the use of microbial-induced peritonitis in mice and liquid chromatography-tandem mass spectrometry–based lipid mediator metabololipidomics, we found that PCTR1 is temporally regulated during self-resolving infection. When administered at peak of inflammation, PCTR1 enhanced macrophage recruitment and phagocytosis of Escherichia coli, decreased polymorphonuclear leukocyte infiltration, and counter-regulated inflammation-initiating lipid mediators, including prostaglandins. In addition, biologically produced PCTR1 promoted human monocyte and macrophage migration in a dose-dependent manner (0.001 to 10.0 nmol/L). We prepared PCTR1 via organic synthesis and confirmed that synthetic PCTR1 increased macrophage and monocyte migration, enhanced macrophage efferocytosis, and accelerated tissue regeneration in planaria. With human macrophage subsets, PCTR1 levels were significantly higher in M2 macrophages than in M1 phenotype, along with members of the resolvin conjugates in tissue regeneration and maresin conjugate families. In contrast, M1 macrophages gave higher levels of cysteinyl leukotrienes. Together, these results demonstrate that PCTR1 is a potent monocyte/macrophage agonist, regulating key anti-inflammatory and pro-resolving processes during bacterial infection. PMID:26878209

  11. A unique anti-CD115 monoclonal antibody which inhibits osteolysis and skews human monocyte differentiation from M2-polarized macrophages toward dendritic cells.

    PubMed

    Haegel, Hélène; Thioudellet, Christine; Hallet, Rémy; Geist, Michel; Menguy, Thierry; Le Pogam, Fabrice; Marchand, Jean-Baptiste; Toh, Myew-Ling; Duong, Vanessa; Calcei, Alexandre; Settelen, Nathalie; Preville, Xavier; Hennequi, Marie; Grellier, Benoit; Ancian, Philippe; Rissanen, Jukka; Clayette, Pascal; Guillen, Christine; Rooke, Ronald; Bonnefoy, Jean-Yves

    2013-01-01

    Cancer progression has been associated with the presence of tumor-associated M2-macrophages (M2-TAMs) able to inhibit anti-tumor immune responses. It is also often associated with metastasis-induced bone destruction mediated by osteoclasts. Both cell types are controlled by the CD115 (CSF-1R)/colony-stimulating factor-1 (CSF-1, M-CSF) pathway, making CD115 a promising target for cancer therapy. Anti-human CD115 monoclonal antibodies (mAbs) that inhibit the receptor function have been generated in a number of laboratories. These mAbs compete with CSF-1 binding to CD115, dramatically affecting monocyte survival and preventing osteoclast and macrophage differentiation, but they also block CD115/CSF-1 internalization and degradation, which could lead to potent rebound CSF-1 effects in patients after mAb treatment has ended. We thus generated and selected a non-ligand competitive anti-CD115 mAb that exerts only partial inhibitory effects on CD115 signaling without blocking the internalization or the degradation of the CD115/CSF-1 complex. This mAb, H27K15, affects monocyte survival only minimally, but downregulates osteoclast differentiation and activity. Importantly, it inhibits monocyte differentiation to CD163(+)CD64(+) M2-polarized suppressor macrophages, skewing their differentiation toward CD14(-)CD1a(+) dendritic cells (DCs). In line with this observation, H27K15 also drastically inhibits monocyte chemotactic protein-1 secretion and reduces interleukin-6 production; these two molecules are known to be involved in M2-macrophage recruitment. Thus, the non-depleting mAb H27K15 is a promising anti-tumor candidate, able to inhibit osteoclast differentiation, likely decreasing metastasis-induced osteolysis, and able to prevent M2 polarization of TAMs while inducing DCs, hence contributing to the creation of more efficient anti-tumor immune responses.

  12. Tumour hypoxia promotes melanoma growth and metastasis via High Mobility Group Box-1 and M2-like macrophages

    PubMed Central

    Huber, Roman; Meier, Barbara; Otsuka, Atsushi; Fenini, Gabriele; Satoh, Takashi; Gehrke, Samuel; Widmer, Daniel; Levesque, Mitchell P.; Mangana, Joanna; Kerl, Katrin; Gebhardt, Christoffer; Fujii, Hiroko; Nakashima, Chisa; Nonomura, Yumi; Kabashima, Kenji; Dummer, Reinhard; Contassot, Emmanuel; French, Lars E.

    2016-01-01

    Hypoxia is a hallmark of cancer that is strongly associated with invasion, metastasis, resistance to therapy and poor clinical outcome. Tumour hypoxia affects immune responses and promotes the accumulation of macrophages in the tumour microenvironment. However, the signals linking tumour hypoxia to tumour-associated macrophage recruitment and tumour promotion are incompletely understood. Here we show that the damage-associated molecular pattern High-Mobility Group Box 1 protein (HMGB1) is released by melanoma tumour cells as a consequence of hypoxia and promotes M2-like tumour-associated macrophage accumulation and an IL-10 rich milieu within the tumour. Furthermore, we demonstrate that HMGB1 drives IL-10 production in M2-like macrophages by selectively signalling through the Receptor for Advanced Glycation End products (RAGE). Finally, we show that HMGB1 has an important role in murine B16 melanoma growth and metastasis, whereas in humans its serum concentration is significantly increased in metastatic melanoma. Collectively, our findings identify a mechanism by which hypoxia affects tumour growth and metastasis in melanoma and depict HMGB1 as a potential therapeutic target. PMID:27426915

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

    PubMed

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

    2017-08-21

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

  14. Efficient identification of murine M2 macrophage peptide targeting ligands by phage display and next-generation sequencing

    PubMed Central

    Liu, Gary W.; Livesay, Brynn R.; Kacherovsky, Nataly A.; Cieslewicz, Maryelise; Lutz, Emi; Waalkes, Adam; Jensen, Michael C.; Salipante, Stephen J.; Pun, Suzie H.

    2015-01-01

    Peptide ligands are used to increase the specificity of drug carriers to their target cells and to facilitate intracellular delivery. One method to identify such peptide ligands, phage display, enables high-throughput screening of peptide libraries for ligands binding to therapeutic targets of interest. However, conventional methods for identifying target binders in a library by Sanger sequencing are low-throughput, labor-intensive, and provide a limited perspective (< 0.01%) of the complete sequence space. Moreover, the small sample space can be dominated by non-specific, preferentially amplifying “parasitic sequences” and plastic-binding sequences, which may lead to the identification of false positives or exclude the identification of target-binding sequences. To overcome these challenges, we employed next-generation Illumina sequencing to couple high-throughput screening and high-throughput sequencing, enabling more comprehensive access to the phage display library sequence space. In this work, we define the hallmarks of binding sequences in next-generation sequencing data, and develop a method that identifies several target-binding phage clones for murine, alternatively-activated (M2) macrophages with a high (100%) success rate: sequences and binding motifs were reproducibly present across biological replicates; binding motifs were identified across multiple unique sequences; and an unselected, amplified library accurately filtered out parasitic sequences. In addition, we validate the Multiple Em for Motif Elicitation tool as an efficient and principled means of discovering binding sequences. PMID:26161996

  15. Activation and dynamic network of the M2 muscarinic receptor

    PubMed Central

    Miao, Yinglong; Nichols, Sara E.; Gasper, Paul M.; Metzger, Vincent T.; McCammon, J. Andrew

    2013-01-01

    G-protein-coupled receptors (GPCRs) mediate cellular responses to various hormones and neurotransmitters and are important targets for treating a wide spectrum of diseases. Although significant advances have been made in structural studies of GPCRs, details of their activation mechanism remain unclear. The X-ray crystal structure of the M2 muscarinic receptor, a key GPCR that regulates human heart rate and contractile forces of cardiomyocytes, was determined recently in an inactive antagonist-bound state. Here, activation of the M2 receptor is directly observed via accelerated molecular dynamics simulation, in contrast to previous microsecond-timescale conventional molecular dynamics simulations in which the receptor remained inactive. Receptor activation is characterized by formation of a Tyr2065.58–Tyr4407.53 hydrogen bond and ∼6-Å outward tilting of the cytoplasmic end of transmembrane α-helix 6, preceded by relocation of Trp4006.48 toward Phe1955.47 and Val1995.51 and flipping of Tyr4307.43 away from the ligand-binding cavity. Network analysis reveals that communication in the intracellular domains is greatly weakened during activation of the receptor. Together with the finding that residue motions in the ligand-binding and G-protein-coupling sites of the apo receptor are correlated, this result highlights a dynamic network for allosteric regulation of the M2 receptor activation. PMID:23781107

  16. Effects of Activated Macrophages on Nocardia asteroides

    PubMed Central

    Filice, Gregory A.; Beaman, Blaine L.; Remington, Jack S.

    1980-01-01

    The mechanism(s) of host resistance against Nocardia asteroides has not been well defined. Since disease due to N. asteroides frequently occurs in patients with impaired cell-mediated immunity, we studied the interaction of N. asteroides with activated and control mouse peritoneal macrophages. Activated macrophages were from mice infected with Toxoplasma gondii or injected with Corynebacterium parvum. N. asteroides in the early stationary phase (>99% in the coccobacillary form) was used for challenge of macrophage monolayers. Growth of two strains of N. asteroides was markedly inhibited in activated macrophages, whereas N. asteroides grew well in control macrophages. Quantitation of macrophage-associated N. asteroides indicated that activated macrophages killed 40 to 50% of N. asteroides within 6 h (P < 0.002). In control macrophage preparations, it appeared as if Nocardia filaments extended from within macrophages to the outside, and many of these filaments appeared to have extended to and then grown through neighboring macrophages. In activated macrophage preparations, Nocardia remained in the coccobacillary form in most macrophages. Control macrophage monolayers were almost completely overgrown with and destroyed by Nocardia 20 h after challenge, whereas activated macrophage monolayers remained intact. Nocardia that grew in control macrophages were not acid-alcohol fast or only weakly so, whereas the few Nocardia that grew in activated macrophages were strongly acid-alcohol fast. Our results indicate that activated macrophages may be important in host defense against N. asteroides. Images Fig. 1 PMID:6991421

  17. Interleukin-17 Indirectly Promotes M2 Macrophage Differentiation through Stimulation of COX-2/PGE2 Pathway in the Cancer Cells

    PubMed Central

    Li, Qingli; Liu, Lunxu; Zhang, Qiuyang; Liu, Sen; Ge, Dongxia; You, Zongbing

    2014-01-01

    Purpose Interleukin-17 (IL-17) is a proinflammatory cytokine that plays important roles in inflammation, autoimmunity, and cancer. The purpose of this study was to determine if IL-17 indirectly regulates macrophage differentiation through up-regulation of cyclooxygenase-2 (COX-2) expression in the cancer cell lines. Materials and Methods Human cervical cancer HeLa, human lung cancer A549, and mouse prostate cancer Myc-CaP/CR cell lines were treated with recombinant IL-17; Western blot analysis, enzyme-linked immunosorbent assay, and quantitative real-time polymerase chain reaction analysis were utilized to examine the cellular responses. Results IL-17 up-regulated expression of COX-2 mRNA and protein in HeLa, A549, and Myc- CaP/CR cell lines. IL-17’s effects were mediated through nuclear factor-κB and ERK1/2 signaling pathways as the inhibitors of these pathways could inhibit IL-17- induced COX-2 expression. The conditional medium obtained from the cancer cells contained prostaglandin E2, the levels of which were increased by IL-17 treatment. When treated with the conditional medium, particularly with the IL-17-induced conditional medium, mouse RAW264.7 macrophages and human THP-1 monocytes expressed higher levels of IL-10 (a marker of M2 macrophages) than inducible nitric oxide synthase or tumor necrosis factor α (markers of M1 macrophages). In contrast, when RAW264.7 and THP-1 cells were treated directly with IL-17, expression of these marker genes was not markedly changed. Conclusion The results of this study suggest that IL-17 indirectly promotes M2 macrophage differentiation through stimulation of the COX-2/PGE2 pathway in the cancer cells, thus IL-17 plays an indirect role in regulating the tumor immune microenvironment. PMID:25038765

  18. Amphiregulin may be a new biomarker of classically activated macrophages.

    PubMed

    Meng, Chen; Liu, Guilin; Mu, Honglan; Zhou, Miaomiao; Zhang, Shihai; Xu, Younian

    2015-10-23

    Amphiregulin (Areg) participates in tissue repair and inflammation regulation. As important effector cells in inflammation, macrophages can be polarized to classically (M1) or alternatively (M2) activated phenotype with diverse functions in immunity. However, the relationship between Areg expression and macrophage activation is poorly understood. Here we report that Areg was significantly expressed in M1 but not in M2 macrophages. This was confirmed by analyses of RT-PCR and ELISA in peritoneal macrophages, and by evaluating protein expression in alveolar macrophages and RAW264.7 cells. Selective inhibitors of TLR4 (CLI-095) and MAP kinase, including Erk1/2 (PD98059), JNK (SP600125) and p38 (SB203580), significantly reduced Areg expression in M1 macrophages, suggesting that M1 macrophages produce Areg mainly through the TLR4-MAPK pathway, which is involved in the mechanism of M1 activation. When compared with productions of classical biomarkers of M1 macrophages, Areg expression was highly consistent in time series. Taken together, Areg may be an effective new biomarker of M1 macrophages.

  19. MicroRNA Cargo of Extracellular Vesicles from Alcohol-exposed Monocytes Signals Naive Monocytes to Differentiate into M2 Macrophages*

    PubMed Central

    Saha, Banishree; Momen-Heravi, Fatemeh; Kodys, Karen; Szabo, Gyongyi

    2016-01-01

    Membrane-coated extracellular vesicles (EVs) released by cells can serve as vehicles for delivery of biological materials and signals. Recently, we demonstrated that alcohol-treated hepatocytes cross-talk with immune cells via exosomes containing microRNA (miRNAs). Here, we hypothesized that alcohol-exposed monocytes can communicate with naive monocytes via EVs. We observed increased numbers of EVs, mostly exosomes, secreted by primary human monocytes and THP-1 monocytic cells in the presence of alcohol in a concentration- and time-dependent manner. EVs derived from alcohol-treated monocytes stimulated naive monocytes to polarize into M2 macrophages as indicated by increased surface expression of CD68 (macrophage marker), M2 markers (CD206 (mannose receptor) and CD163 (scavenger receptor)), secretion of IL-10, and TGFβ and increased phagocytic activity. miRNA profiling of the EVs derived from alcohol-treated THP-1 monocytes revealed high expression of the M2-polarizing miRNA, miR-27a. Treatment of naive monocytes with control EVs overexpressing miR-27a reproduced the effect of EVs from alcohol-treated monocytes on naive monocytes and induced M2 polarization, suggesting that the effect of alcohol EVs was mediated by miR-27a. We found that miR-27a modulated the process of phagocytosis by targeting CD206 expression on monocytes. Importantly, analysis of circulating EVs from plasma of alcoholic hepatitis patients revealed increased numbers of EVs that contained high levels of miR-27a as compared with healthy controls. Our results demonstrate the following: first, alcohol increases EV production in monocytes; second, alcohol-exposed monocytes communicate with naive monocytes via EVs; and third, miR-27a cargo in monocyte-derived EVs can program naive monocytes to polarize into M2 macrophages. PMID:26527689

  20. Therapeutic potential of carbohydrates as regulators of macrophage activation.

    PubMed

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

    2017-09-08

    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. Published by Elsevier Inc.

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

    SciTech Connect

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

    2013-11-15

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

  2. Angiogenic capacity of M1- and M2-polarized macrophages is determined by the levels of TIMP-1 complexed with their secreted proMMP-9

    PubMed Central

    Zajac, Ewa; Schweighofer, Bernhard; Kupriyanova, Tatyana A.; Juncker-Jensen, Anna; Minder, Petra

    2013-01-01

    A proangiogenic function of tissue-infiltrating monocytes/macrophages has long been attributed to their matrix metalloproteinase-9 zymogen (proMMP-9). Herein, we evaluated the capacity of human monocytes, mature M0 macrophages, and M1- and M2-polarized macrophages to induce proMMP-9-mediated angiogenesis. Only M2 macrophages induced angiogenesis at levels comparable with highly angiogenic neutrophils previously shown to release their proMMP-9 in a unique form, free of tissue inhibitor of metalloproteinases-1 (TIMP-1). Macrophage differentiation was accompanied by induction of low-angiogenic, TIMP-1–encumbered proMMP-9. However, polarization toward the M2, but not the M1 phenotype, caused a substantial downregulation of TIMP-1 expression, resulting in production of angiogenic, TIMP-deficient proMMP-9. Correspondingly, the angiogenic potency of M2 proMMP-9 was lost after its complexing with TIMP-1, whereas TIMP-1 silencing in M0/M1 macrophages rendered them both angiogenic. Similar to human cells, murine bone marrow–derived M2 macrophages also shut down their TIMP-1 expression and produced proMMP-9 unencumbered by TIMP-1. Providing proof that angiogenic capacity of murine M2 macrophages depended on their TIMP-free proMMP-9, Mmp9-null M2 macrophages were nonangiogenic, although their TIMP-1 was severely downregulated. Our study provides a unifying molecular mechanism for high angiogenic capacity of TIMP-free proMMP-9 that would be uniquely produced in a pathophysiological microenvironment by influxing neutrophils and/or M2 polarized macrophages. PMID:24174628

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

    PubMed

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

    2017-03-29

    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

  4. Toll-like Receptor-Mediated Responses by Placental Hofbauer Cells (HBCs): A Potential Pro-Inflammatory Role for Fetal M2 Macrophages*

    PubMed Central

    Young, Omar M.; Tang, Zhonghua; Niven-Fairchild, Tracy; Tadesse, Serkalem; Krikun, Graciela; Norwitz, Errol R.; Mor, Gil; Abrahams, Vikki M.; Guller, Seth

    2014-01-01

    Problem Microbial-driven responses in placenta are linked with adverse pregnancy outcomes. The role of Toll-like receptor (TLR) function in Hofbauer cells (HBCs), fetal macrophages of the placental villous core, remains understudied. Method of Study Flow cytometry and immunohistochemistry (IHC) were used to establish the phenotype of HBCs. Regulation of cytokine secretion in response to treatment with TLR agonists, and expression levels of TLRs and co-activators, were compared in HBCs, placental fibroblasts (FIBs), and human umbilical vein endothelial cells (HUVECs) using ELISA and qPCR. Results Although flow cytometry and IHC revealed HBCs to be M2 (anti-inflammatory) macrophages, LPS and Poly (I:C) treatments markedly enhanced IL-6 secretion by HBCs, and expression of TLR-2, TLR-3, TLR-4, CD14, and MD-2 were highest in HBCs. Conclusion These results indicate that although HBCs are M2 macrophages, inflammatory responses are induced through TLR-3 and TLR-4 in this cell type, suggesting a role in microbial-driven placental/fetal inflammation. PMID:25345551

  5. Opuntia ficus-indica seed attenuates hepatic steatosis and promotes M2 macrophage polarization in high-fat diet-fed mice.

    PubMed

    Kang, Jung-Woo; Shin, Jun-Kyu; Koh, Eun-Ji; Ryu, Hyojeong; Kim, Hyoung Ja; Lee, Sun-Mee

    2016-04-01

    Opuntia ficus-indica (L.) is a popular edible plant that possesses considerable nutritional value and exhibits diverse biological actions including anti-inflammatory and antidiabetic activities. In this study, we hypothesized that DWJ504, an extract of O ficus-indica seed, would ameliorate hepatic steatosis and inflammation by regulating hepatic de novo lipogenesis and macrophage polarization against experimental nonalcoholic steatohepatitis. Mice were fed a normal diet or a high-fat diet (HFD) for 10 weeks. DWJ504 (250, 500, and 1000 mg/kg) or vehicle (0.5% carboxymethyl cellulose) were orally administered for the last 4 weeks of the 10-week HFD feeding period. DWJ504 treatment remarkably attenuated HFD-induced increases in hepatic lipid content and hepatocellular damage. DWJ504 attenuated increases in sterol regulatory element-binding protein 1 and carbohydrate-responsive element-binding protein expression and a decrease in carnitine palmitoyltransferase 1A. Although DWJ504 augmented peroxisome proliferator-activated receptor α protein expression, it attenuated peroxisome proliferator-activated receptor γ expression. Moreover, DWJ504 promoted hepatic M2 macrophage polarization as indicated by attenuation of the M1 marker genes and enhancement of M2 marker genes. Finally, DWJ504 attenuated expression of toll-like receptor 4, nuclear factor κB, tumor necrosis factor α, interleukin 6, TIR-domain-containing adapter-inducing interferon β, and interferon β levels. Our results demonstrate that DWJ504 prevented intrahepatic lipid accumulation, induced M2 macrophage polarization, and suppressed the toll-like receptor 4-mediated inflammatory signaling pathway. Thus, DWJ504 has therapeutic potential in the prevention of nonalcoholic fatty liver disease. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Prognostic Implication of M2 Macrophages Are Determined by the Proportional Balance of Tumor Associated Macrophages and Tumor Infiltrating Lymphocytes in Microsatellite-Unstable Gastric Carcinoma

    PubMed Central

    Kim, Kyung-Ju; Wen, Xian-Yu; Yang, Han Kwang; Kim, Woo Ho; Kang, Gyeong Hoon

    2015-01-01

    Tumor associated macrophages are major inflammatory cells that play an important role in the tumor microenvironment. In this study, we investigated the prognostic significance of tumor associated macrophages (TAMs) in MSI-high gastric cancers using immunohistochemistry. CD68 and CD163 were used as markers for total infiltrating macrophages and M2-polarized macrophages, respectively. The density of CD68+ or CD163+ TAMs in four different areas (epithelial and stromal compartments of both the tumor center and invasive front) were analyzed in 143 cases of MSI-high advanced gastric cancers using a computerized image analysis system. Gastric cancers were scored as “0” or “1” in each area when the density of CD68+ and CD163+ TAMs was below or above the median value. Low density of CD68+ or CD163+ macrophages in four combined areas was closely associated with more frequent low-grade histology and the intestinal type tumor of the Lauren classification. In survival analysis, the low density of CD163+ TAMs was significantly associated with poor disease-free survival. In multivariate survival analysis, CD163+ TAMs in four combined areas, stromal and epithelial compartments of both tumor center and invasive front were independent prognostic indicator in MSI-high gastric cancers. In addition, the density of CD163+ TAMs correlated with tumor infiltrating lymphocytes (TILs). Our results indicate that the high density of CD163+ TAMs is an independent prognostic marker heralding prolonged disease-free survival and that the prognostic implication of CD163+ TAMs might be determined by the proportional balance of TAMs and TILs in MSI-high gastric cancers. PMID:26714314

  7. CD4-Positive T Cells and M2 Macrophages Dominate the Peritoneal Infiltrate of Patients with Encapsulating Peritoneal Sclerosis

    PubMed Central

    Habib, Sayed M.; Abrahams, Alferso C.; Korte, Mario R.; Zietse, Robert; de Vogel, Lisette L.; Boer, Walther H.; Dendooven, Amélie; Clahsen-van Groningen, Marian C.; Betjes, Michiel G. H.

    2015-01-01

    Background Encapsulating peritoneal sclerosis (EPS) is a severe complication of peritoneal dialysis (PD). Previously, it has been shown that infiltrating CD4-positive T cells and M2 macrophages are associated with several fibrotic conditions. Therefore, the characteristics of the peritoneal cell infiltrate in EPS may be of interest to understand EPS pathogenesis. In this study, we aim to elucidate the composition of the peritoneal cell infiltrate in EPS patients and relate the findings to clinical outcome. Study Design, Setting, and Participants We studied peritoneal membrane biopsies of 23 EPS patients and compared them to biopsies of 15 PD patients without EPS. The cellular infiltrate was characterized by immunohistochemistry to detect T cells(CD3-positive), CD4-positive (CD4+) and CD8-positive T cell subsets, B cells(CD20-positive), granulocytes(CD15-positive), macrophages(CD68-positive), M1(CD80-positive), and M2(CD163-positive) macrophages. Tissues were analysed using digital image analysis. Kaplan-Meier survival analysis was performed to investigate the survival in the different staining groups. Results The cellular infiltrate in EPS biopsies was dominated by mononuclear cells. For both CD3 and CD68, the median percentage of area stained was higher in biopsies of EPS as opposed to non-EPS patients (p<0.001). EPS biopsies showed a higher percentage of area stained for CD4 (1.29%(0.61-3.20)) compared to CD8 (0.71%(0.46-1.01), p = 0.04), while in the non-EPS group these cells were almost equally represented (respectively 0.28%(0.05-0.83) versus 0.22%(0.17-0.43), p = 0.97). The percentage of area stained for both CD80 and CD163 was higher in EPS than in non-EPS biopsies (p<0.001), with CD163+ cells being the most abundant phenotype. Virtually no CD20-positive and CD15-positive cells were present in biopsies of a subgroup of EPS patients. No relation was found between the composition of the mononuclear cell infiltrate and clinical outcome. Conclusions A

  8. Fibroblast Growth Factor-9 Enhances M2 Macrophage Differentiation and Attenuates Adverse Cardiac Remodeling in the Infarcted Diabetic Heart

    PubMed Central

    Singla, Dinender K.; Singla, Reetu D.; Abdelli, Latifa S.; Glass, Carley

    2015-01-01

    Inflammation has been implicated as a perpetrator of diabetes and its associated complications. Monocytes, key mediators of inflammation, differentiate into pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages upon infiltration of damaged tissue. However, the inflammatory cell types, which propagate diabetes progression and consequential adverse disorders, remain unclear. The current study was undertaken to assess monocyte infiltration and the role of fibroblast growth factor-9 (FGF-9) on monocyte to macrophage differentiation and cardioprotection in the diabetic infarcted heart. Db/db diabetic mice were assigned to sham, myocardial infarction (MI), and MI+FGF-9 groups. MI was induced by permanent coronary artery ligation and animals were subjected to 2D transthoracic echocardiography two weeks post-surgery. Immunohistochemical and immunoassay results from heart samples collected suggest significantly increased infiltration of monocytes (Mean ± SEM; MI: 2.02% ± 0.23% vs. Sham 0.75% ± 0.07%; p<0.05) and associated pro-inflammatory cytokines (TNF-α, MCP-1, and IL-6), adverse cardiac remodeling (Mean ± SEM; MI: 33% ± 3.04% vs. Sham 2.2% ± 0.33%; p<0.05), and left ventricular dysfunction (Mean ± SEM; MI: 35.4% ± 1.25% vs. Sham 49.19% ± 1.07%; p<0.05) in the MI group. Importantly, treatment of diabetic infarcted myocardium with FGF-9 resulted in significantly decreased monocyte infiltration (Mean ± SEM; MI+FGF-9: 1.39% ± 0.1% vs. MI: 2.02% ± 0.23%; p<0.05), increased M2 macrophage differentiation (Mean ± SEM; MI+FGF-9: 4.82% ± 0.86% vs. MI: 0.85% ± 0.3%; p<0.05) and associated anti-inflammatory cytokines (IL-10 and IL-1RA), reduced adverse remodeling (Mean ± SEM; MI+FGF-9: 11.59% ± 1.2% vs. MI: 33% ± 3.04%; p<0.05), and improved cardiac function (Fractional shortening, Mean ± SEM; MI+FGF-9: 41.51% ± 1.68% vs. MI: 35.4% ± 1.25%; p<0.05). In conclusion, our data suggest FGF-9 possesses novel therapeutic potential in its ability to

  9. Electrospun poly(hydroxybutyrate) scaffolds promote engraftment of human skin equivalents via macrophage M2 polarization and angiogenesis.

    PubMed

    Castellano, Delia; Sanchis, Ana; Blanes, María; Pérez Del Caz, Mª Dolores; Ruiz-Saurí, Amparo; Piquer-Gil, Marina; Pelacho, Beatriz; Marco, Bruno; Garcia, Nahuel; Ontoria-Oviedo, Imelda; Cambra, Vicente; Prosper, Felipe; Sepúlveda, Pilar

    2017-01-23

    Human dermo-epidermal skin equivalents (DE) comprising in vitro expanded autologous keratinocytes and fibroblasts are a good option for massive burn treatment. However, the lengthy expansion time required to obtain sufficient surface to cover an extensive burn together with the challenging surgical procedure limits their clinical use. The integration of DE and biodegradable scaffolds has been proposed in an effort to enhance their mechanical properties. Here, it is shown that poly(hydroxybutyrate) electrospun scaffolds (PHB) present good biocompatibility both in vitro and in vivo and are superior to poly-ε-caprolactone electrospun scaffolds as a substrate for skin reconstruction. Implantation of PHB scaffolds in healthy rats polarized macrophages to an M2-type that promoted constructive in vivo remodelling. Moreover, implantation of DE-PHB composites in a NOD/SCID mouse xenograft model resulted in engraftment accompanied by an increase in angiogenesis that favoured the survival of the human graft. Thus, PHB scaffolds are an attractive substrate for further exploration in skin reconstruction procedures, probably due in part to their greater angiogenic and M2 macrophage polarization properties. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

  10. M1 and M2 macrophage recruitment during tendon regeneration induced by amniotic epithelial cell allotransplantation in ovine.

    PubMed

    Mauro, Annunziata; Russo, Valentina; Di Marcantonio, Lisa; Berardinelli, Paolo; Martelli, Alessandra; Muttini, Aurelio; Mattioli, Mauro; Barboni, Barbara

    2016-04-01

    Recently, we have demonstrated that ovine amniotic epithelial cells (oAECs) allotransplanted into experimentally induced tendon lesions are able to stimulate tissue regeneration also by reducing leukocyte infiltration. Amongst leukocytes, macrophages (Mφ) M1 and M2 phenotype cells are known to mediate inflammatory and repairing processes, respectively. In this research it was investigated if, during tendon regeneration induced by AECs allotransplantation, M1Mφ and M2Mφ phenotype cells are recruited and differently distributed within the lesion site. Ovine AECs treated and untreated (Ctr) tendons were explanted at 7, 14, and 28 days and tissue microarchitecture was analyzed together with the distribution and quantification of leukocytes (CD45 positive), Mφ (CD68 pan positive), and M1Mφ (CD86, and IL12b) and M2Mφ (CD206, YM1 and IL10) phenotype related markers. In oAEC transplanted tendons CD45 and CD68 positive cells were always reduced in the lesion site. At day 14, oAEC treated tendons began to recover their microarchitecture, contextually a reduction of M1Mφ markers, mainly distributed close to oAECs, and an increase of M2Mφ markers was evidenced. CD206 positive cells were distributed near the regenerating areas. At day 28 oAECs treated tendons acquired a healthy-like structure with a reduction of M2Mφ. Differently, Ctr tendons maintained a disorganized morphology throughout the experimental time and constantly showed high values of M1Mφ markers. These findings indicate that M2Mφ recruitment could be correlated to tendon regeneration induced by oAECs allotransplantation. Moreover, these results demonstrate oAECs immunomodulatory role also in vivo and support novel insights into their allogeneic use underlying the resolution of tendon fibrosis.

  11. IL-33 and M2a alveolar macrophages promote lung defense against the atypical fungal pathogen Pneumocystis murina.

    PubMed

    Nelson, Michael P; Christmann, Benjamin S; Werner, Jessica L; Metz, Allison E; Trevor, Jennifer L; Lowell, Clifford A; Steele, Chad

    2011-02-15

    We have recently reported that mice deficient in the myeloid Src-family tyrosine kinases Hck, Fgr, and Lyn (Src triple knockout [TKO]) had augmented innate lung clearance of Pneumocystis murina that correlated with a higher ability of alveolar macrophages (AMs) from these mice to kill P. murina. In this article, we show that despite possessing enhanced killing, AMs from naive Src TKO mice did not demonstrate enhanced inflammatory responses to P. murina. We subsequently discovered that both AMs and lungs from P. murina-infected Src TKO mice expressed significantly greater levels of the M2a markers RELM-α and Arg1, and the M2a-associated chemokines CCL17 and CCL22 than did wild-type mice. IL-4 and IL-13, the primary cytokines that promote M2a polarization, were not differentially produced in the lungs between wild-type and Src TKO mice. P. murina infection in Src TKO mice resulted in enhanced lung production of the novel IL-1 family cytokine IL-33. Immunohistochemical analysis of IL-33 in lung tissue revealed localization predominantly in the nucleus of alveolar epithelial cells. We further demonstrate that experimental polarization of naive AMs to M2a resulted in more efficient killing of P. murina compared with untreated AMs, which was further enhanced by the addition of IL-33. Administration of IL-33 to C57BL/6 mice increased lung RELM-α and CCL17 levels, and enhanced clearance of P. murina, despite having no effect on the cellular composition of the lungs. Collectively, these results indicate that M2a AMs are potent effector cells against P. murina. Furthermore, enhancing M2a polarization may be an adjunctive therapy for the treatment of Pneumocystis.

  12. Macrophages migrate in an activation-dependent manner to chemokines involved in neuroinflammation

    PubMed Central

    2014-01-01

    Background In neuroinflammatory diseases, macrophages can play a dual role in the process of tissue damage, depending on their activation status (M1 / M2). M1 macrophages are considered to exert damaging effects to neurons, whereas M2 macrophages are reported to aid regeneration and repair of neurons. Their migration within the central nervous system may be of critical importance in the final outcome of neurodegeneration in neuroinflammatory diseases e.g. multiple sclerosis (MS). To provide insight into this process, we examined the migratory capacity of human monocyte-derived M1 and M2 polarised macrophages towards chemoattractants, relevant for neuroinflammatory diseases like MS. Methods Primary cultures of human monocyte-derived macrophages were exposed to interferon gamma and lipopolysaccharide (LPS) to evoke proinflammatory (M1) activation or IL-4 to evoke anti-inflammatory (M2) activation. In a TAXIScan assay, migration of M0, M1 and M2 towards chemoattractants was measured and quantified. Furthermore the adhesion capacity and the expression levels of integrins as well as chemokine receptors of M0, M1 and M2 were assessed. Alterations in cell morphology were analysed using fluorescent labelling of the cytoskeleton. Results Significant differences were observed between M1 and M2 macrophages in the migration towards chemoattractants. We show that M2 macrophages migrated over longer distances towards CCL2, CCL5, CXCL10, CXCL12 and C1q compared to non-activated (M0) and M1 macrophages. No differences were observed in the adhesion of M0, M1 and M2 macrophages to multiple matrix components, nor in the expression of integrins and chemokine receptors. Significant changes were observed in the cytoskeleton organization upon stimulation with CCL2, M0, M1 and M2 macrophages adopt a spherical morphology and the cytoskeleton is rapidly rearranged. M0 and M2 macrophages are able to form filopodia, whereas M1 macrophages only adapt a spherical morphology. Conclusions

  13. Parkin Regulates the Activity of Pyruvate Kinase M2*

    PubMed Central

    Liu, Kun; Li, Fanzhou; Han, Haichao; Chen, Yue; Mao, Zebin; Luo, Jianyuan; Zhao, Yingming; Zheng, Bin; Gu, Wei; Zhao, Wenhui

    2016-01-01

    Parkin, a ubiquitin E3 ligase, is mutated in most cases of autosomal recessive early onset Parkinson disease. It was discovered that Parkin is also mutated in glioblastoma and other human malignancies and that it inhibits tumor cell growth. Here, we identified pyruvate kinase M2 (PKM2) as a unique substrate for parkin through biochemical purification. We found that parkin interacts with PKM2 both in vitro and in vivo, and this interaction dramatically increases during glucose starvation. Ubiquitylation of PKM2 by parkin does not affect its stability but decreases its enzymatic activity. Parkin regulates the glycolysis pathway and affects the cell metabolism. Our studies revealed the novel important roles of parkin in tumor cell metabolism and provided new insight for therapy of Parkinson disease. PMID:26975375

  14. 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, or M1, macrophages have immune-stimulatory properties and cytotoxic function against tumor cells. Alternatively activated, or M2, macrophages have low cytotoxic function but high tissue-remodeling activity. There are also M2-like cells called tumor-associated macrophages (TAMs) that are responsible for many tumor-promoting activities. Blocking the function of TAMs inhibits tumorigenesis.

  15. CDDO-Me Redirects Activation of Breast Tumor Associated Macrophages

    PubMed Central

    Ball, Michael S.; Shipman, Emilie P.; Kim, Hyunjung; Liby, Karen T.; Pioli, Patricia A.

    2016-01-01

    Tumor-associated macrophages can account for up to 50% of the tumor mass in breast cancer patients and high TAM density is associated with poor clinical prognosis. Because TAMs enhance tumor growth, development, and metastatic potential, redirection of TAM activation may have significant therapeutic benefit. Our studies in primary human macrophages and murine breast TAMs suggest that the synthetic oleanane triterpenoid CDDO-methyl ester (CDDO-Me) reprograms the activation profile of TAMs from tumor-promoting to tumor-inhibiting. We show that CDDO-Me treatment inhibits expression of IL-10 and VEGF in stimulated human M2 macrophages and TAMs but increases expression of TNF-α and IL-6. Surface expression of CD206 and CD163, which are characteristic of M2 activation, is significantly attenuated by CDDO-Me. In contrast, CDDO-Me up-regulates surface expression of HLA-DR and CD80, which are markers of M1 activation, and importantly potentiates macrophage activation of autologous T cells but inhibits endothelial cell vascularization. These results show for the first time that CDDO-Me redirects activation of M2 macrophages and TAMs from immune-suppressive to immune-stimulatory, and implicate a role for CDDO-Me as an immunotherapeutic in the treatment of breast and potentially other types of cancer. PMID:26918785

  16. Attenuated EAN in TNF-α Deficient Mice Is Associated with an Altered Balance of M1/M2 Macrophages

    PubMed Central

    Zhang, Hong-Liang; Hassan, Mohammed Y.; Zheng, Xiang-Yu; Azimullah, Sheikh; Quezada, Hernan Concha; Amir, Naheed; Elwasila, Mohamed; Mix, Eilhard; Adem, Abdu; Zhu, Jie

    2012-01-01

    The role of tumor necrosis factor (TNF)-α and its receptors in neuroautoimmune and neuroinflammatory diseases has been controversial. On the basis of our previous studies, we hereby aimed to further clarify TNF-α’s mechanism of action and to explore the potential role of TNF-α receptor (TNFR)1 as a therapeutic target in experimental autoimmune neuritis (EAN). EAN was induced by immunization with P0 peptide 180–199 in TNF-α knockout (KO) mice and anti-TNFR1 antibodies were used to treat EAN. Particularly, the effects of TNF-α deficiency and TNFR1 blockade on macrophage functions were investigated. The onset of EAN in TNF-α KO mice was markedly later than that in wild type (WT) mice. From day 14 post immunization, the clinical signs of TNF-α KO mice were significantly milder than those of their WT counterparts. Further, we showed that the clinical severity of WT mice treated with anti-TNFR1 antibodies was less severe than that of the control WT mice receiving PBS. Nevertheless, no difference with regard to the clinical signs of EAN or inflammatory infiltration in cauda equina was seen between TNF-α KO and WT mice with EAN after blockade of TNFR1. Although TNF-α deficiency did not alter the proliferation of lymphocytes in response to either antigenic or mitogenic stimuli, it down-regulated the production of interleukin (IL)-12 and nitric oxide (NO), and enhanced the production of IL-10 in macrophages. Increased ratio of regulatory T cells (Tregs) and reduced production of interferon (IFN)-γ in cauda equina infiltrating cells, and elevated levels of IgG2b antibodies against P0 peptide 180–199 in sera were found in TNF-α KO mice with EAN. In conclusion, TNF-α deficiency attenuates EAN via altering the M1/M2 balance of macrophages. PMID:22666471

  17. Attenuated EAN in TNF-α deficient mice is associated with an altered balance of M1/M2 macrophages.

    PubMed

    Zhang, Hong-Liang; Hassan, Mohammed Y; Zheng, Xiang-Yu; Azimullah, Sheikh; Quezada, Hernan Concha; Amir, Naheed; Elwasila, Mohamed; Mix, Eilhard; Adem, Abdu; Zhu, Jie

    2012-01-01

    The role of tumor necrosis factor (TNF)-α and its receptors in neuroautoimmune and neuroinflammatory diseases has been controversial. On the basis of our previous studies, we hereby aimed to further clarify TNF-α's mechanism of action and to explore the potential role of TNF-α receptor (TNFR)1 as a therapeutic target in experimental autoimmune neuritis (EAN). EAN was induced by immunization with P0 peptide 180-199 in TNF-α knockout (KO) mice and anti-TNFR1 antibodies were used to treat EAN. Particularly, the effects of TNF-α deficiency and TNFR1 blockade on macrophage functions were investigated. The onset of EAN in TNF-α KO mice was markedly later than that in wild type (WT) mice. From day 14 post immunization, the clinical signs of TNF-α KO mice were significantly milder than those of their WT counterparts. Further, we showed that the clinical severity of WT mice treated with anti-TNFR1 antibodies was less severe than that of the control WT mice receiving PBS. Nevertheless, no difference with regard to the clinical signs of EAN or inflammatory infiltration in cauda equina was seen between TNF-α KO and WT mice with EAN after blockade of TNFR1. Although TNF-α deficiency did not alter the proliferation of lymphocytes in response to either antigenic or mitogenic stimuli, it down-regulated the production of interleukin (IL)-12 and nitric oxide (NO), and enhanced the production of IL-10 in macrophages. Increased ratio of regulatory T cells (Tregs) and reduced production of interferon (IFN)-γ in cauda equina infiltrating cells, and elevated levels of IgG2b antibodies against P0 peptide 180-199 in sera were found in TNF-α KO mice with EAN. In conclusion, TNF-α deficiency attenuates EAN via altering the M1/M2 balance of macrophages.

  18. All Trans Retinoic Acid, Transforming Growth Factor β and Prostaglandin E2 in Mouse Plasma Synergize with Basophil-Secreted Interleukin-4 to M2 Polarize Murine Macrophages

    PubMed Central

    Elisia, Ingrid; Lam, Vivian; Hsu, Brian E.; Lai, June; Luk, Beryl; Samudio, Ismael; Krystal, Gerald

    2016-01-01

    In previous studies we found that macrophages (MФs) from SH2-containing inositol-5′-phosphatase (SHIP) deficient mice are M2 polarized while their wild type (WT) counterparts are M1 polarized and that this difference in MФ phenotype can be recapitulated during in vitro derivation from bone marrow if mouse plasma (MP), but not fetal calf serum, is added to standard M-CSF-containing cultures. In the current study we investigated the mechanism by which MP skews SHIP-/- but not +/+ MФs to an M2 phenotype. Our results suggest that SHIP-/- basophils constitutively secrete higher levels of IL-4 than SHIP+/+ basophils and this higher level of IL-4 is sufficient to skew both SHIP+/+ and SHIP-/- MФs to an M2 phenotype, but only when MP is present to increase the sensitivity of the MФs to this level of IL-4. MP increases the IL-4 sensitivity of both SHIP+/+ and -/- MФs not by increasing cell surface IL-4 or CD36 receptor levels, but by triggering the activation of Erk and Akt and the production of ROS, all of which play a critical role in sensitizing MФs to IL-4-induced M2 skewing. Studies to identify the factor(s) in MP responsible for promoting IL-4-induced M2 skewing suggests that all-trans retinoic acid (ATRA), TGFβ and prostaglandin E2 (PGE2) all play a role. Taken together, these results indicate that basophil-secreted IL-4 plays an essential role in M2 skewing and that ATRA, TGFβ and PGE2 within MP collaborate to dramatically promote M2 skewing by acting directly on MФs to increase their sensitivity to IL-4. PMID:27977740

  19. Macrophages are recruited to hypoxic tumor areas and acquire a Pro-Angiogenic M2-Polarized phenotype via hypoxic cancer cell derived cytokines Oncostatin M and Eotaxin

    PubMed Central

    Tripathi, Chakrapani; Tewari, Brij Nath; Kanchan, Ranjana Kumari; Baghel, Khemraj Singh; Nautiyal, Naveen; Shrivastava, Richa; Kaur, Harbeer; Bhatt, Madan Lal Bramha; Bhadauria, Smrati

    2014-01-01

    TAMs, a unique and distinct M2-skewed myeloid population of tumor stroma, exhibiting pro-tumor functions is fast emerging as a potential target for anti-cancer immunotherapy. Macrophage-recruitment and M2-polarization represent key TAMs-related phenomenon that are amenable to therapeutic intervention. However successful translation of these approaches into effective therapeutic regimen requires better characterization of tumor-microenvironment derived signals that regulate macrophage recruitment and their polarization. Owing to hypoxic milieu being a persistent feature of tumor-microenvironment and a major contributor to malignancy and treatment resistance, the current study was planned with an aim to decipher tumor cell responses to hypoxia vis-a-vis macrophage homing and phenotype switching. Here, we show that hypoxia-primed cancer cells chemoattract and polarize macrophages to pro-angiogenic M2-polarized subtype via Eotaxin and Oncostatin M. Concordantly, hypoxic regions of human breast-cancer specimen exhibited elevated Eotaxin and Oncostatin M levels with concurrently elevated M2-macrophage content. Blockade of Eotaxin/Oncostatin M not only prevented hypoxic breast-cancer cells from recruiting and polarizing macrophages towards an M2-polarized phenotype and retarded tumor progression in 4T1/BALB/c-syngenic-mice-model of breast-cancer but also enhanced the efficacy of anti-angiogenic Bevacizumab. The findings established these two cytokines as novel targets for devising effective anticancer therapy particularly for tumors that are refractory or develop resistance to anti-angiogenic therapeutics. PMID:25051364

  20. Hyperhomocysteinemia promotes insulin resistance and adipose tissue inflammation in PCOS mice through modulating M2 macrophage polarization via estrogen suppression.

    PubMed

    Qi, Xinyu; Zhang, Bochun; Zhao, Yue; Li, Rong; Chang, Hsun-Ming; Pang, Yanli; Qiao, Jie

    2017-03-15

    It has been shown that serum homocycteine (Hcy) levels are higher in women with polycystic ovary syndrome (PCOS). However, the specific role of hyperhomocycteinemia (HHcy) in the development of PCOS has never been reported. Adipose tissue inflammation is featured by the infiltration of macrophages that plays a critical role in the pathogenesis of glucose and insulin intolerance. In this study, C57BL/6 mice were treated with dehydroepiandrosterone (DHEA) and/or high methionine diet to induce PCOS and HHcy mice models. We showed that DHEA induced a PCOS-like phenotypes, irregular estrous cycles, weight gain, abnormal sex hormone production, glucose and insulin resistance, and polycyctic ovaries. HHcy further intensified the effects DHEA on the metabolic, endocrinal, hormonal, and morphological changes in PCOS-like mice. In addition, the HHcy attenuated the DHEA-induced increase in serum estrogen levels in mice. Furthermore, HHcy may exacerbate the insulin resistance in the PCOS-like mice, most likely through modulating the macrophage M1/M2 polarization pathways via the suppression of estrogen. Most importantly, our clinical data showed that there were increases in serum Hcy levels in PCOS patients. These findings deepen our understanding of the pathological roles of HHcy in the development of PCOS and provide a promising target for PCOS therapy in clinical application.

  1. VAP-1-mediated M2 macrophage infiltration underlies IL-1β- but not VEGF-A-induced lymph- and angiogenesis.

    PubMed

    Nakao, Shintaro; Noda, Kousuke; Zandi, Souska; Sun, Dawei; Taher, Mahdi; Schering, Alexander; Xie, Fang; Mashima, Yukihiko; Hafezi-Moghadam, Ali

    2011-04-01

    Vascular adhesion protein-1 (VAP-1) contributes to inflammatory and angiogenic diseases, including cancer and age-related macular degeneration. It is expressed in blood vessels and contributes to inflammatory leukocyte recruitment. The cytokines IL-1β and vascular endothelial growth factor A (VEGF-A) modulate angiogenesis, lymphangiogenesis, and leukocyte infiltration. The lymphatic endothelium expresses intercellular adhesion molecule-1 and vascular adhesion molecule-1, which facilitate leukocyte transmigration into the lymphatic vessels. However, whether lymphatics express VAP-1 and whether they contribute to cytokine-dependent lymph- and angiogenesis are unknown. We investigated the role of VAP-1 in IL-1β- and VEGF-A-induced lymph- and angiogenesis using the established corneal micropocket assay. IL-1β increased VAP-1 expression in the inflamed cornea. Our in vivo molecular imaging revealed significantly higher VAP-1 expression in neovasculature than in the preexisting vessels. VAP-1 was expressed in blood but not lymphatic vessels in vivo. IL-1β-induced M2 macrophage infiltration and lymph- and angiogenesis were blocked by VAP-1 inhibition. In contrast, VEGF-A-induced lymph- and angiogenesis were unaffected by VAP-1 inhibition. Our results indicate a key role for VAP-1 in lymph- and angiogenesis-related macrophage recruitment. VAP-1 might become a new target for treatment of inflammatory lymph- and angiogenic diseases, including cancer.

  2. Dynamic, M2-Like Remodeling Phenotypes of CD11c+ Adipose Tissue Macrophages During High-Fat Diet–Induced Obesity in Mice

    PubMed Central

    Shaul, Merav E.; Bennett, Grace; Strissel, Katherine J.; Greenberg, Andrew S.; Obin, Martin S.

    2010-01-01

    OBJECTIVE To identify, localize, and determine M1/M2 polarization of epidydimal adipose tissue (eAT) macrophages (Φs) during high-fat diet (HFD)-induced obesity. RESEARCH DESIGN AND METHODS Male C57BL/6 mice were fed an HFD (60% fat kcal) or low-fat diet (LFD) (10% fat kcal) for 8 or 12 weeks. eATMΦs (F4/80+ cells) were characterized by in vivo fluorescent labeling, immunohistochemistry, fluorescence-activated cell sorting, and quantitative PCR. RESULTS Recruited interstitial macrophage galactose-type C-type lectin (MGL)1+/CD11c− and crown-like structure–associated MGL1−/CD11c+ and MGL1med/CD11c+ eATMΦs were identified after 8 weeks of HFD. MGL1med/CD11c+ cells comprised ∼65% of CD11c+ eATMΦs. CD11c+ eATMΦs expressed a mixed M1/M2 profile, with some M1 transcripts upregulated (IL-12p40 and IL-1β), others downregulated (iNOS, caspase-1, MCP-1, and CD86), and multiple M2 and matrix remodeling transcripts upregulated (arginase-1, IL-1Ra, MMP-12, ADAM8, VEGF, and Clec-7a). At HFD week 12, each eATMΦ subtype displayed an enhanced M2 phenotype as compared with HFD week 8. CD11c+ subtypes downregulated IL-1β and genes mediating antigen presentation (I-a, CD80) and upregulated the M2 hallmark Ym-1 and genes promoting oxidative metabolism (PGC-1α) and adipogenesis (MMP-2). MGL1med/CD11c+ eATMΦs upregulated additional M2 genes (IL-13, SPHK1, CD163, LYVE-1, and PPAR-α). MGL1med/CD11c+ ATMΦs expressing elevated PGC-1α, PPAR-α, and Ym-1 transcripts were selectively enriched in eAT of obese mice fed pioglitazone for 6 days, confirming the M2 features of the MGL1med/CD11c+ eATMΦ transcriptional profile and implicating PPAR activation in its elicitation. CONCLUSIONS These results 1) redefine the phenotypic potential of CD11c+ eATMΦs and 2) suggest previously unappreciated phenotypic and functional commonality between murine and human ATMΦs in the development of obesity and its complications. PMID:20185806

  3. Phenotypic, functional, and plasticity features of classical and alternatively activated human macrophages.

    PubMed

    Tarique, Abdullah A; Logan, Jayden; Thomas, Emma; Holt, Patrick G; Sly, Peter D; Fantino, Emmanuelle

    2015-11-01

    Macrophages are dynamic cells that mature under the influence of signals from the local microenvironment into either classically (M1) or alternatively (M2) activated macrophages with specific functional and phenotypic properties. Although the phenotypic identification of M1 and M2 macrophages is well established in mice, this is less clear for human macrophages. In addition, the persistence and reversibility of polarized human phenotypes is not well established. Human peripheral blood monocytes were differentiated into uncommitted macrophages (M0) and then polarized to M1 and M2 phenotypes using LPS/IFN-γ and IL-4/IL-13, respectively. M1 and M2 were identified as CD64(+)CD80(+) and CD11b(+)CD209(+), respectively, by flow cytometry. Polarized M1 cells secreted IP-10, IFN-γ, IL-8, TNF-α, IL-1β, and RANTES, whereas M2 cells secreted IL-13, CCL17, and CCL18. Functionally, M2 cells were highly endocytic. In cytokine-deficient medium, the polarized macrophages reverted back to the M0 state within 12 days. If previously polarized macrophages were given the alternative polarizing stimulus after 6 days of resting in cytokine-deficient medium, a switch in polarization was seen (i.e., M1 macrophages switched to M2 and expressed CD11b(+)CD209(+) and vice versa). In summary, we report phenotypic identification of human M1 and M2 macrophages, their functional characteristics, and their ability to be reprogrammed given the appropriate stimuli.

  4. Evidence That Ly6C(hi) Monocytes are Protective in Acute Ischemic Stroke by Promoting M2 Macrophage Polarization.

    PubMed

    Chu, Hannah X; Broughton, Brad R S; Kim, Hyun Ah; Lee, Seyoung; Drummond, Grant R; Sobey, Christopher G

    2015-07-01

    Ly6C(hi) monocytes are generally thought to exert a proinflammatory role in acute tissue injury, although their impact after injuries to the central nervous system is poorly defined. CC chemokine receptor 2 is expressed on Ly6C(hi) monocytes and plays an essential role in their extravasation and transmigration into the brain after cerebral ischemia. We used a selective CC chemokine receptor 2 antagonist, INCB3344, to assess the effect of Ly6C(hi) monocytes recruited into the brain early after ischemic stroke. Male C57Bl/6J mice underwent occlusion of the middle cerebral artery for 1 hour followed by 23 hours of reperfusion. Mice were administered either vehicle (dimethyl sulfoxide/carboxymethylcellulose) or INCB3344 (10, 30 or 100 mg/kg IP) 1 hour before ischemia and at 2 and 6 hours after ischemia. At 24 hours, we assessed functional outcomes, infarct volume, and quantified the immune cells in blood and brain by flow cytometry or immunofluorescence. Gene expression of selected inflammatory markers was assessed by quantitative polymerase chain reaction. Ly6C(hi) monocytes were increased 3-fold in the blood and 10-fold in the brain after stroke, and these increases were selectively prevented by INCB3344 in a dose-dependent manner. Mice treated with INCB3344 exhibited markedly worse functional outcomes and larger infarct volumes, in association with reduced M2 polarization and increased peroxynitrite production in macrophages, compared with vehicle-treated mice. Our data suggest that Ly6C(hi) monocytes exert an acute protective effect after ischemic stroke to limit brain injury and functional deficit that involves promotion of M2 macrophage polarization. © 2015 American Heart Association, Inc.

  5. Heme Oxygenase-1 Induction Prevents Autoimmune Diabetes in Association With Pancreatic Recruitment of M2-Like Macrophages, Mesenchymal Cells, and Fibrocytes.

    PubMed

    Husseini, Mahmoud; Wang, Gen-Sheng; Patrick, Christopher; Crookshank, Jennifer A; MacFarlane, Amanda J; Noel, J Ariana; Strom, Alexander; Scott, Fraser W

    2015-11-01

    Immunoregulatory and regenerative processes are activated in the pancreas during the development of type 1 diabetes (T1D) but are insufficient to prevent the disease. We hypothesized that the induction of cytoprotective heme oxygenase-1 (HO-1) by cobalt protophoryrin (CoPP) would prevent T1D by promoting anti-inflammatory and pro-repair processes. Diabetes-prone BioBreeding rats received ip CoPP or saline twice per week for 3 weeks, starting at 30 days and were monitored for T1D. Immunohistochemistry, confocal microscopy, quantitative RT-PCR, and microarrays were used to evaluate postinjection pancreatic changes at 51 days, when islet inflammation is first visible. T1D was prevented in CoPP-treated rats (29% vs 73%). Pancreatic Hmox1 was up-regulated along with islet-associated CD68(+)HO-1(+) cells, which were also observed in a striking peri-lobular interstitial infiltrate. Most interstitial cells expressed the mesenchymal marker vimentin and the hematopoietic marker CD34. Spindle-shaped, CD34(+)vimentin(+) cells coexpressed collagen V, characteristic of fibrocytes. M2 macrophage factors Krüppel-like factor 4, CD163, and CD206 were expressed by interstitial cells, consistent with pancreatic upregulation of several M2-associated genes. CoPP upregulated islet-regenerating REG genes and increased neogenic REG3β(+) and insulin(+) clusters. Thus, short-term induction of HO-1 promoted a protective M2-like milieu in the pancreas and recruited mesenchymal cells, M2 macrophages, and fibrocytes that imparted immunoregulatory and pro-repair effects, preventing T1D.

  6. Bone marrow-derived and peritoneal macrophages have different inflammatory response to oxLDL and M1/M2 marker expression – implications for atherosclerosis research

    PubMed Central

    Bisgaard, Line S.; Mogensen, Christina K.; Rosendahl, Alexander; Cucak, Helena; Nielsen, Lars Bo; Rasmussen, Salka E.; Pedersen, Tanja X.

    2016-01-01

    Macrophages are heterogeneous and can polarize into specific subsets, e.g. pro-inflammatory M1-like and re-modelling M2-like macrophages. To determine if peritoneal macrophages (PEMs) or bone marrow derived macrophages (BMDMs) resembled aortic macrophages from ApoE−/− mice, their M1/M2 phenotype, inflammatory status, and lipid metabolism signatures were compared. oxLDL accumulation was similar in PEMs and BMDMs. On protein expression level, BMDMs showed an M2-like CD206highCD11clow profile, while cholesterol loading led to enhanced CD11c expression and reduced MCP-1 secretion. In contrast, PEMs expressed low levels of CD206 and CD11c, and responded to cholesterol loading by increasing CD11c expression and MCP-1 secretion. mRNA expression of M1/M2 markers was higher in PEMS than BMDMs, while lipid metabolism genes were similarly expressed. Whole aorta flow cytometry showed an accumulation of M2-like CD206highCD11clow macrophages in advanced versus early atherosclerotic disease in ApoE−/− mice. In isolated lesions, mRNA levels of the M2 markers Socs2, CD206, Retnla, and IL4 were downregulated with increasing disease severity. Likewise, mRNA expression of lipid metabolism genes (SREBP2, ACSL1, SRB1, DGAT1, and cpt1a) was decreased in advanced versus early lesions. In conclusion, PEMs and BMDMs are phenotypically distinct and differ from macrophages in lesions with respect to expression of M1/M2 markers and lipid metabolism genes. PMID:27734926

  7. Interleukin-33 produced by M2 macrophages and other immune cells contributes to Th2 immune reaction of IgG4-related disease

    PubMed Central

    Furukawa, Sachiko; Moriyama, Masafumi; Miyake, Kensuke; Nakashima, Hitoshi; Tanaka, Akihiko; Maehara, Takashi; Iizuka-Koga, Mana; Tsuboi, Hiroto; Hayashida, Jun-Nosuke; Ishiguro, Noriko; Yamauchi, Masaki; Sumida, Takayuki; Nakamura, Seiji

    2017-01-01

    IgG4-related disease (IgG4-RD) is characterized by elevated serum IgG4 and marked infiltration of IgG4-positive cells in multiple organs. Interleukin-33 (IL-33) is a recently described cytokine that is secreted by damaged epithelial cells, macrophages, and dendritic cells, and potently activates helper T type 2 (Th2) immune responses, which have been suggested to play a major role in IgG4 production of IgG4-RD. Here, we assessed the expression of IL-33 and related molecules in the salivary glands (SGs) of patients with IgG4-RD versus that in patients with Sjögren’s syndrome (SS) and controls. Expression of IL-33 and its receptor (ST2) was strongly detected around ectopic germinal centers (GCs) in the SGs from patients with IgG4-RD, whereas IL-33 was expressed only in epithelial cells in patients with SS and controls. Moreover, IL-33 and CD68+/CD163+ macrophages were mainly distributed around ectopic GCs in patients with IgG4-RD. Double immunofluorescence staining showed that IL-33 expression co-localized with CD68+/CD163+ macrophages. Finally, mRNA expression levels of IL-33 showed a positive correlation to those of Th2 cytokines (IL-4 and IL-13) in patients with IgG4-RD. Our data suggest that IL-33 produced by M2 macrophages might contribute to the pathogenesis of IgG4-RD via aberrant activation of Th2 immune responses. PMID:28205524

  8. Functionally deficient mesenchymal stem cells reside in the bone marrow niche with M2-macrophages and amyloid-β protein adjacent to loose total joint implants.

    PubMed

    Margulies, Bryan S; DeBoyace, Sean D; Parsons, Adrienne M; Policastro, Connor G; Ee, Jessica S S; Damron, Timothy S

    2015-05-01

    We sought to demonstrate whether there is a difference in the local mesenchymal stem cells (MSC) niche obtained from patients undergoing their first total joint replacement surgery versus those patients undergoing a revision surgery for an failing total joint implant. Bone marrow aspirates collected from patients undergoing revision total joint arthroplasty were observed to be less clonal and the expression of PDGFRα, CD51, ALCAM, endoglin, CXCL12, nestin, and nucleostemin were decreased. Revision MSC were also less able to commit to an osteoblast-lineage or an adipocyte-lineage. Further, in revision MSC, OPG, and IL6 expression were increased. Monocytes, derived from revision whole marrow aspirates, were less capable of differentiating into osteoclasts, the cells implicated in the pathologic degradation of bone. Osteoclasts were also not observed in tissue samples collected adjacent to the implants of revision patients; however, the alternatatively activated M2-macrophage phenotype was observed in parallel with pathologic accumulations of amyloid-β, τ-protien and 3-nitrotyrosine. Despite the limited numbers of patients examined, our data suggest that nucleostemin may be a useful functional marker for MSC while the observation of M2-macrophage infiltration around the implant lays the foundation for future investigation into a novel mechanism that we propose is associated with loose total joint implants.

  9. Hypoxia induces macrophage polarization and re-education toward an M2 phenotype in U87 and U251 glioblastoma models

    PubMed Central

    Leblond, Marine M; Gérault, Aurélie N; Corroyer-Dulmont, Aurélien; MacKenzie, Eric T; Petit, Edwige; Bernaudin, Myriam; Valable, Samuel

    2016-01-01

    Hypoxia is a common feature of solid tumors, particularly in glioblastoma (GBM), and known to be a poor prognosis factor in GBM patients. The growth of GBM is also associated with a marked inflammation partially characterized by an accumulation of macrophage (MΦ) of the M2 phenotype. However, the transition between M1 MΦ (antitumoral) and M2 MΦ (protumoral) phenotypes is a dynamic process. We made the assumption that oxygen (O2) availability could be a major regulator of this transition and that the intratumoral O2 gradient is of importance. We evaluated, in vivo, the impact of hypoxia on MΦ tropism and polarization in two models of human GBM, well differentiated by their degree of hypoxia. MΦ migration in the tumor was more pronounced in the more hypoxic tumor of the two GBM models. In the more hypoxic of the models, we have shown that MΦ migrated at the tumor site only when hypoxia takes place. We also demonstrated that the acquisition of the M2 phenotype was clearly an evolving phenomenon with hypoxia as the major trigger for this transition. In support of these in vivo finding, M0 but also M1 MΦ cultured in moderate or severe hypoxia displayed a phenotype close to that of M2 MΦ whose phenotype was further reinforced by severe hypoxia. These results highlight the role of hypoxia in the aggressiveness of GBM, in part, by transforming MΦ such that a protumoral activity is expressed. PMID:26942063

  10. CCL2 Mediates Neuron-Macrophage Interactions to Drive Proregenerative Macrophage Activation Following Preconditioning Injury.

    PubMed

    Kwon, Min Jung; Shin, Hae Young; Cui, Yuexian; Kim, Hyosil; Thi, Anh Hong Le; Choi, Jun Young; Kim, Eun Young; Hwang, Dong Hoon; Kim, Byung Gon

    2015-12-02

    CNS neurons in adult mammals do not spontaneously regenerate axons after spinal cord injury. Preconditioning peripheral nerve injury allows the dorsal root ganglia (DRG) sensory axons to regenerate beyond the injury site by promoting expression of regeneration-associated genes. We have previously shown that peripheral nerve injury increases the number of macrophages in the DRGs and that the activated macrophages are critical to the enhancement of intrinsic regeneration capacity. The present study identifies a novel chemokine signal mediated by CCL2 that links regenerating neurons with proregenerative macrophage activation. Neutralization of CCL2 abolished the neurite outgrowth activity of conditioned medium obtained from neuron-macrophage cocultures treated with cAMP. The neuron-macrophage interactions that produced outgrowth-promoting conditioned medium required CCL2 in neurons and CCR2/CCR4 in macrophages. The conditioning effects were abolished in CCL2-deficient mice at 3 and 7 d after sciatic nerve injury, but CCL2 was dispensable for the initial growth response and upregulation of GAP-43 at the 1 d time point. Intraganglionic injection of CCL2 mimicked conditioning injury by mobilizing M2-like macrophages. Finally, overexpression of CCL2 in DRGs promoted sensory axon regeneration in a rat spinal cord injury model without harmful side effects. Our data suggest that CCL2-mediated neuron-macrophage interaction plays a critical role for amplification and maintenance of enhanced regenerative capacity by preconditioning peripheral nerve injury. Manipulation of chemokine signaling mediating neuron-macrophage interactions may represent a novel therapeutic approach to promote axon regeneration after CNS injury.

  11. Molecular and epigenetic basis of macrophage polarized activation.

    PubMed

    Porta, Chiara; Riboldi, Elena; Ippolito, Alessandro; Sica, Antonio

    2015-08-01

    Macrophages are unique cells for origin, heterogeneity and plasticity. At steady state most of macrophages are derived from fetal sources and maintained in adulthood through self-renewing. Despite sharing common progenitors, a remarkable heterogeneity characterized tissue-resident macrophages indicating that local signals educate them to express organ-specific functions. Macrophages are extremely plastic: chromatin landscape and transcriptional programs can be dynamically re-shaped in response to microenvironmental changes. Owing to their ductility, macrophages are crucial orchestrators of both initiation and resolution of immune responses and key supporters of tissue development and functions in homeostatic and pathological conditions. Herein, we describe current understanding of heterogeneity and plasticity of macrophages using the M1-M2 dichotomy as operationally useful simplification of polarized activation. We focused on the complex network of signaling cascades, metabolic pathways, transcription factors, and epigenetic changes that control macrophage activation. In particular, this network was addressed in sepsis, as a paradigm of a pathological condition determining dynamic macrophage reprogramming.

  12. Combined Administration of ASCs and BMP-12 Promotes an M2 Macrophage Phenotype and Enhances Tendon Healing.

    PubMed

    Gelberman, Richard H; Linderman, Stephen W; Jayaram, Rohith; Dikina, Anna D; Sakiyama-Elbert, Shelly; Alsberg, Eben; Thomopoulos, Stavros; Shen, Hua

    2017-09-01

    formation. Quantitative assessment of gene and protein expression showed effects of treatment: ASC-sheet treatment modulated the postrepair inflammatory response and facilitated healing by increasing regenerative M2 macrophages (M2 marker CD204, twofold of normal, p = 0.030), inflammatory inhibitor (prostaglandin reductase 1 [PTRG1], 1.6-fold of normal, p = 0.026), and proteins involved in tendon formation (periostin [POSTN], 1.9-fold of normal, p = 0.035). Consistently, semiquantitative and qualitative evaluations of repaired tissue showed that ASC-sheet treatment reduced mononuclear cell infiltration (12% less than nontreated tendons, p = 0.021) and introduced CD146+ stem or progenitor cells to the repair site. The combined administration of ASCs and rBMP-12 further stimulated M2 macrophages by increasing IL-4 (116-fold of normal, p = 0.002) and led to the increase of M2 effector matrix metalloproteinase-12 involved in matrix remodeling (twofold of normal, p = 0.016) and reduction of a negative regulator of angiogenesis and cell migration (StAR-related lipid transfer domain protein13 [STARD13]; 84% of normal, p = 0.000), thus facilitating the proliferative stage of tendon repair. ASCs and BMP-12 accelerated the progression of healing in the proliferative stage of tendon repair. The effects of ASCs and BMP-12 on tendon functional recovery should be evaluated in future studies. The cell sheet approach is an effective, biocompatible, and surgeon-friendly approach for cell and growth factor delivery during tendon repair. Combined application of ASCs and BMP-12 may accelerate intrasynovial tendon healing while suppressing the adverse inflammatory response.

  13. Effect of modulation of PPAR-γ activity on Kupffer cells M1/M2 polarization in the development of non-alcoholic fatty liver disease

    PubMed Central

    Luo, Wenjing; Xu, Qinyu; Wang, Qi; Wu, Huimin; Hua, Jing

    2017-01-01

    Abnormal lipid-mediated hepatic inflammatory-immune dysfunction and chronic low grade inflammation play an important role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Macrophage polarization is an important mechanism for the regulation of inflammatory response. Since PPAR-γ has emerged as a master regulator of macrophage polarization, we aimed to investigate the lipid-induced macrophage/Kupffer cell polarization in vivo and in vitro, and explore the association between PPAR-γ activity and macrophages M1/M2 polarization shifting. Here we showed that long-term high-fat diet increased Kupffer cells content with M1-predominant phenotype and increasing production of pro-inflammatory cytokines. Saturated fatty acids polarized Kupffer cells/macrophages to an M1-predominant phenotype while n-3 PUFA polarized Kupffer cells/macrophages to an M2 phenotype, which was associated with activation of NF-κB signal pathway and PPAR-γ respectively. Furthermore, up-regulation of PPAR-γ shifted lipid-induced macrophages polarization from M1-predominant phenotype to M2 phenotype. Macrophages polarization switch was associated with the interaction between PPAR-γ and NF-κBp65 signal pathway. Rosiglitazone restored high-fat diet-induced imblance of Kupffer cells M1/M2 polarization and alleviated hepatic steatosis as well as local pro-inflammatory response. These findings suggest that manipulation of PPAR-γ activity has the potential to balance lipid-induced M1/M2 macrophage/Kupffer cell polarization, and leading to prevent the development of NAFLD. PMID:28300213

  14. Effect of modulation of PPAR-γ activity on Kupffer cells M1/M2 polarization in the development of non-alcoholic fatty liver disease.

    PubMed

    Luo, Wenjing; Xu, Qinyu; Wang, Qi; Wu, Huimin; Hua, Jing

    2017-03-16

    Abnormal lipid-mediated hepatic inflammatory-immune dysfunction and chronic low grade inflammation play an important role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). Macrophage polarization is an important mechanism for the regulation of inflammatory response. Since PPAR-γ has emerged as a master regulator of macrophage polarization, we aimed to investigate the lipid-induced macrophage/Kupffer cell polarization in vivo and in vitro, and explore the association between PPAR-γ activity and macrophages M1/M2 polarization shifting. Here we showed that long-term high-fat diet increased Kupffer cells content with M1-predominant phenotype and increasing production of pro-inflammatory cytokines. Saturated fatty acids polarized Kupffer cells/macrophages to an M1-predominant phenotype while n-3 PUFA polarized Kupffer cells/macrophages to an M2 phenotype, which was associated with activation of NF-κB signal pathway and PPAR-γ respectively. Furthermore, up-regulation of PPAR-γ shifted lipid-induced macrophages polarization from M1-predominant phenotype to M2 phenotype. Macrophages polarization switch was associated with the interaction between PPAR-γ and NF-κBp65 signal pathway. Rosiglitazone restored high-fat diet-induced imblance of Kupffer cells M1/M2 polarization and alleviated hepatic steatosis as well as local pro-inflammatory response. These findings suggest that manipulation of PPAR-γ activity has the potential to balance lipid-induced M1/M2 macrophage/Kupffer cell polarization, and leading to prevent the development of NAFLD.

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

    PubMed Central

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

    2016-01-01

    In ovarian cancer, a high ratio of anti-inflammatory M2 to pro-inflammatory M1 macrophages correlates with poor patient prognosis. The mechanisms driving poor tumor outcome as a result of the presence of M2 macrophages in the tumor microenvironment remain unclear and are challenging to study with current techniques. Therefore, in this study we utilized a micro-culture device previously developed by our lab to model concentrated paracrine signaling in order to address our hypothesis that interactions between M2 macrophages and ovarian cancer cells induce tumor cell proliferation. Using the micro-culture device, we determined that co-culture with M2-differentiated primary macrophages or THP-1 increased OVCA433 proliferation by 10–12%. This effect was eliminated with epidermal growth factor receptor (EGFR) or heparin-bound epidermal growth factor (HB-EGF) neutralizing antibodies and HBEGF expression in peripheral blood mononuclear cells from ovarian cancer patients was 9-fold higher than healthy individuals, suggesting a role for HB-EGF in tumor progression. However, addition of HB-EGF at levels secreted by macrophages or macrophage-conditioned media did not induce proliferation to the same extent, indicating a role for other factors in this process. Matrix metalloproteinase-9, MMP-9, which cleaves membrane-bound HB-EGF, was elevated in co-culture and its inhibition decreased proliferation. Utilizing inhibitors and siRNA against MMP9 in each population, we determined that macrophage-secreted MMP-9 released HB-EGF from macrophages, which increased MMP9 in OVCA433, resulting in a positive feedback loop to drive HB-EGF release and increase proliferation in co-culture. Identification of multi-cellular interactions such as this may provide insight into how to most effectively control ovarian cancer progression. PMID:27888810

  16. A possible mechanism in the recruitment of eosinophils and Th2 cells through CD163(+) M2 macrophages in the lesional skin of eosinophilic cellulitis.

    PubMed

    Fujimura, Taku; Kambayashi, Yumi; Furudate, Sadanori; Kakizaki, Aya; Aiba, Setsuya

    2014-01-01

    M2 macrophages play a critical role in the recruitment of T helper 2 (Th2) regulatory T cells (Treg). To study the role of M2 macrophages and Treg cells in eosinophilic celulitis. We employed immunohistochemical staining for CD163( )and CD206 (macrophages) as well as FoxP3 (Treg), in lesional skin of four cases of eosinophilic cellulitis. CD163(+) CD206(+) M2 macrophages, which were previously reported to produce CCL17 to induce Th2 cells and Treg cells, were predominantly infiltrating the subcutaneous tissues and interstitial area of the dermis. M2 macrophages derived from PBMC showed significantly increased expression of CCL11, CCL17, CCL24 and CCL26 mRNA and production of CCL17 and CCL24, when stimulated by IL-4 or IL- 13. In addition, CCL17-producing cells and CCL24-producing cells were prominent in the lesional skin of EC. Our study sheds light on one of the possible immunological mechanisms of eosinophilic cellulitis.

  17. Alternative (M2) activation of Kupffer cells by PPARδ ameliorates obesity-induced insulin resistance

    PubMed Central

    Odegaard, Justin I.; Ricardo-Gonzalez, Roberto R.; Eagle, Alex Red; Vats, Divya; Morel, Christine R.; Goforth, Matthew H.; Subramanian, Vidya; Mukundan, Lata; Ferrante, Anthony W.; Chawla, Ajay

    2008-01-01

    SUMMARY Macrophage infiltration and activation in metabolic tissues underlie obesity-induced insulin resistance and type 2 diabetes. While inflammatory activation of resident hepatic macrophages potentiates insulin resistance, the functions of alternatively activated Kupffer cells in metabolic disease remain unknown. Here we show that, in response to the Th2 cytokine interleukin-4 (IL-4), peroxisome proliferator activated receptor δ (PPARδ) directs expression of the alternative phenotype in Kupffer cells and adipose tissue macrophages of lean mice. However, adoptive transfer of PPARδ null bone marrow into wild type mice only diminishes alternative activation of hepatic macrophages, causing hepatic dysfunction and systemic insulin resistance. Suppression of hepatic oxidative metabolism is recapitulated by treatment of primary hepatocytes with conditioned media from PPARδ null macrophages, indicating direct involvement of Kupffer cells in liver lipid metabolism. Taken together, these data suggest an unexpected beneficial role for alternatively activated Kupffer cells in metabolic syndrome and type 2 diabetes. PMID:18522831

  18. α-ketoglutarate orchestrates macrophage activation through metabolic and epigenetic reprogramming.

    PubMed

    Liu, Pu-Ste; Wang, Haiping; Li, Xiaoyun; Chao, Tung; Teav, Tony; Christen, Stefan; Di Conza, Giusy; Cheng, Wan-Chen; Chou, Chih-Hung; Vavakova, Magdalena; Muret, Charlotte; Debackere, Koen; Mazzone, Massimiliano; Huang, Hsien-Da; Fendt, Sarah-Maria; Ivanisevic, Julijana; Ho, Ping-Chih

    2017-09-01

    Glutamine metabolism provides synergistic support for macrophage activation and elicitation of desirable immune responses; however, the underlying mechanisms regulated by glutamine metabolism to orchestrate macrophage activation remain unclear. Here we show that the production of α-ketoglutarate (αKG) via glutaminolysis is important for alternative (M2) activation of macrophages, including engagement of fatty acid oxidation (FAO) and Jmjd3-dependent epigenetic reprogramming of M2 genes. This M2-promoting mechanism is further modulated by a high αKG/succinate ratio, whereas a low ratio strengthens the proinflammatory phenotype in classically activated (M1) macrophages. As such, αKG contributes to endotoxin tolerance after M1 activation. This study reveals new mechanistic regulations by which glutamine metabolism tailors the immune responses of macrophages through metabolic and epigenetic reprogramming.

  19. Pancreatic cancer cell/fibroblast co-culture induces M2 like macrophages that influence therapeutic response in a 3D model.

    PubMed

    Kuen, Janina; Darowski, Diana; Kluge, Tobias; Majety, Meher

    2017-01-01

    Pancreatic cancer (PC) remains one of the most challenging solid tumors to treat with a high unmet medical need as patients poorly respond to standard-of-care-therapies. Prominent desmoplastic reaction involving cancer-associated fibroblasts (CAFs) and the immune cells in the tumor microenvironment (TME) and their cross-talk play a significant role in tumor immune escape and progression. To identify the key cellular mechanisms induce an immunosuppressive tumor microenvironment, we established 3D co-culture model with pancreatic cancer cells, CAFs and monocytes. Using this model, we analyzed the influence of tumor cells and fibroblasts on monocytes and their immune suppressive phenotype. Phenotypic characterization of the monocytes after 3D co-culture with tumor/fibroblast spheroids was performed by analyzing the expression of defined cell surface markers and soluble factors. Functionality of these monocytes and their ability to influence T cell phenotype and proliferation was investigated. 3D co-culture of monocytes with pancreatic cancer cells and fibroblasts induced the production of immunosuppressive cytokines which are known to promote polarization of M2 like macrophages and myeloid derived suppressive cells (MDSCs). These co-culture spheroid polarized monocyte derived macrophages (MDMs) were poorly differentiated and had an M2 phenotype. The immunosuppressive function of these co-culture spheroids polarized MDMs was demonstrated by their ability to inhibit CD4+ and CD8+ T cell activation and proliferation in vitro, which we could partially reverse by 3D co-culture spheroid treatment with therapeutic molecules that are able to re-activated spheroid polarized MDMs or block immune suppressive factors such as Arginase-I.

  20. Polarization of Tissue-Resident TFH-Like Cells in Human Hepatoma Bridges Innate Monocyte Inflammation and M2b Macrophage Polarization.

    PubMed

    Chen, Min-Min; Xiao, Xiao; Lao, Xiang-Ming; Wei, Yuan; Liu, Rui-Xian; Zeng, Qiu-Hui; Wang, Jun-Cheng; Ouyang, Fang-Zhu; Chen, Dong-Ping; Chan, Ka-Wo; Shi, Dai-Chao; Zheng, Limin; Kuang, Dong-Ming

    2016-10-01

    The existence, regulation, and functions of IL21(+) immune cells are poorly defined in human cancers. Here, we identified a subset of protumorigenic IL21(+) TFH-like cells in human hepatocellular carcinoma. These cells were the major source of IL21 in tumors and represented about 10% of the CD4(+) T-cell population at levels comparable with the TFH cells present in lymph nodes. However, these TFH-like cells displayed a unique CXCR5(-)PD-1(lo/-)BTLA(-)CD69(hi) tissue-resident phenotype with substantial IFNγ production, which differed from the phenotype of TFH cells. Toll-like receptor 4 (TLR4)-elicited innate monocyte inflammation was important for IL21(+) TFH-like cell induction in tumors, and activation of STAT1 and STAT3 was critical for TFH-like cell polarization in this process. Importantly, the TFH-like cells operated in IL21-IFNγ-dependent pathways to induce plasma cell differentiation and thereby create conditions for protumorigenic M2b macrophage polarization and cancer progression. Thus, induction of TFH-like cells links innate inflammation to immune privilege in tumors. We identified a novel protumorigenic IL21(+) TFH-like cell subset with a CXCR5(-)PD-1(-) BTLA(-)CD69(hi) tissue-resident phenotype in hepatoma. TLR4-mediated monocyte inflammation and subsequent T-cell STAT1 and STAT3 activation are critical for TFH-like cell induction. TFH-like cells operate via IL21-IFNγ pathways to induce plasma cells and create conditions for M2b macrophage polarization. Cancer Discov; 6(10); 1182-95. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 1069. 2016 American Association for Cancer Research.

  1. M1-/M2-macrophages contribute to the development of GST-P-positive preneoplastic lesions in chemically-induced rat cirrhosis.

    PubMed

    Wijesundera, Kavindra Kumara; Izawa, Takeshi; Tennakoon, Anusha Hemamali; Golbar, Hossain M; Tanaka, Miyuu; Kuwamura, Mitsuru; Yamate, Jyoji

    2015-09-01

    Glutathione S-transferase placental form (GST-P) expression in hepatocyte foci is regarded as a preneoplastic change in rats. We aimed to reveal the contribution of polarized macrophages in development of GST-P-positive pseudolobules (PLs) in chemically-induced rat cirrhosis. F344 rats were injected with thioacetamide (100mg/kg BW, twice a week, intraperitoneally). Macrophage immunophenotypes and expression of M1-/M2-related factors were analyzed by immunohistochemistry, real-time RT-PCR and laser microdissection. GST-P-positive foci/clusters were clearly observed at post-first injection week 15. GST-P-positive PLs were distinguishable at weeks 20-32. Microarray analysis revealed upregulation of preneoplastic genes in GST-P-positive PLs at week 32. M1 (CD68(+), Iba1(+))-and M2 (CD163(+), CD204(+), Gal-3(+))-macrophages were greater in number in the GST-P-positive PLs, whereas MHC class II-positive (M1) macrophage number was fewer in the GST-P-positive PLs. Expression of both M1 (IFN-γ, IL-1β, TNF-α, Iba1)- and M2 (IL-4, TGF-β1, IL-10)-related factors were higher in GST-P-positive PLs. Our results showed that both M1- and M2-macrophage populations contribute to the development of hepatic preneoplastic lesions. MHC class II-positive macrophages may be related to anti-tumor progression, since their kinetics showed reverse pattern to other macrophage phenotypes. Copyright © 2015 Elsevier GmbH. All rights reserved.

  2. Conditional deletion of caspase-8 in macrophages alters macrophage activation in a RIPK-dependent manner.

    PubMed

    Cuda, Carla M; Misharin, Alexander V; Khare, Sonal; Saber, Rana; Tsai, FuNien; Archer, Amy M; Homan, Philip J; Haines, G Kenneth; Hutcheson, Jack; Dorfleutner, Andrea; Budinger, G R Scott; Stehlik, Christian; Perlman, Harris

    2015-10-16

    Although caspase-8 is a well-established initiator of apoptosis and suppressor of necroptosis, recent evidence suggests that this enzyme maintains functions beyond its role in cell death. As cells of the innate immune system, and in particular macrophages, are now at the forefront of autoimmune disease pathogenesis, we examined the potential involvement of caspase-8 within this population. Cre (LysM) Casp8 (fl/fl) mice were bred via a cross between Casp8 (fl/fl) mice and Cre (LysM) mice, and RIPK3 (-/-) Cre (LysM) Casp8 (fl/fl) mice were generated to assess the contribution of receptor-interacting serine-threonine kinase (RIPK)3. Immunohistochemical and immunofluorescence analyses were used to examine renal damage. Flow cytometric analysis was employed to characterize splenocyte distribution and activation. Cre (LysM) Casp8 (fl/fl) mice were treated with either Toll-like receptor (TLR) agonists or oral antibiotics to assess their response to TLR activation or TLR agonist removal. Luminex-based assays and enzyme-linked immunosorbent assays were used to measure cytokine/chemokine and immunoglobulin levels in serum and cytokine levels in cell culture studies. In vitro cell culture was used to assess macrophage response to cell death stimuli, TLR activation, and M1/M2 polarization. Data were compared using the Mann-Whitney U test. Loss of caspase-8 expression in macrophages promotes onset of a mild systemic inflammatory disease, which is preventable by the deletion of RIPK3. In vitro cell culture studies reveal that caspase-8-deficient macrophages are prone to a caspase-independent death in response to death receptor ligation; yet, caspase-8-deficient macrophages are not predisposed to unchecked survival, as analysis of mixed bone marrow chimeric mice demonstrates that caspase-8 deficiency does not confer preferential expansion of myeloid populations. Loss of caspase-8 in macrophages dictates the response to TLR activation, as injection of TLR ligands upregulates

  3. Expression of decoy receptor 3 in diffuse sclerosing variant of papillary thyroid carcinoma: correlation with M2 macrophage differentiation and lymphatic invasion.

    PubMed

    Chang, Wei-Chin; Chen, Jui-Yu; Lee, Chen-Hsen; Yang, An-Hang

    2013-06-01

    The diffuse sclerosing variant of papillary thyroid carcinoma (DSV-PTC) is a unique variant of PTC that is characterized by extensive lymphovascular invasion of tumor cells in a background of lymphocytic thyroiditis. The lymphatic emboli contain tumor cells as well as macrophages, but the recruitment of these macrophages is not well understood. The aim of this study was to determine the relationship between the expression of Decoy receptor 3 (DcR3), the recruitment of tumor-associated macrophages (TAMs), and lymphatic invasion in DSV-PTC. We retrospectively examined 14 cases of DSV-PTC using immunohistochemistry studies. The density of TAMs, lymphatic vessel density, lymphatic invasion, tumor emboli area, and DcR3 expression were assessed. Statistical analyses were performed using Fisher's exact test, unpaired t-test, and linear regression. The lymphatic tumor emboli contained a relatively higher density of TAMs than stroma and classical PTC (CPTC) areas. In addition, the number of lymphatic invasions and the size of the tumor emboli area were positively correlated with the number of M2 TAMs. A higher density of M2 TAMs was associated with older patients and larger tumor size. Moreover, DcR3 was expressed only in lymphatic tumor cells and squamous metaplastic tumor cells, but not in macrophages and CPTC. In addition, the preferential expression of DcR3 in tumors was associated with higher levels of M2 TAMs and lymphatic invasion. Despite the fact that the exact relationship between DcR3, M2 macrophages, and lymphatic invasion in DSV-PTC remains to be elucidated, our findings suggest that DcR3 expression in DSV-PTC tumor cells may promote the polarized macrophage differentiation toward the M2 phenotype. This phenomenon may further promote lymphatic invasion of DSV-PTC tumor cells.

  4. Neem leaf glycoprotein regulates function of tumor associated M2 macrophages in hypoxic tumor core: Critical role of IL-10/STAT3 signaling.

    PubMed

    Goswami, Kuntal Kanti; Sarkar, Madhurima; Ghosh, Sarbari; Saha, Akata; Ghosh, Tithi; Guha, Ipsita; Barik, Subhasis; Banerjee, Saptak; Roy, Soumyabrata; Bose, Anamika; Dasgupta, Parthasarathi; Baral, Rathindranath

    2016-12-01

    Heterogeneous tumor microenvironment (TME), broadly divided into tumor core and peripheral sub-microenvironments, differentially polarize normal macrophages into a different form known as tumor associated M2 macrophages (M2TAMs) to promote tumor growth. In view of the extensive immune-editing role of NLGP, here, we have observed that NLGP is effective to convert M2TAMs (CD11b(+)F4/80(high)) to M1 (CD11b(+)F4/80(low)) more prominently in tumor core, along with downregulation of other M2 associated markers, like, ManR, Ym1, Fizz1. High IL-10:IL-12 ratio at tumor core was downregulated in NLGP treated melanoma bearing mice. Decrease in IL-10 by NLGP is again associated with the decrease in hypoxia, as indicated by prominent downregulation of HIF1α and VEGF, particularly at tumor core. Macrophages exposed to hypoxic tumor core lysates in vitro exhibited high IL-10, HIF1α and VEGF expression that was significantly downregulated by NLGP. Further evidences suggest M2TAM to M1 conversion by NLGP is associated with STAT3-regulated IL-10 dependent pathway without affecting the IL-4 dependent one. Such TAM modulatory functions of NLGP might help in the restriction of melanoma growth by increasing the proportion of M1 TAMs in tumor core that helps in prevention of tumor relapse and dissemination of the tumor mass.

  5. Incomplete deletion of IL-4Rα by LysM(Cre) reveals distinct subsets of M2 macrophages controlling inflammation and fibrosis in chronic schistosomiasis.

    PubMed

    Vannella, Kevin M; Barron, Luke; Borthwick, Lee A; Kindrachuk, Kristen N; Narasimhan, Prakash Babu; Hart, Kevin M; Thompson, Robert W; White, Sandra; Cheever, Allen W; Ramalingam, Thirumalai R; Wynn, Thomas A

    2014-09-01

    Mice expressing a Cre recombinase from the lysozyme M-encoding locus (Lyz2) have been widely used to dissect gene function in macrophages and neutrophils. Here, we show that while naïve resident tissue macrophages from IL-4Rαf(lox/delta)LysM(Cre) mice almost completely lose IL-4Rα function, a large fraction of macrophages elicited by sterile inflammatory stimuli, Schistosoma mansoni eggs, or S. mansoni infection, fail to excise Il4rα. These F4/80(hi)CD11b(hi) macrophages, in contrast to resident tissue macrophages, express lower levels of Lyz2 explaining why this population resists LysM(Cre)-mediated deletion. We show that in response to IL-4 and IL-13, Lyz2(lo)IL-4Rα(+) macrophages differentiate into an arginase 1-expressing alternatively-activated macrophage (AAM) population, which slows the development of lethal fibrosis in schistosomiasis. In contrast, we identified Lyz2(hi)IL-4Rα(+) macrophages as the key subset of AAMs mediating the downmodulation of granulomatous inflammation in chronic schistosomiasis. Our observations reveal a limitation on using a LysMCre mouse model to study gene function in inflammatory settings, but we utilize this limitation as a means to demonstrate that distinct populations of alternatively activated macrophages control inflammation and fibrosis in chronic schistosomiasis.

  6. Critical role of p38 MAPK in IL-4-induced alternative activation of peritoneal macrophages.

    PubMed

    Jiménez-Garcia, Lidia; Herránz, Sandra; Luque, Alfonso; Hortelano, Sonsoles

    2015-01-01

    Alternative activation of macrophages plays an important role in a range of physiological and pathological processes. This alternative phenotype, also known as M2 macrophages, is induced by type 2 cytokines such as IL-4. The binding of IL-4 to its receptor leads to activation of two major signaling pathways: STAT-6 and PI3K. However, recent studies have described that p38 MAPK might play a role in IL-4-dependent signaling in some cells, although its role in macrophages is still controversial. In this study, we investigated whether p38 MAPK plays a role in the polarization of macrophages in mice. Our results reveal that IL-4 induces phosphorylation of p38 MAPK in thioglycollate-elicited murine peritoneal macrophages, in addition to STAT-6 and PI3K activation. Furthermore, p38 MAPK inactivation, by gene silencing or pharmacological inhibition, suppressed IL-4-induced typical M2 markers, indicating the involvement of p38 MAPK in the signaling of IL-4 leading to M2-macrophage polarization. Moreover, p38 MAPK inhibition blocked phosphorylation of STAT-6 and Akt, suggesting that p38 MAPK is upstream of these signaling pathways. Finally, we show that in an in vivo model of chitin-induced M2 polarization, p38 MAPK inhibition also diminished activation of M2 markers. Taken together, our data establish a new role for p38 MAPK during IL-4-induced alternative activation of macrophages. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Pulmonary macrophages activity in CO intoxication.

    PubMed

    Pieri, Maria; Giugliano, Pasquale; Vacchiano, Giuseppe

    2016-02-01

    The presence of macrophages and their activation on the pulmonary tissues of 21 subjects deceased after CO intoxication has been studied. A notable number of activated macrophages, especially in the interstitial level, have been evidenced, and such phenomenon supports the hypothesis of a possible association between CO intoxication and pulmonary macrophages activity. The highlighted association could be mediated by changes of the surfactant, by impairing of mitochondrial respiration and by release of pro-inflammatory cytokines. Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.

  8. Macrophage activation in human diseases.

    PubMed

    Schultze, Joachim L; Schmieder, Astrid; Goerdt, S

    2015-08-01

    It is becoming increasingly accepted that macrophages play a crucial role in many diseases associated with chronic inflammation, including atherosclerosis, obesity, diabetes, cancer, skin diseases, and even neurodegenerative diseases. It is therefore not surprising that macrophages in human diseases have gained significant interest during the last years. Molecular analysis combined with more sophisticated murine disease models and the application of genome-wide technologies has resulted in a much better understanding of the role of macrophages in human disease. We highlight important gain of knowledge during the last years for tumor-associated macrophages, and for macrophages in atherosclerosis, obesity and wound healing. Albeit these exciting findings certainly pave the way to novel diagnostics and therapeutics, several hurdles still need to be overcome. We propose a general outline for future research and development in disease-related macrophage biology based on integrating (1) genome-wide technologies, (2) direct human sampling, and (3) a dedicated use of in vivo model systems. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Tim-3 promotes tumor-promoting M2 macrophage polarization by binding to STAT1 and suppressing the STAT1-miR-155 signaling axis.

    PubMed

    Jiang, Xingwei; Zhou, Tingting; Xiao, Yan; Yu, Jiahui; Dou, Shuaijie; Chen, Guojiang; Wang, Renxi; Xiao, He; Hou, Chunmei; Wang, Wei; Shi, Qingzhu; Feng, Jiannan; Ma, Yuanfang; Shen, Beifen; Li, Yan; Han, Gencheng

    2016-01-01

    T cell Ig mucin-3 (Tim-3), an immune checkpoint inhibitor, shows therapeutic potential. However, the molecular mechanism by which Tim-3 regulates immune responses remains to be determined. In particular, very little is known about how Tim-3 works in innate immune cells. Here, we demonstrated that Tim-3 is involved in the development of tumor-promoting M2 macrophages in colon cancer. Manipulation of the Tim-3 pathway significantly affected the polarization status of intestinal macrophages and the progression of colon cancer. The Tim-3 signaling pathway in macrophages was explored using microarray, co-immunoprecipitation, gene mutation, and high-content analysis. For the first time, we demonstrated that Tim-3 polarizes macrophages by directly binding to STAT1 via residue Y256 and Y263 in its intracellular tail and inhibiting the STAT1-miR-155-SOCS1 signaling axis. We also identified a new signaling adaptor of Tim-3 in macrophages, and, by modulating the Tim-3 pathway, demonstrated the feasibility of altering macrophage polarization as a potential tool for treating this kind of disease.

  10. IL-10 Cytokine Released from M2 Macrophages Is Crucial for Analgesic and Anti-inflammatory Effects of Acupuncture in a Model of Inflammatory Muscle Pain

    PubMed Central

    da Silva, Morgana D.; Bobinski, Franciane; Sato, Karina L.; Kolker, Sandra J.; Sluka, Kathleen A.; Santos, Adair R. S.

    2014-01-01

    Muscle pain is a common medical problem that is difficult to treat. One nonpharmacological treatment used is acupuncture, a procedure in which fine needles are inserted into body points with the intent of relieving pain and other symptoms. Here we investigated the effects of manual acu-puncture (MA) on modulating macrophage phenotype and interleukin-10 (IL-10) concentrations in animals with muscle inflammation. Carrageenan, injected in the gastrocnemius muscle of mice, induces an inflammatory response characterized by mechanical hyperalgesia and edema. The inflammation is initially a neutrophilic infiltration that converts to a macrophage-dominated inflammation by 48 h. MA of the Sanyinjiao or Spleen 6 (SP6) acupoint reduces nociceptive behaviors, heat, and mechanical hyperalgesia and enhanced escape/avoidance and the accompanying edema. SP6 MA increased muscle IL-10 levels and was ineffective in reducing pain behaviors and edema in IL-10 knockout (IL-10−/−) mice. Repeated daily treatments with SP6 MA induced a phenotypic switch of muscle macrophages with reduced M1 macrophages (pro-inflammatory cells) and an increase of M2 macrophages (anti-inflammatory cells and important IL-10 source). These findings provide new evidence that MA produces a phenotypic switch in macrophages and increases IL-10 concentrations in muscle to reduce pain and inflammation. PMID:24961568

  11. Endogenous Epoxygenases Are Modulators of Monocyte/Macrophage Activity

    PubMed Central

    Sugden, Mary C.; Holness, Mark J.; Swales, Karen E.; Warner, Timothy D.; Edin, Matthew L.; Zeldin, Darryl C.; Gilroy, Derek W.; Bishop-Bailey, David

    2011-01-01

    Background Arachidonic acid is metabolized through three major metabolic pathways, the cyclooxygenase, lipoxygenase and CYP450 enzyme systems. Unlike cyclooxygenase and lipoxygenases, the role of CYP450 epoxygenases in monocyte/macrophage-mediated responses is not known. Methodology/Principal Findings When transfected in vitro, CYP2J2 is an efficient activator of anti-inflammatory pathways through the nuclear receptor peroxisome proliferator-activated receptor (PPAR) α. Human monocytes and macrophages contain PPARα and here we show they express the epoxygenases CYP2J2 and CYP2C8. Inhibition of constitutive monocyte epoxygenases using the epoxygenase inhibitor SKF525A induces cyclooxygenase (COX)-2 expression and activity, and the release of TNFα, and can be reversed by either add back of the endogenous epoxygenase products and PPARα ligand 11,12- epoxyeicosatrienoic acid (EET) or the addition of the selective synthetic PPARα ligand GW7647. In alternatively activated (IL-4-treated) monocytes, in contrast to classically activated cells, epoxygenase inhibition decreased TNFα release. Epoxygenases can be pro-inflammatory via superoxide anion production. The suppression of TNFα by SKF525A in the presence of IL-4 was associated with a reduction in superoxide anion generation and reproduced by the superoxide dismutase MnCl2. Similar to these acute activation studies, in monocyte derived macrophages, epoxygenase inhibition elevates M1 macrophage TNFα mRNA and further decreases M2 macrophage TNFα. Conclusions/Significance In conclusion, epoxygenase activity represents an important endogenous pathway which limits monocyte activation. Moreover endogenous epoxygenases are immuno-modulators regulating monocyte/macrophage activation depending on the underlying activation state. PMID:22028915

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

    SciTech Connect

    Hasegawa-Moriyama, Maiko; Ohnou, Tetsuya; Godai, Kohei; Kurimoto, Tae; Nakama, Mayo; Kanmura, Yuichi

    2012-09-14

    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 functions 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

  13. BTB and CNC homolog 1 (Bach1) deficiency ameliorates TNBS colitis in mice: role of M2 macrophages and heme oxygenase-1.

    PubMed

    Harusato, Akihito; Naito, Yuji; Takagi, Tomohisa; Uchiyama, Kazuhiko; Mizushima, Katsura; Hirai, Yasuko; Higashimura, Yasuki; Katada, Kazuhiro; Handa, Osamu; Ishikawa, Takeshi; Yagi, Nobuaki; Kokura, Satoshi; Ichikawa, Hiroshi; Muto, Akihiko; Igarashi, Kazuhiko; Yoshikawa, Toshikazu

    2013-01-01

    BTB and CNC homolog 1 (Bach1) is a transcriptional repressor of heme oxygenase-1 (HO-1), which plays an important role in the protection of cells and tissues against acute and chronic inflammation. However, the role of Bach1 in the gastrointestinal mucosal defense system remains little understood. HO-1 supports the suppression of experimental colitis and localizes mainly in macrophages in colonic mucosa. This study was undertaken to elucidate the Bach1/HO-1 system's effects on the pathogenesis of experimental colitis. This study used C57BL/6 (wild-type) and homozygous Bach1-deficient C57BL/6 mice in which colonic damage was induced by the administration of an enema of 2,4,6-trinitrobenzene sulfonic acid (TNBS). Subsequently, they were evaluated macroscopically, histologically, and biochemically. Peritoneal macrophages from the respective mice were isolated and analyzed. Then, wild-type mice were injected with peritoneal macrophages from the respective mice. Acute colitis was induced similarly. TNBS-induced colitis was inhibited in Bach1-deficient mice. TNBS administration increased the expression of HO-1 messenger RNA and protein in colonic mucosa in Bach1-deficient mice. The expression of HO-1 mainly localized in F4/80-immunopositive and CD11b-immunopositive macrophages. Isolated peritoneal macrophages from Bach1-deficient mice highly expressed HO-1 and also manifested M2 macrophage markers, such as Arginase-1, Fizz-1, Ym1, and MRC1. Furthermore, TNBS-induced colitis was inhibited by the transfer of Bach1-deficient macrophages into wild-type mice. Deficiency of Bach1 ameliorated TNBS-induced colitis. Bach1-deficient macrophages played a key role in protection against colitis. Targeting of this mechanism is applicable to cell therapy for human inflammatory bowel disease.

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

    PubMed

    López-García, Sonia; Castañeda-Sanchez, Jorge Ismael; Jiménez-Arellanes, Adelina; Domínguez-López, Lilia; Castro-Mussot, Maria Eugenia; Hernández-Sanchéz, Javier; Luna-Herrera, Julieta

    2015-08-06

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

  15. Cardiac fibroblast cytokine profiles induced by proinflammatory or profibrotic stimuli promote monocyte recruitment and modulate macrophage M1/M2 balance in vitro.

    PubMed

    Humeres, Claudio; Vivar, Raúl; Boza, Pia; Muñoz, Claudia; Bolivar, Samir; Anfossi, Renatto; Osorio, Jose Miguel; Olivares-Silva, Francisco; García, Lorena; Díaz-Araya, Guillermo

    2016-10-27

    Macrophage polarization plays an essential role in cardiac remodeling after injury, evolving from an initial accumulation of proinflammatory M1 macrophages to a greater balance of anti-inflammatory M2 macrophages. Whether cardiac fibroblasts themselves influence this process remains an intriguing question. In this work, we present evidence for a role of cardiac fibroblasts (CF) as regulators of macrophage recruitment and skewing. Adult rat CF, were treated with lipopolysaccharide (LPS) or TGF-β1, to evaluate ICAM-1 and VCAM-1 expression using Western blot and proinflammatory/profibrotic cytokine secretion using LUMINEX. We performed in vitro migration and adhesion assays of rat spleen monocytes to layers of TGF-β1- or LPS-pretreated CF. Finally, TGF-β1- or LPS-pretreated CF were co-cultured with monocyte, to evaluate their effects on macrophage polarization, using flow cytometry and cytokine secretion. There was a significant increase in monocyte adhesion to LPS- or TGF-β1-stimulated CF, associated with increased CF expression of ICAM-1 and VCAM-1. siRNA silencing of either ICAM-1 or VCAM-1 inhibited monocyte adhesion to LPS-pretreated CF; however, monocyte adhesion to TGF-β1-treated CF was dependent on only VCAM-1 expression. Pretreatment of CF with LPS or TGF-β1 increased monocyte migration to CF, and this effect was completely abolished with an MCP-1 antibody blockade. LPS-treated CF secreted elevated levels of TNF-α and MCP-1, and when co-cultured with monocyte, LPS-treated CF stimulated increased macrophage M1 polarization and secretion of proinflammatory cytokines (TNF-α, IL-12 and MCP-1). On the other hand, CF stimulated with TGF-β1 produced an anti-inflammatory cytokine profile (high IL-10 and IL-5, low TNF-α). When co-cultured with monocytes, the TGF-β1 stimulated fibroblasts skewed monocyte differentiation towards M2 macrophages accompanied by increased IL-10 and decreased IL-12 levels. Taken together, our results show for the first time that

  16. IL-6 promotes M2 macrophage polarization by modulating purinergic signaling and regulates the lethal release of nitric oxide during Trypanosoma cruzi infection.

    PubMed

    Sanmarco, Liliana M; Ponce, Nicolás E; Visconti, Laura M; Eberhardt, Natalia; Theumer, Martin G; Minguez, Ángel R; Aoki, Maria P

    2017-04-01

    The production of nitric oxide (NO) is a key defense mechanism against intracellular pathogens but it must be tightly controlled in order to avoid excessive detrimental oxidative stress. In this study we described a novel mechanism through which interleukin (IL)-6 mediates the regulation of NO release induced in response to Trypanosoma cruzi infection. Using a murine model of Chagas disease, we found that, in contrast to C57BL/6 wild type (WT) mice, IL-6-deficient (IL6KO) mice exhibited a dramatic increase in plasma NO levels concomitant with a significantly higher amount of circulating IL-1β and inflammatory monocytes. Studies on mouse macrophages and human monocytes, revealed that IL-6 decreased LPS-induced NO production but this effect was abrogated in the presence of anti-IL-1β and in macrophages deficient in the NLRP3 inflammasome. In accordance, while infected WT myocardium exhibited an early shift from microbicidal/M1 to anti-inflammatory/M2 macrophage phenotype, IL6KO cardiac tissue never displayed a dominant M2 macrophage profile that correlated with decreased expression of ATP metabolic machinery and a lower cardiac parasite burden. The deleterious effects of high NO production-induced oxidative stress were evidenced by enhanced cardiac malondialdehyde levels, myocardial cell death and mortality. The survival rate was improved by the treatment of IL-6-deficient mice with a NO production-specific inhibitor. Our data revealed that IL-6 regulates the excessive release of NO through IL-1β inhibition and determines the establishment of an M2 macrophage profile within infected heart tissue.

  17. A lovastatin-elicited genetic program inhibits M2 macrophage polarization and enhances T cell infiltration into spontaneous mouse mammary tumors.

    PubMed

    Mira, Emilia; Carmona-Rodríguez, Lorena; Tardáguila, Manuel; Azcoitia, Iñigo; González-Martín, Alicia; Almonacid, Luis; Casas, Josefina; Fabriás, Gemma; Mañes, Santos

    2013-12-01

    Beyond their ability to inhibit cholesterol biosynthesis, the statins have pleiotropic effects that include anti-inflammatory and immunomodulatory activities. Statins could have clinical utility, alone or in combination with other chemotherapeutics, in the treatment of cancer. The mechanisms that underlie the anti-tumor activity of the statins are nonetheless poorly defined. No studies have analyzed how they alter the tumor-associated leukocyte infiltrate, a central factor that influences tumor stroma and cancer evolution. Here we used HER2/neu transgenic (Tg-neu) mice to analyze the effect of lovastatin (Lov) on the inflammatory reaction of spontaneous mammary tumors. Lov treatment of tumor-bearing Tg-neu mice did not alter growth of established tumors, but significantly reduced the number of new oncogenic lesions in these mice. Moreover, Lov inhibited the growth of newly implanted Tg-neu tumors in immunocompetent but not in immunodeficient mice. We found that Lov enhanced tumor infiltration by effector T cells, and reduced the number of immunosuppressive and pro-angiogenic M2-like tumor-associated macrophages (TAM). Concomitantly, the drug improved the structure and function of the tumor vasculature, measured as enhanced tumor oxygenation and penetration of cytotoxic drugs. Microarray analysis identified a Lov-elicited genetic program in Tg-neu tumors that might explain these effects; we observed Lov-induced downregulation of placental growth factor, which triggers aberrant angiogenesis and M2-like TAM polarization. Our results identify a role for lovastatin in the shaping and re-education of the inflammatory infiltrate in tumors, with functional consequences in angiogenesis and antitumor immunity.

  18. EPA protects against muscle damage in the mdx mouse model of Duchenne muscular dystrophy by promoting a shift from the M1 to M2 macrophage phenotype.

    PubMed

    Carvalho, Samara Camaçari de; Apolinário, Leticia Montanholi; Matheus, Selma Maria Michelin; Santo Neto, Humberto; Marques, Maria Julia

    2013-11-15

    In dystrophic mdx mice and in Duchenne muscular dystrophy, inflammation contributes to myonecrosis. Previously, we demonstrated that eicosapentaenoic acid (EPA) decreased inflammation and necrosis in dystrophic muscle. In the present study, we examined the effects of EPA and the corticoid deflazacort (DFZ) as modulators of M1 (iNOS-expressing cells) and M2 (CD206-expressing cells) macrophages. Mdx mice (14 days old) received EPA or DFZ for 16 days. The diaphragm, biceps brachii and quadriceps muscles were studied. Immunofluorescence, immunoblotting and ELISA assays showed that EPA increased interleucin-10, reduced interferon-γ and was more effective than DFZ in promoting a shift from M1 to M2.

  19. Alternatively activated macrophages determine repair of the infarcted adult murine heart

    PubMed Central

    Shiraishi, Manabu; Shintani, Yasunori; Shintani, Yusuke; Ishida, Hidekazu; Saba, Rie; Yamaguchi, Atsushi; Adachi, Hideo; Yashiro, Kenta

    2016-01-01

    Alternatively activated (also known as M2) macrophages are involved in the repair of various types of organs. However, the contribution of M2 macrophages to cardiac repair after myocardial infarction (MI) remains to be fully characterized. Here, we identified CD206+F4/80+CD11b+ M2-like macrophages in the murine heart and demonstrated that this cell population predominantly increases in the infarct area and exhibits strengthened reparative abilities after MI. We evaluated mice lacking the kinase TRIB1 (Trib1–/–), which exhibit a selective depletion of M2 macrophages after MI. Compared with control animals, Trib1–/– mice had a catastrophic prognosis, with frequent cardiac rupture, as the result of markedly reduced collagen fibril formation in the infarct area due to impaired fibroblast activation. The decreased tissue repair observed in Trib1–/– mice was entirely rescued by an external supply of M2-like macrophages. Furthermore, IL-1α and osteopontin were suggested to be mediators of M2-like macrophage–induced fibroblast activation. In addition, IL-4 administration achieved a targeted increase in the number of M2-like macrophages and enhanced the post-MI prognosis of WT mice, corresponding with amplified fibroblast activation and formation of more supportive fibrous tissues in the infarcts. Together, these data demonstrate that M2-like macrophages critically determine the repair of infarcted adult murine heart by regulating fibroblast activation and suggest that IL-4 is a potential biological drug for treating MI. PMID:27140396

  20. Macrophage activation and its role in repair and pathology after spinal cord injury.

    PubMed

    Gensel, John C; Zhang, Bei

    2015-09-04

    The injured spinal cord does not heal properly. In contrast, tissue repair and functional recovery occur after skin or muscle injuries. The reason for this dichotomy in wound repair is unclear but inflammation, and specifically macrophage activation, likely plays a key role. Macrophages have the ability to promote the repair of injured tissue by regulating transitions through different phase of the healing response. In the current review we compare and contrast the healing and inflammatory responses between spinal cord injuries and tissues that undergo complete wound resolution. Through this comparison, we identify key macrophage phenotypes that are inaptly triggered or absent after spinal cord injury and discuss spinal cord stimuli that contribute to this maladaptive response. Sequential activation of classic, pro-inflammatory, M1 macrophages and alternatively activated, M2a, M2b, and M2c macrophages occurs during normal healing and facilitates transitions through the inflammatory, proliferative, and remodeling phases of repair. In contrast, in the injured spinal cord, pro-inflammatory macrophages potentiate a prolonged inflammatory phase and remodeling is not properly initiated. The desynchronized macrophage activation after spinal cord injury is reminiscent of the inflammation present in chronic, non-healing wounds. By refining the role macrophages play in spinal cord injury repair we bring to light important areas for future neuroinflammation and neurotrauma research. This article is part of a Special Issue entitled SI: Spinal cord injury.

  1. Statin attenuates experimental anti-glomerular basement membrane glomerulonephritis together with the augmentation of alternatively activated macrophages.

    PubMed

    Fujita, Emiko; Shimizu, Akira; Masuda, Yukinari; Kuwahara, Naomi; Arai, Takashi; Nagasaka, Shinya; Aki, Kaoru; Mii, Akiko; Natori, Yasuhiro; Iino, Yasuhiko; Katayama, Yasuo; Fukuda, Yuh

    2010-09-01

    Macrophages are heterogeneous and include classically activated M1 and alternatively activated M2 macrophages, characterized by pro- and anti-inflammatory functions, respectively. Macrophages that express heme oxygenase-1 also exhibit anti-inflammatory effects. We assessed the anti-inflammatory effects of statin in experimental anti-glomerular basement membrane glomerulonephritis and in vitro, focusing on the macrophage heterogeneity. Rats were induced anti-glomerular basement membrane glomerulonephritis and treated with atorvastatin (20 mg/kg/day) or vehicle (control). Control rats showed infiltration of macrophages in the glomeruli at day 3 and developed crescentic glomerulonephritis by day 7, together with increased mRNA levels of the M1 macrophage-associated cytokines, interferon-gamma, tumor necrosis factor-alpha, and interleukin-12. In contrast, statin reduced the level of proteinuria, reduced infiltration of macrophages in glomeruli with suppression of monocyte chemotactic protein-1 expression, and inhibited the formation of necrotizing and crescentic lesions. The number of glomerular ED3-positive macrophages decreased with down-regulation of M1 macrophage-associated cytokines. Furthermore, statin augmented ED2-positive M2 macrophages with up-regulation of the M2 macrophage-associated chemokines and cytokines, chemokine (C-C motif) Iigand-17 and interleukin-10. Statin also increased the glomerular interleukin-10-expressing heme oxygenase-1-positive macrophages. Statin inhibited macrophage development, and suppressed ED3-positive macrophages, but augmented ED2-positive macrophages in M2-associated cytokine environment in vitro. We conclude that the anti-inflammatory effects of statin in glomerulonephritis are mediated through inhibition of macrophage infiltration as well as augmentation of anti-inflammatory macrophages.

  2. High salt reduces the activation of IL-4– and IL-13–stimulated macrophages

    PubMed Central

    Binger, Katrina J.; Gebhardt, Matthias; Heinig, Matthias; Rintisch, Carola; Schroeder, Agnes; Neuhofer, Wolfgang; Hilgers, Karl; Manzel, Arndt; Schwartz, Christian; Kleinewietfeld, Markus; Voelkl, Jakob; Schatz, Valentin; Linker, Ralf A.; Lang, Florian; Voehringer, David; Wright, Mark D.; Hubner, Norbert; Dechend, Ralf; Jantsch, Jonathan; Titze, Jens; Müller, Dominik N.

    2015-01-01

    A high intake of dietary salt (NaCl) has been implicated in the development of hypertension, chronic inflammation, and autoimmune diseases. We have recently shown that salt has a proinflammatory effect and boosts the activation of Th17 cells and the activation of classical, LPS-induced macrophages (M1). Here, we examined how the activation of alternative (M2) macrophages is affected by salt. In stark contrast to Th17 cells and M1 macrophages, high salt blunted the alternative activation of BM-derived mouse macrophages stimulated with IL-4 and IL-13, M(IL-4+IL-13) macrophages. Salt-induced reduction of M(IL-4+IL-13) activation was not associated with increased polarization toward a proinflammatory M1 phenotype. In vitro, high salt decreased the ability of M(IL-4+IL-13) macrophages to suppress effector T cell proliferation. Moreover, mice fed a high salt diet exhibited reduced M2 activation following chitin injection and delayed wound healing compared with control animals. We further identified a high salt–induced reduction in glycolysis and mitochondrial metabolic output, coupled with blunted AKT and mTOR signaling, which indicates a mechanism by which NaCl inhibits full M2 macrophage activation. Collectively, this study provides evidence that high salt reduces noninflammatory innate immune cell activation and may thus lead to an overall imbalance in immune homeostasis. PMID:26485286

  3. Dysregulation of Macrophage Activation Profiles by Engineered Nanoparticles

    PubMed Central

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

    2013-01-01

    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 pre-treatment. Neither silica nor SPIO treatment elicited direct cytotoxic or pro-inflammatory effects in bone marrow-derived macrophages. However, pre-treatment of macrophages with SPIO caused extensive reprogramming of nearly 500 genes regulated in response to LPS challenge, hallmarked by exaggerated activation of oxidative stress response pathways and suppressed activation of both pro- and anti-inflammatory pathways. Silica pre-treatment 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 a 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. Nanotoxicology screening strategies

  4. Dysregulation of Macrophage Activation Profiles by Engineered Nanoparticles

    SciTech Connect

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

    2013-08-27

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

  5. Morphine Modulates Interleukin-4- or Breast Cancer Cell-induced Pro-metastatic Activation of Macrophages.

    PubMed

    Khabbazi, Samira; Goumon, Yannick; Parat, Marie-Odile

    2015-06-16

    Interactions between cancer cells and stromal cells in the tumour microenvironment play a key role in the control of invasiveness, metastasis and angiogenesis. Macrophages display a range of activation states in specific pathological contexts and alternatively activated (M2) macrophages can promote tumour aggressiveness. Opioids are able to modulate tumour growth and metastasis. We tested whether morphine modulates the activation of macrophages induced by (i) interleukin-4 (IL-4), the prototypical M2 polarization-inducing cytokine, or (ii) coculture with breast cancer cells. We showed that IL-4 causes increased MMP-9 production and expression of the alternative activation markers arginase-1 and MRC-1. Morphine prevented IL-4-induced increase in MMP-9 in a naloxone- and methylnaltrexone-reversible fashion. Morphine also prevented IL-4-elicited alternative activation of RAW264.7 macrophages. Expression of MMP-9 and arginase-1 were increased when RAW264.7 were subjected to paracrine activation by 4T1 cells, and this effect was prevented by morphine via an opioid receptor-mediated mechanism. Morphine further decreased 4T1 breast cancer cell invasion elicited by co-culture with RAW264.7. Reduction of MMP-9 expression and alternative activation of macrophages by morphine was confirmed using mouse bone marrow-derived macrophages. Taken together, our results indicate that morphine may modulate tumour aggressiveness by regulating macrophage protease production and M2 polarization within the tumour microenvironment.

  6. Adipose tissue macrophages in insulin-resistant subjects are associated with collagen VI and fibrosis and demonstrate alternative activation

    PubMed Central

    Spencer, Michael; Yao-Borengasser, Aiwei; Unal, Resat; Rasouli, Neda; Gurley, Catherine M.; Zhu, Beibei; Peterson, Charlotte A.

    2010-01-01

    Adipose tissue macrophages are associated with insulin resistance and are linked to changes in the extracellular matrix. To better characterize adipose macrophages, the extracellular matrix, and adipocyte-macrophage interactions, gene expression from adipose tissue and the stromal vascular fraction was assessed for markers of inflammation and fibrosis, and macrophages from obese and lean subjects were counted and characterized immunohistochemically. Coculture experiments examined the effects of adipocyte-macrophage interaction. Collagen VI gene expression was associated with insulin sensitivity and CD68 (r = −0.56 and 0.60, P < 0.0001) and with other markers of inflammation and fibrosis. Compared with adipose tissue from lean subjects, adipose tissue from obese subjects contained increased areas of fibrosis, which correlated inversely with insulin sensitivity (r = −0.58, P < 0.02) and positively with macrophage number (r = 0.70, P < 0.01). Although macrophages in crownlike structures (CLS) were more abundant in obese adipose tissue, the majority of macrophages were associated with fibrosis and were not organized in CLS. Macrophages in CLS were predominantly M1, but most other macrophages, particularly those in fibrotic areas, were M2 and also expressed CD150, a marker of M2c macrophages. Coculture of THP-1 macrophages with adipocytes promoted the M2 phenotype, with a lower level of IL-1 expression and a higher ratio of IL-10 to IL-12. Transforming growth factor-β (TGF-β) was more abundant in M2 macrophages and was further increased by coculture with adipocytes. Downstream effectors of TGF-β, such as plasminogen activator inhibitor-1, collagen VI, and phosphorylated Smad, were increased in macrophages and adipocytes. Thus adipose tissue of insulin-resistant humans demonstrated increased fibrosis, M2 macrophage abundance, and TGF-β activity. PMID:20841504

  7. Adipose tissue macrophages in insulin-resistant subjects are associated with collagen VI and fibrosis and demonstrate alternative activation.

    PubMed

    Spencer, Michael; Yao-Borengasser, Aiwei; Unal, Resat; Rasouli, Neda; Gurley, Catherine M; Zhu, Beibei; Peterson, Charlotte A; Kern, Philip A

    2010-12-01

    Adipose tissue macrophages are associated with insulin resistance and are linked to changes in the extracellular matrix. To better characterize adipose macrophages, the extracellular matrix, and adipocyte-macrophage interactions, gene expression from adipose tissue and the stromal vascular fraction was assessed for markers of inflammation and fibrosis, and macrophages from obese and lean subjects were counted and characterized immunohistochemically. Coculture experiments examined the effects of adipocyte-macrophage interaction. Collagen VI gene expression was associated with insulin sensitivity and CD68 (r = -0.56 and 0.60, P < 0.0001) and with other markers of inflammation and fibrosis. Compared with adipose tissue from lean subjects, adipose tissue from obese subjects contained increased areas of fibrosis, which correlated inversely with insulin sensitivity (r = -0.58, P < 0.02) and positively with macrophage number (r = 0.70, P < 0.01). Although macrophages in crownlike structures (CLS) were more abundant in obese adipose tissue, the majority of macrophages were associated with fibrosis and were not organized in CLS. Macrophages in CLS were predominantly M1, but most other macrophages, particularly those in fibrotic areas, were M2 and also expressed CD150, a marker of M2c macrophages. Coculture of THP-1 macrophages with adipocytes promoted the M2 phenotype, with a lower level of IL-1 expression and a higher ratio of IL-10 to IL-12. Transforming growth factor-β (TGF-β) was more abundant in M2 macrophages and was further increased by coculture with adipocytes. Downstream effectors of TGF-β, such as plasminogen activator inhibitor-1, collagen VI, and phosphorylated Smad, were increased in macrophages and adipocytes. Thus adipose tissue of insulin-resistant humans demonstrated increased fibrosis, M2 macrophage abundance, and TGF-β activity.

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

    PubMed

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

    2015-08-01

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

  9. Monocytes and macrophages, implications for breast cancer migration and stem cell-like activity and treatment

    PubMed Central

    Ward, Rebecca; Sims, Andrew H.; Lee, Alexander; Lo, Christina; Wynne, Luke; Yusuf, Humza; Gregson, Hannah; Lisanti, Michael P.; Sotgia, Federica; Landberg, Göran; Lamb, Rebecca

    2015-01-01

    Macrophages are a major cellular constituent of the tumour stroma and contribute to breast cancer prognosis. The precise role and treatment strategies to target macrophages remain elusive. As macrophage infiltration is associated with poor prognosis and high grade tumours we used the THP-1 cell line to model monocyte-macrophage differentiation in co-culture with four breast cancer cell lines (MCF7, T47D, MDA-MB-231, MDA-MB-468) to model in vivo cellular interactions. Polarisation into M1 and M2 subtypes was confirmed by specific cell marker expression of ROS and HLA-DR, respectively. Co-culture with all types of macrophage increased migration of ER-positive breast cancer cell lines, while M2-macrophages increased mammosphere formation, compared to M1-macrophages, in all breast cancer cells lines. Treatment of cells with Zoledronate in co-culture reduced the “pro-tumourigenic” effects (increased mammospheres/migration) exerted by macrophages. Direct treatment of breast cancer cells in homotypic culture was unable to reduce migration or mammosphere formation. Macrophages promote “pro-tumourigenic” cellular characteristics of breast cancer cell migration and stem cell activity. Zoledronate targets macrophages within the microenvironment which in turn, reduces the “pro-tumourigenic” characteristics of breast cancer cells. Zoledronate offers an exciting new treatment strategy for both primary and metastatic breast cancer. PMID:26008983

  10. Classically and alternatively activated bone marrow derived macrophages differ in cytoskeletal functions and migration towards specific CNS cell types

    PubMed Central

    2011-01-01

    Background Macrophages play an important role in neuroinflammatory diseases such as multiple sclerosis (MS) and spinal cord injury (SCI), being involved in both damage and repair. The divergent effects of macrophages might be explained by their different activation status: classically activated (CA/M1), pro-inflammatory, macrophages and alternatively activated (AA/M2), growth promoting, macrophages. Little is known about the effect of macrophages with these phenotypes in the central nervous system (CNS) and how they influence pathogenesis. The aim of this study was therefore to determine the characteristics of these phenotypically different macrophages in the context of the CNS in an in vitro setting. Results Here we show that bone marrow derived CA and AA macrophages have a distinct migratory capacity towards medium conditioned by various cell types of the CNS. AA macrophages were preferentially attracted by the low weight (< 10 kD) fraction of neuronal conditioned medium, while CA macrophages were attracted in higher numbers by astrocyte- and oligodendrocyte conditioned medium. Intrinsic motility was twice as high in AA macrophages compared to CA macrophages. The adhesion to extracellular matrix molecules (ECM) was significantly enhanced in CA macrophages compared to control and AA macrophages. The actin cytoskeleton was differentially organized between CA and AA macrophages, possibly due to greater activity of the GTPases RhoA and Rac in CA macrophages. Phagocytosis of myelin and neuronal fragments was increased in CA macrophages compared to AA macrophages. The increase in myelin phagocytosis was associated with higher expression of CR3/MAC-1 in CA macrophages. Conclusion In conclusion, since AA macrophages are more motile and are attracted by NCM, they are prone to migrate towards neurons in the CNS. CA macrophages have a lower motility and a stronger adhesion to ECM. In neuroinflammatory diseases the restricted migration and motility of CA macrophages might

  11. Cx3cr1 deficiency in mice attenuates hepatic granuloma formation during acute schistosomiasis by enhancing the M2-type polarization of macrophages.

    PubMed

    Ran, Lin; Yu, Qilin; Zhang, Shu; Xiong, Fei; Cheng, Jia; Yang, Ping; Xu, Jun-Fa; Nie, Hao; Zhong, Qin; Yang, Xueli; Yang, Fei; Gong, Quan; Kuczma, Michal; Kraj, Piotr; Gu, Weikuan; Ren, Bo-Xu; Wang, Cong-Yi

    2015-07-01

    Acute schistosomiasis is characterized by pro-inflammatory responses against tissue- or organ-trapped parasite eggs along with granuloma formation. Here, we describe studies in Cx3cr1(-/-) mice and demonstrate the role of Cx3cr1 in the pathoetiology of granuloma formation during acute schistosomiasis. Mice deficient in Cx3cr1 were protected from granuloma formation and hepatic injury induced by Schistosoma japonicum eggs, as manifested by reduced body weight loss and attenuated hepatomegaly along with preserved liver function. Notably, S. japonicum infection induced high levels of hepatic Cx3cr1 expression, which was predominantly expressed by infiltrating macrophages. Loss of Cx3cr1 rendered macrophages preferentially towards M2 polarization, which then led to a characteristic switch of the host immune defense from a conventional Th1 to a typical Th2 response during acute schistosomiasis. This immune switch caused by Cx3cr1 deficiency was probably associated with enhanced STAT6/PPAR-γ signaling and increased expression of indoleamine 2,3-dioxygenase (IDO), an enzyme that promotes M2 polarization of macrophages. Taken together, our data provide evidence suggesting that CX3CR1 could be a viable therapeutic target for treatment of acute schistosomiasis.

  12. Cx3cr1 deficiency in mice attenuates hepatic granuloma formation during acute schistosomiasis by enhancing the M2-type polarization of macrophages

    PubMed Central

    Ran, Lin; Yu, Qilin; Zhang, Shu; Xiong, Fei; Cheng, Jia; Yang, Ping; Xu, Jun-Fa; Nie, Hao; Zhong, Qin; Yang, Xueli; Yang, Fei; Gong, Quan; Kuczma, Michal; Kraj, Piotr; Gu, Weikuan; Ren, Bo-Xu; Wang, Cong-Yi

    2015-01-01

    ABSTRACT Acute schistosomiasis is characterized by pro-inflammatory responses against tissue- or organ-trapped parasite eggs along with granuloma formation. Here, we describe studies in Cx3cr1−/− mice and demonstrate the role of Cx3cr1 in the pathoetiology of granuloma formation during acute schistosomiasis. Mice deficient in Cx3cr1 were protected from granuloma formation and hepatic injury induced by Schistosoma japonicum eggs, as manifested by reduced body weight loss and attenuated hepatomegaly along with preserved liver function. Notably, S. japonicum infection induced high levels of hepatic Cx3cr1 expression, which was predominantly expressed by infiltrating macrophages. Loss of Cx3cr1 rendered macrophages preferentially towards M2 polarization, which then led to a characteristic switch of the host immune defense from a conventional Th1 to a typical Th2 response during acute schistosomiasis. This immune switch caused by Cx3cr1 deficiency was probably associated with enhanced STAT6/PPAR-γ signaling and increased expression of indoleamine 2,3-dioxygenase (IDO), an enzyme that promotes M2 polarization of macrophages. Taken together, our data provide evidence suggesting that CX3CR1 could be a viable therapeutic target for treatment of acute schistosomiasis. PMID:26035381

  13. Polarisation of Tumor-Associated Macrophages toward M2 Phenotype Correlates with Poor Response to Chemoradiation and Reduced Survival in Patients with Locally Advanced Cervical Cancer

    PubMed Central

    Martinelli, Enrica; Pedone Anchora, Luigi; Ferrandina, Gabriella; Tropeano, Giovanna; Fagotti, Anna; Scambia, Giovanni

    2015-01-01

    Objective we investigate the prognostic role of pre-treatment ratio between Type 1 (M1) and Type 2 (M2) tumor-associated macrophages (TAMs) in locally advanced cervical cancer (LACC) patients treated with chemoradiation (CT/RT). Methods 84 consecutive LACC patients treated with cisplatin-based CT/RT for a total dose of 50.0 Gy, followed by radical surgery were analysed. Double-staining immunohistochemistry of CD163/p-STAT, CD68/pSTAT1, CD163/c-MAF, and CD68/c-MAF was performed on tumor samples taken at the time of diagnosis. TAMs with CD163+pSTAT1+, or CD68+pSTAT1+ were defined M1; CD163+c-MAF+ or CD68+c-MAF+ defined the M2 phenotype. The number of M1 and M2 cells was counted at low magnification by evaluating for each case the same tumour area. The ratio between M1 and M2 (M1/M2) was finally calculated. Results At diagnosis, we observed a direct correlation between the number of circulating monocytes and of TAMs (p-value = 0.001). Patients with high M1/M2 experienced more frequently complete pathologic response (no residual tumor) to CT/RT, compared to cases with low M1/M2 (55.0% Vs 29.5%; p-value = 0.029). At multivariate analysis M1/M2 (OR = 2.067; p-value = 0.037) emerged as independent predictor of pathologic response to CT/RT. Women with high M1/M2 showed a longer 5-yrs Disease-free (67.2% Vs. 44.3%; p-value = 0.019), and 5-yrs Overall (69.3% Vs. 46.9%; p-value = 0.037) survival, compared to cases with low M1/M2. The presence of a high M1/M2 ratio was independently associated with an unfavourable survival outcome in multivariate analysis. Conclusions polarisation of TAMs toward a M2 phenotype, as reflected by a lower M1/M2 ratio, is an independent predictor of poor response to CT/RT, and shorter survival in LACC. PMID:26335330

  14. 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.

  15. 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.

  16. Role of Macrophage (M1 and M2) in Titanium-Dioxide Nanoparticle-Induced Oxidative Stress and Inflammatory Response in Rat.

    PubMed

    Kumar, Sumit; Meena, Ramovatar; Paulraj, R

    2016-12-01

    Titanium-dioxide nanoparticles (TNP) are used in various consumable goods. Evidence has demonstrated the cytotoxicity of TNPs, but exact mechanism is yet to be elucidated. The present study has been aimed at finding out the mechanism of TNP-induced toxicity in biological system. Different doses of anatase-TNPs administrated intravenously to Wistar rats for once a week for 1 month and properties of TH cells, macrophages, cytokines secretion, oxidative damage, apoptotic pathway, and hematological and pathological changes were investigated as downstream events of TNP-mediated cytotoxicity. Result suggests that TNPs induce TH1 and TH2 response as measured by immunophenotyping (interferon gamma (IFN-γ) and interleukin (IL)-4) of TH cells, causing induction of M1 (nitric oxide (NO), nitric oxide synthase (iNOS), NF-kappaB (NF-κB), cyclooxygenase-2 (COX-2), IL-1, IL-6, and TNF-α) and M2 (Arg-1, Ym1) macrophages response. At lower dose, TH1 or M1 response counteracted by TH2 or M2 response, resulting in insignificant oxidative damage. However, with increasing dose of TNPs, the M1 response was increased over M2 response resulting in significant tissue damage. The M1-induced inflammatory response was found to cause DNA and chromosomal damage resulting apoptosis induction via upregulation of Bax/Bcl-2 ratio and subsequent loss of mitochondrial membrane potential and cyto c release in splenocytes. The TNP-led inflammatory response also causes damage at different tissue levels.

  17. Characterization of the tumor microenvironment in primary cutaneous CD30-positive lymphoproliferative disorders: a predominance of CD163-positive M2 macrophages.

    PubMed

    De Souza, Aieska; Tinguely, Marianne; Burghart, Daniel R; Berisha, Arbeneshe; Mertz, Kirsten D; Kempf, Werner

    2016-07-01

    The tumor microenvironment is essential for tumor survival, growth and progression. There are only a few studies on the tumor microenvironment in cutaneous CD30-positive lymphoproliferative disorders. We assessed the composition of the tumor microenvironment using immunohistochemistry studies in skin biopsies from cases diagnosed with lymphomatoid papulosis (LyP: 18 specimens), primary cutaneous anaplastic large-cell lymphoma (PC-ALCL: 8 specimens), and reactive diseases harboring CD30-positive cells (18 specimens). The predominant cells present in LyP and PC-ALCL were CD163+ M2 macrophages (44.7%, 35%), followed by CD8+ tumor infiltrating lymphocytes (11%, 15%), FOXP3+ T-regulatory cells (9%, 4.5%) and programmed cell death 1(PD-1) + lymphocytes (2.2%, 6.8%). In contrast, CD30-positive reactive inflammatory and infectious disorders were characterized by higher numbers of CD123+ plasmacytoid dendritic cells (6.3%) when compared to LyP (1%), and PC-ALCL (1.1%). Key differences exist between the microenvironment of CD30-positive lymphoproliferative disorders and reactive conditions harboring CD30-positive lymphocytes. The high number of tumor associated macrophages, and the close vicinity of these immune cells to the CD30-positive tumor cells might suggest that tumor associated macrophages have direct influence on tumorigenesis in LyP and ALCL. Therefore, modulation of M2 macrophages may represent a new therapeutic strategy in cutaneous CD30-positive lymphoproliferative disorders. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Signaling of Prostaglandin E Receptors, EP3 and EP4 Facilitates Wound Healing and Lymphangiogenesis with Enhanced Recruitment of M2 Macrophages in Mice

    PubMed Central

    Hosono, Kanako; Isonaka, Risa; Kawakami, Tadashi; Narumiya, Shuh; Majima, Masataka

    2016-01-01

    Lymphangiogenesis plays an important role in homeostasis, metabolism, and immunity, and also occurs during wound-healing. Here, we examined the roles of prostaglandin E2 (PGE2) receptor (EP) signaling in enhancement of lymphangiogenesis in wound healing processes. The hole-punch was made in the ears of male C57BL/6 mice using a metal ear punch. Healing process and lymphangiogenesis together with macrophage recruitment were analyzed in EP knockout mice. Lymphangiogenesis was up-regulated in the granulation tissues at the margins of punched-hole wounds in mouse ears, and this increase was accompanied by increased expression levels of COX-2 and microsomal prostaglandin E synthase-1. Administration of celecoxib, a COX-2 inhibitor, suppressed lymphangiogenesis in the granulation tissues and reduced the induction of the pro-lymphangiogenic factors, vascular endothelial growth factor (VEGF) -C and VEGF-D. Topical applications of selective EP receptor agonists enhanced the expressions of lymphatic vessel endothelial hyaluronan receptor-1 and VEGF receptor-3. The wound-healing processes and recruitment of CD11b-positive macrophages, which produced VEGF-C and VEGF-D, were suppressed under COX-2 inhibition. Mice lacking either EP3 or EP4 exhibited reduced wound-healing, lymphangiogenesis and recruitment of M2 macrophages, compared with wild type mice. Proliferation of cultured human lymphatic endothelial cells was not detected under PGE2 stimulation. Lymphangiogenesis and recruitment of M2 macrophages that produced VEGF-C/D were suppressed in mice treated with a COX-2 inhibitor or lacking either EP3 or EP4 during wound healing. COX-2 and EP3/EP4 signaling may be novel targets to control lymphangiogenesis in vivo. PMID:27711210

  19. PI3K/Akt Signaling Pathway Modulates Influenza Virus Induced Mouse Alveolar Macrophage Polarization to M1/M2b

    PubMed Central

    Zhao, Xiangfeng; Dai, Jianping; Xiao, Xuejun; Wu, Liqi; Zeng, Jun; Sheng, Jiangtao; Su, Jinghua; Chen, Xiaoxuan; Wang, Gefei; Li, Kangsheng

    2014-01-01

    Macrophages polarized to M1 (pro-inflammation) or M2 (anti-inflammation) phenotypes in response to environmental signals. In this study, we examined the polarization of alveolar macrophage (AM), following induction by different influenza virus strains (ST169 (H1N1), ST602 (H3N2) and HKG9 (H9N2)). Macrophages from other tissues or cell line exert alternative responding pattern, and AM is necessary for investigating the respiratory system. AM polarized toward the M1 phenotype after 4 hours of infection by all three virus strains, and AM to presented M2b phenotype after 8 hours induction, and immunosuppressive phenotype after 24 hours of induction. Protein expression assay showed similar results as the gene expression analysis for phenotype verification. The ELISA assay showed that TNF-α secretion was up-regulated after 4 and 8 hours of infection by influenza viruses, and it returned to basal levels after 24 hours of infection. IL-10 expression was elevated after 8 and 24 hours of infection. Immunofluorescence showed that iNOS expression was up-regulated but not Arg1 expression. Influenza virus notably increased phospho-Akt but not phospho-Erk1/2 or phospho-p38, and the AM polarization pattern have been changed by LY294002 (PI3K inhibitor). In conclusion, our results demonstrate the dynamic polarization of AM induced by influenza viruses, and suggested that PI3K/Akt signaling pathway modulates AM polarization to M1/M2b. PMID:25105760

  20. Macrophage phenotypes in atherosclerosis.

    PubMed

    Colin, Sophie; Chinetti-Gbaguidi, Giulia; Staels, Bart

    2014-11-01

    Initiation and progression of atherosclerosis depend on local inflammation and accumulation of lipids in the vascular wall. Although many cells are involved in the development and progression of atherosclerosis, macrophages are fundamental contributors. For nearly a decade, the phenotypic heterogeneity and plasticity of macrophages has been studied. In atherosclerotic lesions, macrophages are submitted to a large variety of micro-environmental signals, such as oxidized lipids and cytokines, which influence the phenotypic polarization and activation of macrophages resulting in a dynamic plasticity. The macrophage phenotype spectrum is characterized, at the extremes, by the classical M1 macrophages induced by T-helper 1 (Th-1) cytokines and by the alternative M2 macrophages induced by Th-2 cytokines. M2 macrophages can be further classified into M2a, M2b, M2c, and M2d subtypes. More recently, additional plaque-specific macrophage phenotypes have been identified, termed as Mox, Mhem, and M4. Understanding the mechanisms and functional consequences of the phenotypic heterogeneity of macrophages will contribute to determine their potential role in lesion development and plaque stability. Furthermore, research on macrophage plasticity could lead to novel therapeutic approaches to counteract cardiovascular diseases such as atherosclerosis. The present review summarizes our current knowledge on macrophage subsets in atherosclerotic plaques and mechanism behind the modulation of the macrophage phenotype.

  1. Model-Based Characterization of Inflammatory Gene Expression Patterns of Activated Macrophages

    PubMed Central

    Ehlting, Christian; Thomas, Maria; Zanger, Ulrich M.; Sawodny, Oliver; Häussinger, Dieter; Bode, Johannes G.

    2016-01-01

    Macrophages are cells with remarkable plasticity. They integrate signals from their microenvironment leading to context-dependent polarization into classically (M1) or alternatively (M2) activated macrophages, representing two extremes of a broad spectrum of divergent phenotypes. Thereby, macrophages deliver protective and pro-regenerative signals towards injured tissue but, depending on the eliciting damage, may also be responsible for the generation and aggravation of tissue injury. Although incompletely understood, there is emerging evidence that macrophage polarization is critical for these antagonistic roles. To identify activation-specific expression patterns of chemokines and cytokines that may confer these distinct effects a systems biology approach was applied. A comprehensive literature-based Boolean model was developed to describe the M1 (LPS-activated) and M2 (IL-4/13-activated) polarization types. The model was validated using high-throughput transcript expression data from murine bone marrow derived macrophages. By dynamic modeling of gene expression, the chronology of pathway activation and autocrine signaling was estimated. Our results provide a deepened understanding of the physiological balance leading to M1/M2 activation, indicating the relevance of co-regulatory signals at the level of Akt1 or Akt2 that may be important for directing macrophage polarization. PMID:27464342

  2. Respiratory syncytial virus M2-1 protein induces the activation of nuclear factor kappa B

    SciTech Connect

    Reimers, Kerstin . E-mail: reimers.kerstin@mh-hannover.de; Buchholz, Katja; Werchau, Hermann

    2005-01-20

    Respiratory syncytial virus (RSV) induces the production of a number of cytokines and chemokines by activation of nuclear factor kappa B (NF-{kappa}B). The activation of NF-{kappa}B has been shown to depend on viral replication in the infected cells. In this study, we demonstrate that expression of RSV M2-1 protein, a transcriptional processivity and anti-termination factor, is sufficient to activate NF-{kappa}B in A549 cells. Electromobility shift assays show increased NF-{kappa}B complexes in the nuclei of M2-1-expressing cells. M2-1 protein is found in nuclei of M2-1-expressing cells and in RSV-infected cells. Co-immunoprecipitations of nuclear extracts of M2-1-expressing cells and of RSV-infected cells revealed an association of M2-1 with Rel A protein. Furthermore, the activation of NF-{kappa}B depends on the C-terminus of the RSV M2-1 protein, as shown by NF-{kappa}B-induced gene expression of a reporter gene construct.

  3. A humanized anti-M2 scFv shows protective in vitro activity against influenza

    SciTech Connect

    Bradbury, Andrew M; Velappan, Nileena; Schmidt, Jurgen G

    2008-01-01

    M2 is one of the most conserved influenza proteins, and has been widely prospected as a potential universal vaccine target, with protection predominantly mediated by antibodies. In this paper we describe the creation of a humanized single chain Fv from 14C2, a potent monoclonal antibody against M2. We show that the humanized scFv demonstrates similar activity to the parental mAb: it is able to recognize M2 in its native context on cell surfaces and is able to show protective in vitro activity against influenza, and so represents a potential lead antibody candidate for universal prophylactic or therapeutic intervention in influenza.

  4. Membrane-Tethered MUC1 Mucin Counter-Regulates the Phagocytic Activity of Macrophages.

    PubMed

    Kato, Kosuke; Uchino, Reina; Lillehoj, Erik P; Knox, Kenneth; Lin, Yong; Kim, K Chul

    2016-04-01

    MUC1 (MUC in human; Muc in animals) is a transmembrane mucin glycoprotein expressed in mucosal epithelial cells and hematopoietic cells. MUC1 is involved in the resolution of inflammation during airway Pseudomonas aeruginosa (Pa) infection by suppressing Toll-like receptor signaling in airway epithelial cells. Although alveolar macrophages are recognized as critical mediators of cell-mediated immunity against microorganisms inhaled into the airways, the role of MUC1 in regulating their response is unknown. The aims of this study were to determine whether macrophages express MUC1, and, if so, whether MUC1 expression might be associated with macrophage M0/M1/M2 differentiation or phagocytic activity. Human and mouse MUC1/Muc1 expression was drastically up-regulated in classically activated (M1) macrophages compared with nonactivated (M0) and alternatively activated (M2) macrophages. M1 polarization and Pa stimulation each increased MUC1 ectodomain shedding from the macrophage surface in a TNF-α-converting enzyme-dependent manner. MUC1/Muc1 deficiency in M0 macrophages increased adhesion and phagocytosis of Pa and Escherichia coli compared with MUC1/Muc1-expressing cells, and attenuation of phagocytosis by MUC1 was augmented after polarization into M1 macrophages compared with M0 macrophages. Finally, MUC1/Muc1 deficiency in macrophages increased reactive oxygen species production and TNF-α release in response to Pa compared with MUC1/Muc1-sufficient cells. These results indicate that MUC1/Muc1 expression by macrophages is predominantly in the M1 subtype, and that MUC1/Muc1 expression in these cells decreases their phagocytic activity in an antiinflammatory manner.

  5. Involvement of Proteasome and Macrophages M2 in the Protection Afforded by Telmisartan against the Acute Myocardial Infarction in Zucker Diabetic Fatty Rats with Metabolic Syndrome

    PubMed Central

    Di Filippo, C.; Rossi, C.; Ferraro, B.; Maisto, R.; De Angelis, A.; Ferraraccio, F.; Rotondo, A.; D'Amico, M.

    2014-01-01

    This study investigated the involvement of proteasome and macrophages M2 in the protection afforded by telmisartan against the acute myocardial infarction in Zucker diabetic fatty (ZDF) rats with metabolic syndrome. ZDF rats were treated for three weeks with telmisartan at doses of 7 and 12 mg/kg/day. After treatment, rats were subjected to a 25 min occlusion of the left descending coronary artery followed by 2 h reperfusion (I/R). At the end of the I/R period, biochemical, immunohistochemical, and echocardiographic evaluations were done. Telmisartan treatment (7 mg/kg and 12 mg/kg) reduced the myocardial infarct size, the expression of proteasome subunits 20S and 26S, and the protein ubiquitin within the heart. The compound has led to an increased M2 macrophage phenotype within the cardiac specimens and a modification of the cardiac cytokine and chemokine profile. This was functionally translated in improved cardiac performance as evidenced by echography after 2 h reperfusion. 7 mg/kg/day telmisartan was sufficient to improve the left ventricular ejection fraction LVEF of the rat heart recorded after I/R (e.g., vehicle 38 ± 2.2%; telmisartan 54 ± 2.7%) and was sufficient to improve the diastolic function and the myocardial performance index up to values of 0.6 ± 0.01 measured after I/R. PMID:25110402

  6. Measurement of Macrophage Activation by Chemiluminescence.

    DTIC Science & Technology

    1986-05-01

    cytotoxicity and oxidative metabolism including those reactions responsible for chemiluminescence (11-13). This activation has been correlated with an...order to assay this event, enhancers of chemiluminescence such as lucigenin and luminol are required to amplify the reaction . Enhancement of CL by...Maleic vinyl ether activation of murine *. macrophages against lung metastasizing tumors. Cancer Res 41:3901-3906 1981 4. Fidler I.J., Sone, S., Fogler

  7. Intracellular multiplication of Paracoccidioides brasiliensis in macrophages: killing and restriction of multiplication by activated macrophages.

    PubMed Central

    Brummer, E; Hanson, L H; Restrepo, A; Stevens, D A

    1989-01-01

    The effect of coculturing yeast-form Paracoccidioides brasiliensis with murine cells was studied. Coculture of resident peritoneal or pulmonary macrophages with P. brasiliensis for 72 h dramatically enhanced fungal multiplication 19.3 +/- 2.4- and 4.7 +/- 0.8-fold, respectively, compared with cocultures with lymph node cells or complete tissue culture medium alone. Support of P. brasiliensis multiplication by resident peritoneal macrophages was macrophage dose dependent. Lysates of macrophages, supernatants from macrophage cultures, or McVeigh-Morton broth, like complete tissue culture medium, did not support multiplication of P. brasiliensis in 72-h cultures. Time course microscopic studies of cocultures in slide wells showed that macrophages ingested P. brasiliensis cells and that the ingested cells multiplied intracellularly. In sharp contrast to resident macrophages, lymphokine-activated peritoneal and pulmonary macrophages not only prevented multiplication but reduced inoculum CFU by 96 and 100%, respectively, in 72 h. Microscopic studies confirmed killing and digestion of P. brasiliensis ingested by activated macrophages in 48 h. These findings indicate that resident macrophages are permissive for intracellular multiplication of P. brasiliensis and that this could be a factor in pathogenicity. By contrast, activated macrophages are fungicidal for P. brasiliensis. Images PMID:2744848

  8. Adipogenic role of alternatively activated macrophages in β-adrenergic remodeling of white adipose tissue.

    PubMed

    Lee, Yun-Hee; Kim, Sang-Nam; Kwon, Hyun-Jung; Maddipati, Krishna Rao; Granneman, James G

    2016-01-01

    De novo brown adipogenesis involves the proliferation and differentiation of progenitors, yet the mechanisms that guide these events in vivo are poorly understood. We previously demonstrated that treatment with a β3-adrenergic receptor (ADRB3) agonist triggers brown/beige adipogenesis in gonadal white adipose tissue following adipocyte death and clearance by tissue macrophages. The close physical relationship between adipocyte progenitors and tissue macrophages suggested that the macrophages that clear dying adipocytes might generate proadipogenic factors. Flow cytometric analysis of macrophages from mice treated with CL 316,243 identified a subpopulation that contained elevated lipid and expressed CD44. Lipidomic analysis of fluorescence-activated cell sorting-isolated macrophages demonstrated that CD44+ macrophages contained four- to five-fold higher levels of the endogenous peroxisome-proliferator activated receptor gamma (PPARγ) ligands 9-hydroxyoctadecadienoic acid (HODE), and 13-HODE compared with CD44- macrophages. Gene expression profiling and immunohistochemistry demonstrated that ADRB3 agonist treatment upregulated expression of ALOX15, the lipoxygenase responsible for generating 9-HODE and 13-HODE. Using an in vitro model of adipocyte efferocytosis, we found that IL-4-primed tissue macrophages accumulated lipid from dying fat cells and upregulated expression of Alox15. Furthermore, treatment of differentiating adipocytes with 9-HODE and 13-HODE potentiated brown/beige adipogenesis. Collectively, these data indicate that noninflammatory removal of adipocyte remnants and coordinated generation of PPARγ ligands by M2 macrophages provides localized adipogenic signals to support de novo brown/beige adipogenesis.

  9. Arginase activity in alternatively activated macrophages protects PI3Kp110δ deficient mice from dextran sodium sulfate induced intestinal inflammation.

    PubMed

    Weisser, Shelley B; Kozicky, Lisa K; Brugger, Hayley K; Ngoh, Eyler N; Cheung, Bonnie; Jen, Roger; Menzies, Susan C; Samarakoon, Asanga; Murray, Peter J; Lim, C James; Johnson, Pauline; Boucher, Jean-Luc; van Rooijen, Nico; Sly, Laura M

    2014-11-01

    Alternatively activated or M2 macrophages have been reported to protect mice from intestinal inflammation, but the mechanism of protection has not been elucidated. In this study, we demonstrate that mice deficient in the p110δ catalytic subunit activity of class I phosphatidylinositol 3-kinase (PI3Kp110δ) have increased clinical disease activity and histological damage during dextran sodium sulfate (DSS) induced colitis. Increased disease severity in PI3Kp110δ-deficient mice is dependent on professional phagocytes and correlates with reduced numbers of arginase I+ M2 macrophages in the colon and increased production of inflammatory nitric oxide. We further demonstrate that PI3Kp110δ-deficient macrophages are defective in their ability to induce arginase I when skewed to an M2 phenotype with IL-4. Importantly, adoptive transfer of IL-4-treated macrophages derived from WT mice, but not those from PI3Kp110δ-deficient mice, protects mice during DSS-induced colitis. Moreover, M2 macrophages mediated protection is lost when mice are cotreated with inhibitors that block arginase activity or during adoptive transfer of arginase I deficient M2 macrophages. Taken together, our data demonstrate that arginase I activity is required for M2 macrophages mediated protection during DSS-induced colitis in PI3Kp110δ-deficient mice.

  10. Leptin Deficiency Shifts Mast Cells toward Anti-Inflammatory Actions and Protects Mice from Obesity and Diabetes by Polarizing M2 Macrophages.

    PubMed

    Zhou, Yi; Yu, Xueqing; Chen, Huimei; Sjöberg, Sara; Roux, Joséphine; Zhang, Lijun; Ivoulsou, Al-Habib; Bensaid, Farid; Liu, Cong-Lin; Liu, Jian; Tordjman, Joan; Clement, Karine; Lee, Chih-Hao; Hotamisligil, Gokhan S; Libby, Peter; Shi, Guo-Ping

    2015-12-01

    Mast cells (MCs) contribute to the pathogenesis of obesity and diabetes. This study demonstrates that leptin deficiency slants MCs toward anti-inflammatory functions. MCs in the white adipose tissue (WAT) of lean humans and mice express negligible leptin. Adoptive transfer of leptin-deficient MCs expanded ex vivo mitigates diet-induced and pre-established obesity and diabetes in mice. Mechanistic studies show that leptin-deficient MCs polarize macrophages from M1 to M2 functions because of impaired cell signaling and an altered balance between pro- and anti-inflammatory cytokines, but do not affect T cell differentiation. Rampant body weight gain in ob/ob mice, a strain that lacks leptin, associates with reduced MC content in WAT. In ob/ob mice, genetic depletion of MCs exacerbates obesity and diabetes, and repopulation of ex vivo expanded ob/ob MCs ameliorates these diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Leptin deficiency shifts mast cells toward anti-inflammatory actions and protects mice from obesity and diabetes by polarizing M2 macrophages

    PubMed Central

    Zhou, Yi; Yu, Xueqing; Chen, Huimei; Sjöberg, Sara; Roux, Joséphine; Zhang, Lijun; Ivoulsou, Al-Habib; Bensaid, Farid; Liu, Conglin; Liu, Jian; Tordjman, Joan; Clement, Karine; Lee, Chih-Hao; Hotamisligil, Gokhan S.; Libby, Peter; Shi, Guo-Ping

    2015-01-01

    SUMMARY Mast cells (MCs) contribute to the pathogenesis of obesity and diabetes. This study demonstrates that leptin deficiency slants MCs toward anti-inflammatory functions. MCs in the white adipose tissues (WAT) of lean humans and mice express negligible leptin. Adoptive transfer of leptin-deficient MCs expanded ex vivo mitigates diet-induced and pre-established obesity and diabetes in mice. Mechanistic studies show that leptin-deficient MCs polarize macrophages from M1 to M2 functions because of impaired cell signaling and an altered balance between pro- and anti-inflammatory cytokines, but do not affect T-cell differentiation. Rampant body weight gain in ob/ob mice, a strain that lacks leptin, associates with reduced MC content in WAT. In ob/ob mice, genetic depletion of MCs exacerbates obesity and diabetes, and repopulation of ex vivo expanded ob/ob MCs ameliorates these diseases. PMID:26481668

  12. Redefining the transcriptional regulatory dynamics of classically and alternatively activated macrophages by deepCAGE transcriptomics

    PubMed Central

    Roy, Sugata; Schmeier, Sebastian; Arner, Erik; Alam, Tanvir; Parihar, Suraj P.; Ozturk, Mumin; Tamgue, Ousman; Kawaji, Hideya; de Hoon, Michiel J. L.; Itoh, Masayoshi; Lassmann, Timo; Carninci, Piero; Hayashizaki, Yoshihide; Forrest, Alistair R. R.; Bajic, Vladimir B.; Guler, Reto; Consortium, FANTOM; Brombacher, Frank; Suzuki, Harukazu

    2015-01-01

    Classically or alternatively activated macrophages (M1 and M2, respectively) play distinct and important roles for microbiocidal activity, regulation of inflammation and tissue homeostasis. Despite this, their transcriptional regulatory dynamics are poorly understood. Using promoter-level expression profiling by non-biased deepCAGE we have studied the transcriptional dynamics of classically and alternatively activated macrophages. Transcription factor (TF) binding motif activity analysis revealed four motifs, NFKB1_REL_RELA, IRF1,2, IRF7 and TBP that are commonly activated but have distinct activity dynamics in M1 and M2 activation. We observe matching changes in the expression profiles of the corresponding TFs and show that only a restricted set of TFs change expression. There is an overall drastic and transient up-regulation in M1 and a weaker and more sustainable up-regulation in M2. Novel TFs, such as Thap6, Maff, (M1) and Hivep1, Nfil3, Prdm1, (M2) among others, were suggested to be involved in the activation processes. Additionally, 52 (M1) and 67 (M2) novel differentially expressed genes and, for the first time, several differentially expressed long non-coding RNA (lncRNA) transcriptome markers were identified. In conclusion, the finding of novel motifs, TFs and protein-coding and lncRNA genes is an important step forward to fully understand the transcriptional machinery of macrophage activation. PMID:26117544

  13. High numbers of macrophages, especially M2-like (CD163-positive), correlate with hyaluronan accumulation and poor outcome in breast cancer.

    PubMed

    Tiainen, Satu; Tumelius, Ritva; Rilla, Kirsi; Hämäläinen, Kirsi; Tammi, Markku; Tammi, Raija; Kosma, Veli-Matti; Oikari, Sanna; Auvinen, Päivi

    2015-05-01

    High amounts of tumour-associated macrophages (TAMs) and hyaluronan (HA) correlate with tumour aggressiveness in breast cancer, but the relationship between these parameters is unclear. The aim of this study was to assay the numbers of TAMs in 278 human breast cancer cases, and their correlations with HA-related factors, clinical variables, and outcome. The immunoreactivities for CD163 and CD68 were considered as indicators for M2-like and all TAMs, respectively. The numbers of TAMs were counted in at least four hot spots, and averaged to represent the numbers of TAMs in each section. In the statistical analyses, the numbers were graded as either low or high according to the median. High numbers of TAMs correlated with a high tumour HA content, HA synthases, CD44 positivity, and poor outcome. The number of CD163-positive cells represented a strong independent prognostic factor. There was also a significant correlation between obesity and a high number of CD163-positive cells. Concurrent increases in TAMs and HA in breast cancer indicate that the accumulation of HA facilitates macrophage infiltration and inflammatory responses during human breast cancer progression. © 2014 John Wiley & Sons Ltd.

  14. Pivotal Advance: Tumor-mediated induction of myeloid-derived suppressor cells and M2-polarized macrophages by altering intracellular PGE₂ catabolism in myeloid cells.

    PubMed

    Eruslanov, Evgeniy; Daurkin, Irina; Ortiz, Javier; Vieweg, Johannes; Kusmartsev, Sergei

    2010-11-01

    Recent studies suggest that tumor-infiltrated myeloid cells frequently up-regulate COX-2 expression and have enhanced PGE₂ metabolism. This may affect the maturation and immune function of tumor-infiltrated antigen-presenting cells. In vitro studies demonstrate that tumor-derived factors can skew GM-CSF-driven differentiation of T(h)1-oriented myeloid APCs into M2-oriented Ly6C(+)F4/80(+) MDSCs or Ly6C(-)F4/80(+) arginase-expressing macrophages. These changes enable myeloid cells to produce substantial amounts of IL-10, VEGF, and MIP-2. The tumor-mediated inhibition of APC differentiation was associated with the up-regulated expression of PGE₂-forming enzymes COX-2, mPGES1 in myeloid cells, and the simultaneous repression of PGE(2)-catabolizing enzyme 15-PGDH. The presence of tumor-derived factors also led to a reduced expression of PGT but promoted the up-regulation of MRP4, which works as a PGE₂ efflux receptor. Addition of COX-2 inhibitor to the BM cell cultures could prevent the tumor-induced skewing of myeloid cell differentiation, partially restoring cell phenotype and down-regulating the arginase expression in the myeloid APCs. Our study suggests that tumors impair the intracellular PGE(2) catabolism in myeloid cells through simultaneous stimulation of PGE(2)-forming enzymes and inhibition of PGE₂-degrading systems. This tumor-induced dichotomy drives the development of M2-oriented, arginase-expressing macrophages or the MDSC, which can be seen frequently among tumor-infiltrated myeloid cells.

  15. RGS4 regulates partial agonism of the M2 muscarinic receptor-activated K+ currents

    PubMed Central

    Chen, I-Shan; Furutani, Kazuharu; Inanobe, Atsushi; Kurachi, Yoshihisa

    2014-01-01

    Partial agonists are used clinically to avoid overstimulation of receptor-mediated signalling, as they produce a submaximal response even at 100% receptor occupancy. The submaximal efficacy of partial agonists is due to conformational change of the agonist–receptor complex, which reduces effector activation. In addition to signalling activators, several regulators help control intracellular signal transductions. However, it remains unclear whether these signalling regulators contribute to partial agonism. Here we show that regulator of G-protein signalling (RGS) 4 is a determinant for partial agonism of the M2 muscarinic receptor (M2R). In rat atrial myocytes, pilocarpine evoked smaller G-protein-gated K+ inwardly rectifying (KG) currents than those evoked by ACh. In a Xenopus oocyte expression system, pilocarpine acted as a partial agonist in the presence of RGS4 as it did in atrial myocytes, while it acted like a full agonist in the absence of RGS4. Functional couplings within the agonist–receptor complex/G-protein/RGS4 system controlled the efficacy of pilocarpine relative to ACh. The pilocarpine–M2R complex suppressed G-protein-mediated activation of KG currents via RGS4. Our results demonstrate that partial agonism of M2R is regulated by the RGS4-mediated inhibition of G-protein signalling. This finding helps us to understand the molecular components and mechanism underlying the partial agonism of M2R-mediated physiological responses. PMID:24421355

  16. Macrophage in chronic kidney disease

    PubMed Central

    Flaquer, Maria; Cruzado, Josep M.

    2016-01-01

    Chronic kidney disease (CKD) has become a major health problem worldwide. This review describes the role of macrophages in CKD and highlights the importance of anti-inflammatory M2 macrophage activation in both renal fibrosis and wound healing processes. Furthermore, the mechanisms by which M2 macrophages induce renal repair and regeneration are still under debate and currently demand more attention. The M1/M2 macrophage balance is related to the renal microenvironment and could influence CKD progression. In fact, an inflammatory renal environment and M2 plasticity can be the major hurdles to establishing macrophage cell-based therapies in CKD. M2 macrophage cell-based therapy is promising if the M2 phenotype remains stable and is ‘fixed’ by in vitro manipulation. However, a greater understanding of phenotype polarization is still required. Moreover, better strategies and targets to induce reparative macrophages in vivo should guide future investigations in order to abate kidney diseases. PMID:27994852

  17. Activation of the α7 nicotinic receptor promotes lipopolysaccharide-induced conversion of M1 microglia to M2

    PubMed Central

    Zhang, Qichun; Lu, Ying; Bian, Huimin; Guo, Liwei; Zhu, Huaxu

    2017-01-01

    The α7 subtype of the nicotinic acetylcholine receptor (α7 nAChR) plays an essential role in the cholinergic anti-inflammatory pathway that regulates macrophage/microglia function in inflammation. Similar to M1 and M2 macrophages, M1 and M2 microglia exhibit pro-inflammation and anti-inflammation properties, respectively. In the present study, we analyzed function-associated phenotypes to detect the transformation of microglia with activation of α7 nAChRs. We used lentivirus-mediated shRNA to knockdown the expression of α7 nAChR in BV-2 microglia incubated with lipopolysaccharides (LPS, 0.1 μg/mL) and measured the acetylcholine (Ach, 1 μg/mL)-mediated release of cytokines, such as IL-1β, IL-4, IL-6, and IL-10, in the culture supernatant via radioimmunoassay. After stimulation with Ach, the expression of typical biomarkers for different microglia phenotypes, Iba-1 and Arg-1, was determined by cellular immunofluorescence. Furthermore, the expression of signaling molecules, including p38, JAK2/STAT3, PI3K/Akt and miR-124, was analyzed via western blotting and real-time PCR. We found that Ach inhibited LPS-induced IL-1β and IL-6 elevation and promoted IL-4 and IL-10 production and that knockdown of the α7 nAChR abolished these effects of Ach. In addition, Ach decreased LPS-induced Iba-1 expression and increased Arg-1 levels in an α7 nAChR-dependent manner. The LPS-inhibited activation of JAK2/STAT3 and PI3K/Akt was also rescued by Ach, an effect that was blocked by knockdown of the α7 nAChR. In contrast, Ach triggered the phosphorylation of JAK2 and STAT3 that was otherwise inactivated by LPS in BV-2 cells. Finally, the levels of miR-124 and downstream targets C/EBPα and PU.1 were significantly enhanced in LPS-treated BV-2 microglia, and the effect of Ach on this signaling pathway was blocked by α7 nAChR knockdown as expected. Overall, our data demonstrate that activation ofα7 nAChRs inhibits the transformation of M1 microglia and promotes the M2

  18. Modulation of macrophage activation by prostaglandins

    PubMed Central

    Carnuccio, R.; D'Acquisto, F.; Rosa, M. Di

    1996-01-01

    The effect of prostaglandtn E2, iloprost and cAMP on both nitric oxide and tumour necrosis factor-α release in J774 macrophages has been studied. Both prostaglandin E2 and iloprost inhibited, in a concentration-dependent fashion, the lipopolysaccharide-induced generation of nitric oxide and tumour necrosis factor-α. The inhibitory effect of these prostanoids seems to be mediated by an increase of the second messenger cAMP since it was mimicked by dibutyryl cAMP and potentiated by the selective type IV phosphodiesterase inhibitor RO-20-1724. Our results suggest that the inhibition of nitric oxide release by prostaglandin E2 and iloprost in lipopolysaccharide-activated J774 macrophages may be secondary to the inhibition of tumour necrosis factor-α generation, which in turn is likely to be mediated by cAMP. PMID:18475691

  19. ROS sets the stage for macrophage differentiation.

    PubMed

    Covarrubias, Anthony; Byles, Vanessa; Horng, Tiffany

    2013-08-01

    While M1 macrophages are highly pro-inflammatory and microbicidal, M2 macrophages and the related tumor associated macrophages (TAMs) regulate tissue remodeling and angiogenesis and can display immunomodulatory activity. In July issue of Cell Research, Zhang et al. show that ROS production, critical for the activation and functions of M1 macrophages, is necessary for the differentiation of M2 macrophages and TAMs, and that antioxidant therapy blocks TAM differentiation and tumorigenesis in mouse models of cancer.

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

    PubMed

    Cheng, Yahui; Tian, Yuanyao; Xia, Jialu; Wu, Xiaoqin; Yang, Yang; Li, Xiaofeng; Huang, Cheng; Meng, Xiaoming; Ma, Taotao; Li, Jun

    2017-02-15

    Activation of Kupffer cells (KCs) plays a pivotal role in the pathogenesis of liver fibrosis. The progression and reversal of CCl4-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 CCl4-induced mice but increased to a near-normal level in reversed mice. Moreover, PTEN was significantly decreased in IL4-induced RAW 264.7 cells in vitro and lower expression of PTEN was observed in M2 macrophages in vivo. In addition, loss- and gain-of-function studies suggested that PTEN regulates M2 macrophages polarization via activation of PI3K/Akt/STAT6 signaling, but had a limited effect on M1 macrophages polarization in vitro. Additionally, Ly294002, a chemical inhibitor of PI3K/Akt, could dramatically down-regulate the hallmarks of M2 macrophages. In conclusion, PTEN mediates macrophages activation by PI3K/Akt/STAT6 signaling pathway, which provides novel compelling evidences on the potential of PTEN in liver injury and opens new cellular target for the pharmacological therapy of liver fibrosis.

  1. 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

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

    PubMed

    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

    2016-02-19

    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.

  3. Guanylate-binding protein 5 is a marker of interferon-γ-induced classically activated macrophages

    PubMed Central

    Fujiwara, Yukio; Hizukuri, Yoshiyuki; Yamashiro, Kyoko; Makita, Naoyuki; Ohnishi, Koji; Takeya, Motohiro; Komohara, Yoshihiro; Hayashi, Yasuhiro

    2016-01-01

    Macrophage activation is the main immunological process occurring during the development of several diseases, and the heterogeneity of macrophage activation or differentiation has been suggested to be involved in disease progression. In the present study, we attempted to identify molecules specifically expressed on human classically activated macrophages (M1) to investigate the significance of the M1-like phenotype in human diseases. Human monocyte-derived macrophages were differentiated into M1, M2a, M2b and M2c phenotypes, and also M1(−) (the M1 phenotype differentiated with interferon-γ) to eliminate the strong effects of lipopolysaccharides (LPS) on the gene expression profile. The gene expression profiles of those macrophage phenotypes were analyzed by a cDNA microarray analysis and were used for a bioinformatics examination to identify the markers of the M1 phenotype that are expressed in both M1 and M1(−). The gene expression profiles of murine macrophages were also evaluated. We identified guanylate-binding protein 5 (GBP5), which is associated nucleotide-binding domain and leucine-rich repeat containing gene family, pyrin domain containing 3 (NLRP3)-mediated inflammasome assembly in the M1 macrophages of both humans and mice. Notably, the expression of GBP5 protein was detected in cultured M1(−) as well as in M1 macrophages by western blotting, which means that GBP5 is a more generalized marker of the M1 phenotype compared with the M1 markers that can be induced by LPS stimulation. GBP5 is a useful candidate marker of the M1 phenotype. PMID:27990286

  4. Guanylate-binding protein 5 is a marker of interferon-γ-induced classically activated macrophages.

    PubMed

    Fujiwara, Yukio; Hizukuri, Yoshiyuki; Yamashiro, Kyoko; Makita, Naoyuki; Ohnishi, Koji; Takeya, Motohiro; Komohara, Yoshihiro; Hayashi, Yasuhiro

    2016-11-01

    Macrophage activation is the main immunological process occurring during the development of several diseases, and the heterogeneity of macrophage activation or differentiation has been suggested to be involved in disease progression. In the present study, we attempted to identify molecules specifically expressed on human classically activated macrophages (M1) to investigate the significance of the M1-like phenotype in human diseases. Human monocyte-derived macrophages were differentiated into M1, M2a, M2b and M2c phenotypes, and also M1(-) (the M1 phenotype differentiated with interferon-γ) to eliminate the strong effects of lipopolysaccharides (LPS) on the gene expression profile. The gene expression profiles of those macrophage phenotypes were analyzed by a cDNA microarray analysis and were used for a bioinformatics examination to identify the markers of the M1 phenotype that are expressed in both M1 and M1(-). The gene expression profiles of murine macrophages were also evaluated. We identified guanylate-binding protein 5 (GBP5), which is associated nucleotide-binding domain and leucine-rich repeat containing gene family, pyrin domain containing 3 (NLRP3)-mediated inflammasome assembly in the M1 macrophages of both humans and mice. Notably, the expression of GBP5 protein was detected in cultured M1(-) as well as in M1 macrophages by western blotting, which means that GBP5 is a more generalized marker of the M1 phenotype compared with the M1 markers that can be induced by LPS stimulation. GBP5 is a useful candidate marker of the M1 phenotype.

  5. Direct imaging of macrophage activation during PDT treatment

    NASA Astrophysics Data System (ADS)

    Song, Sheng; Zhou, Feifan; Chen, Wei R.; Xing, Da

    2012-03-01

    Mounting evidence describes a more complex progress of macrophage activation during photodynamic therapy (PDT), which performing distinct immunological functions and different physiologies on surrounding cells and tissues. Macrophage-targeted PDT has been applied in the selective killing of cells involved in inflammation and tumor. We have previously shown that PDT-mediated tumor cells apoptosis can induce a higher level immune response than necrosis, and enhance the macrophage activation. However, the molecular mechanism of macrophage activation during PDT-induced apoptotic cells (AC) still unclear. Here, we use confocal microscopy to image the phagocytosis of tumor cells by macrophages. We also observed that PDT-treated AC can activate Toll-like receptors (TLRs) which are present on macrophages surface. Besides, the increase in nitric oxide (NO) formation in macrophages was detected in real time by a laser scanning microscopy. This study provided more details for understanding the molecular mechanism of the immune response induced by PDT-treated AC.

  6. Direct imaging of macrophage activation during PDT treatment

    NASA Astrophysics Data System (ADS)

    Song, Sheng; Zhou, Feifan; Chen, Wei R.; Xing, Da

    2011-11-01

    Mounting evidence describes a more complex progress of macrophage activation during photodynamic therapy (PDT), which performing distinct immunological functions and different physiologies on surrounding cells and tissues. Macrophage-targeted PDT has been applied in the selective killing of cells involved in inflammation and tumor. We have previously shown that PDT-mediated tumor cells apoptosis can induce a higher level immune response than necrosis, and enhance the macrophage activation. However, the molecular mechanism of macrophage activation during PDT-induced apoptotic cells (AC) still unclear. Here, we use confocal microscopy to image the phagocytosis of tumor cells by macrophages. We also observed that PDT-treated AC can activate Toll-like receptors (TLRs) which are present on macrophages surface. Besides, the increase in nitric oxide (NO) formation in macrophages was detected in real time by a laser scanning microscopy. This study provided more details for understanding the molecular mechanism of the immune response induced by PDT-treated AC.

  7. The predominance of alternatively activated macrophages following challenge with cell wall peptide-polysaccharide after prior infection with Sporothrix schenckii.

    PubMed

    Alegranci, Pamela; de Abreu Ribeiro, Livia Carolina; Ferreira, Lucas Souza; Negrini, Thais de Cássia; Maia, Danielle Cardoso Geraldo; Tansini, Aline; Gonçalves, Amanda Costa; Placeres, Marisa Campos Polesi; Carlos, Iracilda Zeppone

    2013-08-01

    Sporotrichosis is a subcutaneous mycosis that is caused by the dimorphic fungus Sporothrix schenckii. This disease generally occurs within the skin and subcutaneous tissues, causing lesions that can spread through adjacent lymphatic vessels and sometimes leading to systemic diseases in immunocompromised patients. Macrophages are crucial for proper immune responses against a variety of pathogens. Furthermore, macrophages can play different roles in response to different microorganisms and forms of activation, and they can be divided into "classic" or "alternatively" activated populations, as also known as M1 and M2 macrophages. M1 cells can lead to tissue injury and contribute to pathogenesis, whereas M2 cells promote angiogenesis, tissue remodeling, and repair. The aim of this study was to investigate the roles of M1 and M2 macrophages in a sporotrichosis model. Toward this end, we performed phenotyping of peritoneal exudate cells and evaluated the concomitant production of several immunomediators, including IL-12, IL-10, TGF-β, nitric oxide, and arginase-I activity, which were stimulated ex vivo with cell wall peptide-polysaccharide. Our results showed the predominance of the M2 macrophage population, indicated by peaks of arginase-I activity as well as IL-10 and TGF-β production during the 6th and 8th weeks after infection. These results were consistent with cellular phenotyping that revealed increases in CD206-positive cells over this period. This is the first report of the participation of M2 macrophages in sporotrichosis infections.

  8. Conformational variability of the glycine receptor M2 domain in response to activation by different agonists.

    PubMed

    Pless, Stephan A; Dibas, Mohammed I; Lester, Henry A; Lynch, Joseph W

    2007-12-07

    Models describing the structural changes mediating Cys loop receptor activation generally give little attention to the possibility that different agonists may promote activation via distinct M2 pore-lining domain structural rearrangements. We investigated this question by comparing the effects of different ligands on the conformation of the external portion of the homomeric alpha1 glycine receptor M2 domain. Conformational flexibility was assessed by tethering a rhodamine fluorophore to cysteines introduced at the 19' or 22' positions and monitoring fluorescence and current changes during channel activation. During glycine activation, fluorescence of the label attached to R19'C increased by approximately 20%, and the emission peak shifted to lower wavelengths, consistent with a more hydrophobic fluorophore environment. In contrast, ivermectin activated the receptors without producing a fluorescence change. Although taurine and beta-alanine were weak partial agonists at the alpha1R19'C glycine receptor, they induced large fluorescence changes. Propofol, which drastically enhanced these currents, did not induce a glycine-like blue shift in the spectral emission peak. The inhibitors strychnine and picrotoxin elicited fluorescence and current changes as expected for a competitive antagonist and an open channel blocker, respectively. Glycine and taurine (or beta-alanine) also produced an increase and a decrease, respectively, in the fluorescence of a label attached to the nearby L22'C residue. Thus, results from two separate labeled residues support the conclusion that the glycine receptor M2 domain responds with distinct conformational changes to activation by different agonists.

  9. THE ENHANCEMENT OF MACROPHAGE BACTERIOSTASIS BY PRODUCTS OF ACTIVATED LYMPHOCYTES

    PubMed Central

    Fowles, Robert E.; Fajardo, Ileana M.; Leibowitch, Jacques L.; David, John R.

    1973-01-01

    It was reported previously that the incubation of normal guinea pig macrophages with partially purified products of activated lymphocytes resulted in altered macrophage function including increased cell adherence to culture vessels, spreading, phagocytosis, and glucose carbon-1 oxidation. Studies reported here demonstrate that such macrophages also exhibit enhanced bacteriostasis. Lymphocytes were stimulated with concanavalin A, the culture supernatant was chromatographed over Sephadex G-100 and the fraction of mol wt 25,000–55,000, rich in lymphocyte mediators, was cultured with normal guinea pig macrophages for 1–3 days. Macrophages incubated with fractions from unstimulated lymphocyte cultures served as controls. The resulting macrophage monolayers were infected with Listeria monocytogenes. Macrophages incubated with mediator-rich fractions exhibited 2- to 10-fold enhanced bacteriostasis compared to controls. Further studies indicate that this enhancement was attributable to intrinsic changes in the macrophages and not simply a consequence of the number of macrophages on the monolayers. The studies support the concept that macrophage bacteriostasis can be enhanced by lymphocyte mediators. However, macrophages, which have been preincubated directly with sensitive lymphocytes and antigen exhibit even greater bacteriostasis and sometimes bactericidal capacity, suggesting that either a labile lymphocyte factor or direct lymphocyte macrophage interaction may also be involved in bactericidal activity. PMID:4200649

  10. 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

  11. D-penicillamine-induced autoimmunity: relationship to macrophage activation.

    PubMed

    Li, Jinze; Uetrecht, Jack P

    2009-09-01

    Idiosyncratic drug reactions represent a serious health problem, and they remain unpredictable largely due to our limited understanding of the mechanisms involved. Penicillamine-induced autoimmunity in Brown Norway (BN) rats represents one model of an idiosyncratic reaction, and this drug can also cause autoimmune reactions in humans. We previously demonstrated that penicillamine binds to aldehydes on the surface of macrophages. There is evidence that an imine bond formed by aldehyde groups on macrophages and amine groups on T cells is one type of interaction between these two cells that is involved in the induction of an immune response. We proposed that the binding of penicillamine with aldehyde groups on macrophages could lead to their activation and in some patients could lead to autoimmunity. In this study, the transcriptome profile of spleen macrophages 6 h after penicillamine treatment was used to detect effects of penicillamine on macrophages with a focus on 20 genes known to be macrophage activation biomarkers. One biological consequence of macrophage activation was investigated by determining mRNA levels for IL-15 and IL-1 beta which are crucial for NK cell activation, as well as levels of mRNA for selected cytokines in spleen NK cells. Up-regulation of the macrophage activating cytokines, IFN-gamma and GM-CSF, and down-regulation of IL-13 indicated activation of NK cells, which suggests a positive feedback loop between macrophages and NK cells. Furthermore, treatment of a murine macrophage cell line, RAW264.7, with penicillamine increased the production of TNF-alpha, IL-6, and IL-23, providing additional evidence that penicillamine activates macrophages. Hydralazine and isoniazid cause a lupus-like syndrome in humans and also bind to aldehyde groups. These drugs were also found to activate RAW264.7 macrophages. Together, these data support the hypothesis that drugs that bind irreversibly with aldehydes lead to macrophage activation, which in some

  12. Hofbauer cells of M2a, M2b and M2c polarization may regulate feto-placental angiogenesis.

    PubMed

    Loegl, J; Hiden, U; Nussbaumer, E; Schliefsteiner, C; Cvitic, S; Lang, I; Wadsack, C; Huppertz, B; Desoye, G

    2016-11-01

    The human placenta comprises a special type of tissue macrophages, the Hofbauer cells (HBC), which exhibit M2 macrophage phenotype. Several subtypes of M2-polarized macrophages (M2a, M2b and M2c) exist in almost all tissues. Macrophage polarization depends on the way of macrophage activation and leads to the expression of specific cell surface markers and the acquisition of specific functions, including tissue remodeling and the promotion of angiogenesis. The placenta is a highly vascularized and rapidly growing organ, suggesting a role of HBC in feto-placental angiogenesis. We here aimed to characterize the specific polarization and phenotype of HBC and investigated the role of HBC in feto-placental angiogenesis. Therefore, HBC were isolated from third trimester placentas and their phenotype was determined by the presence of cell surface markers (FACS analysis) and secretion of cytokines (ELISA). HBC conditioned medium (CM) was analyzed for pro-angiogenic factors, and the effect of HBC CM on angiogenesis, proliferation and chemoattraction of isolated primary feto-placental endothelial cells (fpEC) was determined in vitro Our results revealed that isolated HBC possess an M2 polarization, with M2a, M2b and M2c characteristics. HBC secreted the pro-angiogenic molecules VEGF and FGF2. Furthermore, HBC CM stimulated the in vitro angiogenesis of fpEC. However, compared with control medium, chemoattraction of fpEC toward HBC CM was reduced. Proliferation of fpEC was not affected by HBC CM. These findings demonstrate a paracrine regulation of feto-placental angiogenesis by HBC in vitro Based on our collective results, we propose that the changes in HBC number or phenotype may affect feto-placental angiogenesis. © 2016 Society for Reproduction and Fertility.

  13. Functional modifications of macrophage activity after sublethal irradiation. [Toxoplasma gondii

    SciTech Connect

    Swartz, R.P.

    1982-01-01

    The modifications of macrophage activity following sublethal irradiation, both in vivo and in vitro, were studied using spreading and C3b-receptor-mediated ingestion assays. Nonelicited peritoneal washout cells were examined for changes in activity and selected population characteristics. The cells from irradiated mice were from a resident peritoneal population and not immigrating cells. The macrophage population showed enhanced activity early with a refractory period (24-48) when the macrophages were unresponsive to stimulation by irradiated lymphocytes. The enhanced activity was inversely dose dependent on macrophage. The lymphocytes showed a regulatory function(s) on the time post irradiation at which they were examined. Early lymphocytes exhibited the ability to enhance the activity of normal macrophages while lymphocytes removed 24 hours post irradiation could suppress the activity of already activated macrophages. The effect(s) of the various lymphocyte populations were reproduced with cell-free supernatants which was indicative of the production of lymphokines. Separation on nylon wool columns indicated that the activity resided primarily in the T-cell population of lymphocytes. In vitro irradiation indicated that stimulation of the lymphocytes is macrophage dependent. Additional work indicated that sublethally irradiated macrophages did not inhibit replication of the coccidian protozoon Toxoplasma gondii although they did show increased phagocytosis. Examination of the serum from whole body irradiated mice showed the presence of a postirradiation substance which enhanced the phagocytosis of normal macrophages. It was not present in the serum of normal mice and was not endotoxin.

  14. The Macrophage Switch in Obesity Development

    PubMed Central

    Castoldi, Angela; Naffah de Souza, Cristiane; Câmara, Niels Olsen Saraiva; Moraes-Vieira, Pedro M.

    2016-01-01

    Immune cell infiltration in (white) adipose tissue (AT) during obesity is associated with the development of insulin resistance. In AT, the main population of leukocytes are macrophages. Macrophages can be classified into two major populations: M1, classically activated macrophages, and M2, alternatively activated macrophages, although recent studies have identified a broad range of macrophage subsets. During obesity, AT M1 macrophage numbers increase and correlate with AT inflammation and insulin resistance. Upon activation, pro-inflammatory M1 macrophages induce aerobic glycolysis. By contrast, in lean humans and mice, the number of M2 macrophages predominates. M2 macrophages secrete anti-inflammatory cytokines and utilize oxidative metabolism to maintain AT homeostasis. Here, we review the immunologic and metabolic functions of AT macrophages and their different facets in obesity and the metabolic syndrome. PMID:26779183

  15. Glioma sensitive or chemoresistant to temozolomide differentially modulate macrophage protumor activities.

    PubMed

    Azambuja, Juliana H; da Silveira, Elita F; de Carvalho, Taíse R; Oliveira, Pathise S; Pacheco, Simone; do Couto, Carlus T; Beira, Fátima T; Stefanello, Francieli M; Spanevello, Rosélia M; Braganhol, Elizandra

    2017-11-01

    Glioblastomas are the most devastating brain tumor characterized by chemoresistance development and poor prognosis. Macrophages are a component of tumor microenvironment related to glioma malignancy. The relation among inflammation, innate immunity and cancer is accepted; however, molecular and cellular mechanisms mediating this relation and chemoresistance remain unresolved. Here we evaluated whether glioma sensitive or resistant to temozolomide (TMZ) modulate macrophage polarization and inflammatory pathways associated. The impact of glioma-macrophage crosstalk on glioma proliferation was also investigated. GL261 glioma chemoresistance was developed by exposing cells to increasing TMZ concentrations over a period of 6months. Mouse peritoneal macrophages were exposed to glioma-conditioned medium or co-cultured directly with glioma sensitive (GL) or chemoresistant (GLTMZ). Macrophage polarization, in vitro and in vivo glioma proliferation, redox parameters, ectonucleotidase activity and ATP cytotoxicity were performed. GLTMZ cells were more effective than GL in induce M2-like macrophage polarization and in promote a strong immunosuppressive environment characterized by high IL-10 release and increased antioxidant potential, which may contribute to glioma chemoresistance and proliferation. Interestingly, macrophage-GLTMZ crosstalk enhanced in vitro and in vivo proliferation of chemoresistant cells, decreased ectonucleotidase activities, which was followed by increased macrophage sensitivity to ATP induced death. Results suggest a differential macrophage modulation by GLTMZ cells, which may favor the maintenance of immunosuppressive tumor microenvironment and glioma proliferation. The induction of immunosuppressive environment and macrophage education by chemoresistant gliomas may be important for tumor recovery after chemotherapy and could be considered to overcome chemoresistance development. Copyright © 2017. Published by Elsevier B.V.

  16. Inflammatory Stroke Extracellular Vesicles Induce Macrophage Activation.

    PubMed

    Couch, Yvonne; Akbar, Naveed; Davis, Simon; Fischer, Roman; Dickens, Alex M; Neuhaus, Ain A; Burgess, Annette I; Rothwell, Peter M; Buchan, Alastair M

    2017-08-01

    Extracellular vesicles (EVs) are protein-lipid complexes released from cells, as well as actively exocytosed, as part of normal physiology, but also during pathological processes such as those occurring during a stroke. Our aim was to determine the inflammatory potential of stroke EVs. EVs were quantified and analyzed in the sera of patients after an acute stroke (<24 hours; OXVASC [Oxford Vascular Study]). Isolated EV fractions were subjected to untargeted proteomic analysis by liquid chromatography mass-spectrometry/mass-spectrometry and then applied to macrophages in culture to investigate inflammatory gene expression. EV number, but not size, is significantly increased in stroke patients when compared to age-matched controls. Proteomic analysis reveals an overall increase in acute phase proteins, including C-reactive protein. EV fractions applied to monocyte-differentiated macrophage cultures induced inflammatory gene expression. Together these data show that EVs from stroke patients are proinflammatory in nature and are capable of inducing inflammation in immune cells. © 2017 American Heart Association, Inc.

  17. Immunomodulation by Blastomyces dermatitidis: functional activity of murine peritoneal macrophages.

    PubMed Central

    McDaniel, L S; Cozad, G C

    1983-01-01

    Cell-mediated immunity plays the dominant role in the immune response of mice to Blastomyces dermatitidis infections. Since macrophages play an important role in cell-mediated immunity, the interactions between sensitized murine peritoneal macrophages and the yeast phase of B. dermatitidis were investigated. Scanning electron microscopy showed that the sensitized macrophages readily phagocytized B. dermatitidis yeast cells. In addition, there appeared to be activation of metabolic pathways within the sensitized macrophages, as indicated by increased chemiluminescence activity during phagocytosis. Sensitized macrophages were significantly better at controlling intracellular proliferation of the yeast cells when compared to nonsensitized cells. This was determined by disruption of macrophages and plating for viable yeasts. Scanning electron microscope observations offered further substantiation. Experiments with Candida albicans indicated that B. dermatitidis non-specifically activated macrophages. At 2 h postphagocytosis, 30% fewer C. albicans in B. dermatitidis-activated macrophages were able to form germ tubes. These studies demonstrated the multiple potential of activated macrophages with regard to their functional activity. Images PMID:6840859

  18. Telmisartan prevention of LPS-induced microglia activation involves M2 microglia polarization via CaMKKβ-dependent AMPK activation.

    PubMed

    Xu, Yuan; Xu, Yazhou; Wang, Yurong; Wang, Yunjie; He, Ling; Jiang, Zhenzhou; Huang, Zhangjian; Liao, Hong; Li, Jia; Saavedra, Juan M; Zhang, Luyong; Pang, Tao

    2015-11-01

    Brain inflammation plays an important role in the pathophysiology of many psychiatric and neurological diseases. During brain inflammation, microglia cells are activated, producing neurotoxic molecules and neurotrophic factors depending on their pro-inflammatory M1 and anti-inflammatory M2 phenotypes. It has been demonstrated that Angiotensin II type 1 receptor blockers (ARBs) ameliorate brain inflammation and reduce M1 microglia activation. The ARB telmisartan suppresses glutamate-induced upregulation of inflammatory genes in cultured primary neurons. We wished to clarify whether telmisartan, in addition, prevents microglia activation through polarization to an anti-inflammatory M2 phenotype. We found that telmisartan promoted M2 polarization and reduced M1 polarization in LPS-stimulated BV2 and primary microglia cells, effects partially dependent on PPARγ activation. The promoting effects of telmisartan on M2 polarization, were attenuated by an AMP-activated protein kinase (AMPK) inhibitor or AMPK knockdown, indicating that AMPK activation participates on telmisartan effects. Moreover, in LPS-stimulated BV2 cells, telmisartan enhancement of M2 gene expression was prevented by the inhibitor STO-609 and siRNA of calmodulin-dependent protein kinase kinase β (CaMKKβ), an upstream kinase of AMPK. Furthermore, telmisartan enhanced brain AMPK activation and M2 gene expression in a mouse model of LPS-induced neuroinflammation. In addition, telmisartan reduced the LPS-induced sickness behavior in this in vivo model, and this effect was prevented by prior administration of an AMPK inhibitor. Our results indicate that telmisartan can be considered as a novel AMPK activator, suppressing microglia activation by promoting M2 polarization. Telmisartan may provide a novel, safe therapeutic approach to treat brain disorders associated with enhanced inflammation. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Cyclophilin A Aggravates Collagen-Induced Arthritis via Promoting Classically Activated Macrophages.

    PubMed

    Dongsheng, Zhai; Zhiguang, Fu; Junfeng, Jia; Zifan, Lu; Li, Wang

    2017-07-29

    Activated macrophages exhibiting diverse phenotypes and various functions contribute to the pathogenesis or amelioration of different diseases like cancer, inflammation, and infectious and autoimmune diseases. However, the mechanisms of macrophage polarization in inflamed joint and its effects on rheumatoid arthritis (RA) are still not clarified. This study is designed to explore the effects of cyclophilin A (CypA) on macrophage polarization and describe the underlying mechanisms. Collagen-induced arthritis (CIA) was employed to address the pro-arthritic effects of CypA. Flow cytometry was performed to investigate the populations of M1 and M2 macrophages in synovial tissues of the mice. Knockdown or overexpression of CypA macrophage cells was used to study the functions of CypA on macrophage polarization. Western blot was carried out to examine the potential signaling pathways. We found that CypA aggravated the severity of CIA in mice, as assessed by the increase of clinical score of inflammation, cartilage damage, and bone erosion. Moreover, the level of cytokines, such as IL-6, IL-1β, and IL-17, and the number of pro-inflammatory macrophages in synovial fluid were significantly elevated. In accordance with our observation, CypA dysregulation could actually influence the M1 macrophages polarization and pro-inflammatory cytokines production. Further mechanism study disclosed that CypA could regulate the transcriptional activity of NF-κB, the pivotal transcriptional factor regulating M1 polarization, dependent of its PPIase activity. Our findings provide evidence that PPIase CypA promoted macrophages polarization toward pro-inflammatory M1 phenotype via transcriptional activating NF-κB, thus leading to aggravated arthritis.

  20. Titanium particles that have undergone phagocytosis by macrophages lose the ability to activate other macrophages.

    PubMed

    Xing, Zhiqing; Schwab, Luciana P; Alley, Carie F; Hasty, Karen A; Smith, Richard A

    2008-04-01

    Titanium particles derived from the wear of the orthopaedic implant surfaces can activate macrophages to secrete cytokines and stimulate osteoclastic bone resorption, causing osteolysis around orthopaedic implants. However, what happens to the titanium particles after being phagocytosed by macrophages is not known. We prepared titanium particles (as received, clean, and LPS-coated), and exposed them to macrophages in culture. Free particles were washed away after 24 h and the intracellular particles were kept in culture for additional 48 h until being harvested by lysing the cells. Particles that had been cell treated or noncell treated were examined by scanning electronic microscopy to analyze the shape, size, and concentration of the particles. The cell treated and noncell treated particles were exposed to macrophages in culture with a particle to cell ratio of 300:1. After 18 h, the levels of TNF-alpha in culture medium and the viability of the cells were examined. Clean particles did not stimulate TNF-alpha secretion by macrophages, while LPS-coated particles dramatically increased that response. Phagocytosis by macrophages did not change the shape and size of the particles, but depleted the ability of the particles to stimulate TNF-alpha secretion by macrophages. This indicates that macrophages are capable of rendering titanium particles inactive without degrading the particles, possibly by altering the surface chemistry of the particles.

  1. Contribution of alternatively activated macrophages to allergic lung inflammation: a tale of mice and men.

    PubMed

    Dasgupta, Preeta; Keegan, Achsah D

    2012-01-01

    The concept that macrophages play an active role in inflammatory responses began its development in the late 1800s with the now iconic studies by Elie Metchnikoff using starfish larvae and Daphnia [reviewed in Kaufmann SHE: Nat Immunol 2008;9:705-712 and Cavaillon JM: J Leukoc Biol 2011;90:413-424]. Based on his observation of the phagocyte response to a foreign body (rose thorn) and yeast, he proposed that phagocytes acted in host defense and were active participants in the inflammatory process. Flash forward more than 100 years and we find that these basic tenets hold true. However, it is now appreciated that macrophages come in many different flavors and can adopt a variety of nuanced phenotypes depending on the tissue environment in which the macrophage is found. In this brief review, we discuss the role of one type of macrophage termed the alternatively activated macrophage (AAM), also known as the M2 type of macrophage, in regulating allergic lung inflammation and asthma. Recent studies using mouse models of allergic lung inflammation and samples from human asthma patients contribute to the emerging concept that AAMs are not just bystanders of the interleukin (IL)-4- and IL-13-rich environment found in allergic asthma but are also active players in orchestrating allergic lung disease.

  2. Complement expression in retinal pigment epithelial cells is modulated by activated macrophages.

    PubMed

    Luo, Chang; Zhao, Jiawu; Madden, Angelina; Chen, Mei; Xu, Heping

    2013-07-01

    Complement activation is involved in a variety of retinal diseases. We have shown previously that a number of complement components and regulators can be produced locally in the eye, and that retinal pigment epithelial (RPE) cells are the major source of complement expression at the retina-choroidal interface. The expression of complement components by RPE cells is regulated by inflammatory cytokines. Under aging or inflammatory conditions, microglia and macrophages accumulate in the subretinal space, where they are in close contact with RPE cells. In this study, we investigated the effect of activated macrophages on complement expression by RPE cells. Mouse RPE cells were treated with the supernatants from un-activated bone marrow-derived macrophages (BM-DMs), the classically activated BM-DMs (M1) and different types of the alternatively activated BM-DMs (M2a by IL-4, M2b by immune complex and lipopolysaccharide (LPS), M2c by IL-10). The expression of inflammatory cytokines and complement genes by RPE cells were determined by real-time RT-PCR. The protein expression of CFB, C3, C1INH, and C1r was examined by Western blot. Our results show that un-stimulated RPE cells express a variety of complement-related genes, and that the expression levels of complement regulators, including C1r, factor H (CFH), DAF1, CD59, C1INH, Crry, and C4BP genes are significantly higher than those of complement component genes (C2, C4, CFB, C3, and C5). Macrophage supernatants increased inflammatory cytokine (IL-1β, IL-6, iNOS), chemokine (CCL2) and complement expression in RPE cells. The supernatants from M0, M2a and M2c macrophages mildly up-regulated (2-3.5-fold) CFB, CFH and C3 gene expression in RPE cells, whereas the supernatants from M1 and M2b macrophages massively increased (10-30-fold) CFB and C3 gene expression in RPE cells. The expression of other genes, including C1r, C2, C4, CFH, Masp1, C1INH, and C4BP in RPE cells was also increased by the supernatants of M1 and M2b

  3. Collagenase Production by Endotoxin-Activated Macrophages

    PubMed Central

    Wahl, Larry M.; Wahl, Sharon M.; Mergenhagen, Stephan E.; Martin, George R.

    1974-01-01

    Peritoneal exudate macrophages, when exposed to bacterial lipopolysaccharide in culture, were found to produce collagenase (EC 3.4.24.3). This enzyme was not detected in extracts of the macrophages or in media from nonstimulated macrophage cultures. Lipidcontaining fractions of the lipopolysaccharide, including a glycolipid from the rough mutant of Salmonella minnesota (R595) and lipid A, were potent stimulators of collagenase production. The lipid-free polysaccharide fraction had no effect. Cycloheximide prevented the production of collagenase by endotoxin-treated macrophages, suggesting that it was newly synthesized. Images PMID:4372628

  4. Exopolysaccharide from Trichoderma pseudokoningii induces macrophage activation.

    PubMed

    Wang, Guodong; Zhu, Lei; Yu, Bo; Chen, Ke; Liu, Bo; Liu, Jun; Qin, Guozheng; Liu, Chunyan; Liu, Huixia; Chen, Kaoshan

    2016-09-20

    In this study, we evaluated the immunomodulatory activity of an exopolysaccharide (EPS) derived from Trichoderma pseudokoningii and investigated the molecular mechanism of EPS-mediated activation of macrophages. Results revealed that EPS could significantly induce the production of nitric oxide (NO), tumor necrosis factor (TNF)-α and interleukin (IL)-1β and enhance phagocytic activity in RAW 264.7 cells. Immunofluorescence staining indicated that EPS promoted the nuclear translocation of nuclear factor (NF)-κB p65 subunit. Western blot analysis showed that EPS increased the expression of inducible nitric oxide synthase (iNOS) protein, the degradation of IκB-α and the phosphorylation of mitogen-activated protein kinases (MAPKs). Furthermore, pretreatment of RAW 264.7 cells with specific inhibitors of NF-κB and MAPKs significantly attenuated EPS-induced TNF-α and IL-1β production. EPS also induced the inhibition of cytokine secretion by special antibodies against Toll-like receptor-4 (TLR4) and Dectin-1. These data suggest that EPS from Trichoderma pseudokoningii activates RAW 264.7 cells through NF-κB and MAPKs signaling pathways via TLR4 and Dectin-1.

  5. Cellular and Molecular Mechanisms Underpinning Macrophage Activation during Remyelination

    PubMed Central

    Lloyd, Amy F.; Miron, Veronique E.

    2016-01-01

    Remyelination is an example of central nervous system (CNS) regeneration, whereby myelin is restored around demyelinated axons, re-establishing saltatory conduction and trophic/metabolic support. In progressive multiple sclerosis, remyelination is limited or fails altogether which is considered to contribute to axonal damage/loss and consequent disability. Macrophages have critical roles in both CNS damage and regeneration, such as remyelination. This diverse range in functions reflects the ability of macrophages to acquire tissue microenvironment-specific activation states. This activation is dynamically regulated during efficient regeneration, with a switch from pro-inflammatory to inflammation-resolution/pro-regenerative phenotypes. Although, some molecules and pathways have been implicated in the dynamic activation of macrophages, such as NFκB, the cellular and molecular mechanisms underpinning plasticity of macrophage activation are unclear. Identifying mechanisms regulating macrophage activation to pro-regenerative phenotypes may lead to novel therapeutic strategies to promote remyelination in multiple sclerosis. PMID:27446913

  6. Whole-cell MALDI-TOF MS: a new tool to assess the multifaceted activation of macrophages.

    PubMed

    Ouedraogo, Richard; Daumas, Aurélie; Ghigo, Eric; Capo, Christian; Mege, Jean-Louis; Textoris, Julien

    2012-10-22

    Whole-cell MALDI-TOF MS is routinely used to identify bacterial species in clinical samples. This technique has also proven to allow identification of intact mammalian cells, including macrophages. Here, we wondered whether this approach enabled the assessment human macrophages plasticity. The whole-cell MALDI-TOF spectra of macrophages stimulated with IFN-γ and IL-4, two inducers of M1 and M2 macrophage polarisation, consisted of peaks ranging from 2 to 12 kDa. The spectra of unstimulated and stimulated macrophages were clearly different. The fingerprints induced by the M1 agonists, IFN-γ, TNF, LPS and LPS+IFN-γ, and the M2 agonists, IL-4, TGF-β1 and IL-10, were specific and readily identifiable. Thus, whole-cell MALDI-TOF MS was able to characterise M1 and M2 macrophage subtypes. In addition, the fingerprints induced by extracellular (group B Streptococcus, Staphylococcus aureus) or intracellular (BCG, Orientia tsutsugamushi, Coxiella burnetii) bacteria were bacterium-specific. The whole-cell MALDI-TOF MS fingerprints therefore revealed the multifaceted activation of human macrophages. This approach opened a new avenue of studies to assess the immune response in the clinical setting, by monitoring the various activation patterns of immune cells in pathological conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis

    SciTech Connect

    Anastasiou, Dimitrios; Yu, Yimin; Israelsen, William J.; Jiang, Jian-Kang; Boxer, Matthew B.; Hong, Bum Soo; Tempel, Wolfram; Dimov, Svetoslav; Shen, Min; Jha, Abhishek; Yang, Hua; Mattaini, Katherine R.; Metallo, Christian M.; Fiske, Brian P.; Courtney, Kevin D.; Malstrom, Scott; Khan, Tahsin M.; Kung, Charles; Skoumbourdis, Amanda P.; Veith, Henrike; Southall, Noel; Walsh, Martin J.; Brimacombe, Kyle R.; Leister, William; Lunt, Sophia Y.; Johnson, Zachary R.; Yen, Katharine E.; Kunii, Kaiko; Davidson, Shawn M.; Christofk, Heather R.; Austin, Christopher P.; Inglese, James; Harris, Marian H.; Asara, John M.; Stephanopoulos, Gregory; Salituro, Francesco G.; Jin, Shengfang; Dang, Lenny; Auld, Douglas S.; Park, Hee-Won; Cantley, Lewis C.; Thomas, Craig J.; Vander Heiden, Matthew G.

    2012-08-26

    Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. The interaction of PKM2 with phosphotyrosine-containing proteins inhibits enzyme activity and increases the availability of glycolytic metabolites to support cell proliferation. This suggests that high pyruvate kinase activity may suppress tumor growth. We show that expression of PKM1, the pyruvate kinase isoform with high constitutive activity, or exposure to published small-molecule PKM2 activators inhibits the growth of xenograft tumors. Structural studies reveal that small-molecule activators bind PKM2 at the subunit interaction interface, a site that is distinct from that of the endogenous activator fructose-1,6-bisphosphate (FBP). However, unlike FBP, binding of activators to PKM2 promotes a constitutively active enzyme state that is resistant to inhibition by tyrosine-phosphorylated proteins. This data supports the notion that small-molecule activation of PKM2 can interfere with anabolic metabolism.

  8. Pyruvate kinase M2 activators promote tetramer formation and suppress tumorigenesis

    PubMed Central

    Anastasiou, Dimitrios; Yu, Yimin; Israelsen, William J.; Jiang, Jian-kang; Boxer, Matthew B.; Hong, Bum Soo; Tempel, Wolfram; Dimov, Svetoslav; Shen, Min; Jha, Abhishek; Yang, Hua; Mattaini, Katherine R.; Metallo, Christian M.; Fiske, Brian P.; Courtney, Kevin D.; Malstrom, Scott; Khan, Tahsin M.; Kung, Charles; Skoumbourdis, Amanda P.; Veith, Henrike; Southall, Noel; Walsh, Martin J.; Brimacombe, Kyle R.; Leister, William; Lunt, Sophia Y.; Johnson, Zachary R.; Yen, Katharine E.; Kunii, Kaiko; Davidson, Shawn M.; Christofk, Heather R.; Austin, Christopher P.; Inglese, James; Harris, Marian H.; Asara, John M.; Stephanopoulos, Gregory; Salituro, Francesco G.; Jin, Shengfang; Dang, Lenny; Auld, Douglas S.; Park, Hee-Won; Cantley, Lewis C.; Thomas, Craig J.; Vander Heiden, Matthew G.

    2012-01-01

    Cancer cells engage in a metabolic program to enhance biosynthesis and support cell proliferation. The regulatory properties of pyruvate kinase M2 (PKM2) influence altered glucose metabolism in cancer. PKM2 interaction with phosphotyrosine-containing proteins inhibits enzyme activity and increases availability of glycolytic metabolites to support cell proliferation. This suggests that high pyruvate kinase activity may suppress tumor growth. We show that expression of PKM1, the pyruvate kinase isoform with high constitutive activity, or exposure to published small molecule PKM2 activators inhibit growth of xenograft tumors. Structural studies reveal that small molecule activators bind PKM2 at the subunit interaction interface, a site distinct from that of the endogenous activator fructose-1,6-bisphosphate (FBP). However, unlike FBP, binding of activators to PKM2 promotes a constitutively active enzyme state that is resistant to inhibition by tyrosine-phosphorylated proteins. These data support the notion that small molecule activation of PKM2 can interfere with anabolic metabolism. PMID:22922757

  9. Trichoderma asperelloides Spores Downregulate dectin1/2 and TLR2 Receptors of Mice Macrophages and Decrease Candida parapsilosis Phagocytosis Independent of the M1/M2 Polarization.

    PubMed

    Dos Santos, Andréa G; Mendes, Érica A; de Oliveira, Rafael P; Faria, Ana M C; de Sousa, Aurizangela O; Pirovani, Carlos P; de Araújo, Fernanda F; de Carvalho, Andréa T; Costa, Marliete Carvalho; Assis Santos, Daniel; Montoya, Quimi V; Rodrigues, Andre; Dos Santos, Jane L

    2017-01-01

    The intensive use of pesticides to control pests in agriculture has promoted several issues relating to environment. As chemical pesticides remain controversial, biocontrol agents originating from fungi could be an alternative. Among them, we highlight biocontrol agents derived from the fungi genus Trichoderma, which have been documented in limiting the growth of other phytopathogenic fungus in the roots and leaves of several plant species. An important member of this genus is Trichoderma asperelloides, whose biocontrol agents have been used to promote plant growth while also treating soil diseases caused by microorganisms in both greenhouses and outdoor crops. To evaluate the safety of fungal biological agents for human health, tests to detect potentially adverse effects, such as allergenicity, toxicity, infectivity and pathogenicity, are crucial. In addition, identifying possible immunomodulating properties of fungal biocontrol agents merits further investigation. Thus, the aim of this study was to evaluate the effects of T. asperelloides spores in the internalization of Candida parapsilosis yeast by mice phagocytes, in order to elucidate the cellular and molecular mechanism of this interaction, as a model to understand possible in vivo effects of this fungus. For this, mice were exposed to a fungal spore suspension through-intraperitoneal injection, euthanized and cells from the peripheral blood and peritoneal cavity were collected for functional, quantitative and phenotypic analysis, throughout analysis of membrane receptors gene expression, phagocytosis ability and cells immunophenotyping M1 (CCR7 and CD86) and M2 (CCR2 and CD206). Our analyses showed that phagocytes exposed to fungal spores had reduced phagocytic capacity, as well as a decrease in the quantity of neutrophils and monocytes in the peripheral blood and peritoneal cavity. Moreover, macrophages exposed to T. asperelloides spores did not display the phenotypic profile M1/M2, and had reduced

  10. Haematopoietic prolyl hydroxylase-1 deficiency promotes M2 macrophage polarization and is both necessary and sufficient to protect against experimental colitis.

    PubMed

    Van Welden, Sophie; De Vos, Martine; Wielockx, Ben; Tavernier, Simon J; Dullaers, Melissa; Neyt, Sara; Descamps, Benedicte; Devisscher, Lindsey; Devriese, Sarah; Van den Bossche, Lien; Holvoet, Tom; Baeyens, Ann; Correale, Carmen; D'Alessio, Silvia; Vanhove, Christian; De Vos, Filip; Verhasselt, Bruno; Breier, Georg; Lambrecht, Bart N; Janssens, Sophie; Carmeliet, Peter; Danese, Silvio; Elewaut, Dirk; Laukens, Debby; Hindryckx, Pieter

    2017-03-01

    Prolyl hydroxylase domain-containing proteins (PHDs) regulate the adaptation of cells to hypoxia. Pan-hydroxylase inhibition is protective in experimental colitis, in which PHD1 plays a prominent role. However, it is currently unknown how PHD1 targeting regulates this protection and which cell type(s) are involved. Here, we demonstrated that Phd1 deletion in endothelial and haematopoietic cells (Phd1(f/)(f) Tie2:cre) protected mice from dextran sulphate sodium (DSS)-induced colitis, with reduced epithelial erosions, immune cell infiltration, and colonic microvascular dysfunction, whereas the response of Phd2(f/+) Tie2:cre and Phd3(f/)(f) Tie2:cre mice to DSS was similar to that of their littermate controls. Using bone marrow chimeras and cell-specific cre mice, we demonstrated that ablation of Phd1 in haematopoietic cells but not in endothelial cells was both necessary and sufficient to inhibit experimental colitis. This effect relied, at least in part, on skewing of Phd1-deficient bone marrow-derived macrophages towards an anti-inflammatory M2 phenotype. These cells showed an attenuated nuclear factor-κB-dependent response to lipopolysaccharide (LPS), which in turn diminished endothelial chemokine expression. In addition, Phd1 deficiency in dendritic cells significantly reduced interleukin-1β production in response to LPS. Taken together, our results further support the development of selective PHD1 inhibitors for ulcerative colitis, and identify haematopoietic cells as their primary target. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  11. Advanced Glycation End Products Enhance Macrophages Polarization into M1 Phenotype through Activating RAGE/NF-κB Pathway

    PubMed Central

    Jin, Xian; Yao, Tongqing; Zhou, Zhong'e; Zhu, Jian; Zhang, Song; Hu, Wei; Shen, Chengxing

    2015-01-01

    Atherosclerotic lesions are accelerated in patients with diabetes. M1 (classically activated in contrast to M2 alternatively activated) macrophages play key roles in the progression of atherosclerosis. Since advanced glycation end products (AGEs) are major pathogenic factors and active inflammation inducers in diabetes mellitus, this study assessed the effects of AGEs on macrophage polarization. The present study showed that AGEs significantly promoted macrophages to express IL-6 and TNF-α. M1 macrophage markers such as iNOS and surface markers including CD11c and CD86 were significantly upregulated while M2 macrophage markers such as Arg1 and CD206 remained unchanged after AGEs stimulation. AGEs significantly increased RAGE expression in macrophages and activated NF-κB pathway, and the aforementioned effects were partly abolished by administration of anti-RAGE antibody or NF-κB inhibitor PDTC. In conclusion, our results suggest that AGEs enhance macrophage differentiation into proinflammatory M1 phenotype at least partly via RAGE/NF-κB pathway activation. PMID:26114112

  12. Advanced Glycation End Products Enhance Macrophages Polarization into M1 Phenotype through Activating RAGE/NF-κB Pathway.

    PubMed

    Jin, Xian; Yao, Tongqing; Zhou, Zhong'e; Zhu, Jian; Zhang, Song; Hu, Wei; Shen, Chengxing

    2015-01-01

    Atherosclerotic lesions are accelerated in patients with diabetes. M1 (classically activated in contrast to M2 alternatively activated) macrophages play key roles in the progression of atherosclerosis. Since advanced glycation end products (AGEs) are major pathogenic factors and active inflammation inducers in diabetes mellitus, this study assessed the effects of AGEs on macrophage polarization. The present study showed that AGEs significantly promoted macrophages to express IL-6 and TNF-α. M1 macrophage markers such as iNOS and surface markers including CD11c and CD86 were significantly upregulated while M2 macrophage markers such as Arg1 and CD206 remained unchanged after AGEs stimulation. AGEs significantly increased RAGE expression in macrophages and activated NF-κB pathway, and the aforementioned effects were partly abolished by administration of anti-RAGE antibody or NF-κB inhibitor PDTC. In conclusion, our results suggest that AGEs enhance macrophage differentiation into proinflammatory M1 phenotype at least partly via RAGE/NF-κB pathway activation.

  13. Control of tumor-associated macrophage alternative activation by MIF

    PubMed Central

    Yaddanapudi, Kavitha; Putty, Kalyani; Rendon, Beatriz E.; Lamont, Gwyneth J.; Faughn, Jonathan D.; Satoskar, Abhay; Lasnik, Amanda; Eaton, John W.; Mitchell, Robert A.

    2013-01-01

    Tumor stromal alternatively activated macrophages are important determinants of anti-tumor T lymphocyte responses, intratumoral neovascularization and metastatic dissemination. Our recent efforts to investigate the mechanism of macrophage migration inhibitory factor (MIF) in antagonizing anti-melanoma immune responses reveal that macrophage-derived MIF participates in macrophage alternative activation in melanoma-bearing mice. Both peripheral and tumor-associated macrophages (TAMs) isolated from melanoma bearing MIF-deficient mice display elevated pro-inflammatory cytokine expression and reduced anti-inflammatory, immunosuppressive and pro-angiogenic gene products compared to macrophages from tumor bearing MIF wildtype mice. Moreover, TAMs and myeloid-derived suppressor cells (MDSCs) from MIF-deficient mice exhibit reduced T lymphocyte immunosuppressive activities than do those from their wildtype littermates. Corresponding with reduced tumor immunosuppression and neoangiogenic potential by TAMs, MIF-deficiency confers protection against transplantable subcutaneous melanoma outgrowth and melanoma lung metastatic colonization. Finally, we report for the first time that our previously discovered MIF small molecule antagonist, 4-iodo-6-phenylpyrimidine (4-IPP), recapitulates MIF-deficiency in vitro and in vivo and attenuates tumor polarized macrophage alternative activation, immunosuppression, neoangiogenesis and melanoma tumor outgrowth. These studies describe an important functional contribution by MIF to tumor-associated macrophage alternative activation and provide justification for immunotherapeutic targeting of MIF in melanoma patients. PMID:23390297

  14. Glycyrrhizic Acid Promotes M1 Macrophage Polarization in Murine Bone Marrow-Derived Macrophages Associated with the Activation of JNK and NF-κB

    PubMed Central

    Mao, Yulong; Wang, Baikui; Xu, Xin; Du, Wei; Li, Weifen; Wang, Youming

    2015-01-01

    The roots and rhizomes of Glycyrrhiza species (licorice) have been widely used as natural sweeteners and herbal medicines. The aim of this study is to investigate the effect of glycyrrhizic acid (GA) from licorice on macrophage polarization. Both phenotypic and functional activities of murine bone marrow-derived macrophages (BMDMs) treated by GA were assessed. Our results showed that GA obviously increased the cell surface expression of CD80, CD86, and MHCII molecules. Meanwhile, GA upregulated the expression of CCR7 and the production of TNF-α, IL-12, IL-6, and NO (the markers of classically activated (M1) macrophages), whereas it downregulated the expression of MR, Ym1, and Arg1 (the markers of alternatively activated (M2) macrophage). The functional tests showed that GA dramatically enhanced the uptake of FITC-dextran and E. coli K88 by BMDMs and decreased the intracellular survival of E. coli K88 and S. typhimurium. Moreover, we demonstrated that JNK and NF-κB activation are required for GA-induced NO and M1-related cytokines production, while ERK1/2 pathway exhibits a regulatory effect via induction of IL-10. Together, these findings indicated that GA promoted polarization of M1 macrophages and enhanced its phagocytosis and bactericidal capacity. The results expanded our knowledge about the role of GA in macrophage polarization. PMID:26664149

  15. Glycyrrhizic Acid Promotes M1 Macrophage Polarization in Murine Bone Marrow-Derived Macrophages Associated with the Activation of JNK and NF-κB.

    PubMed

    Mao, Yulong; Wang, Baikui; Xu, Xin; Du, Wei; Li, Weifen; Wang, Youming

    2015-01-01

    The roots and rhizomes of Glycyrrhiza species (licorice) have been widely used as natural sweeteners and herbal medicines. The aim of this study is to investigate the effect of glycyrrhizic acid (GA) from licorice on macrophage polarization. Both phenotypic and functional activities of murine bone marrow-derived macrophages (BMDMs) treated by GA were assessed. Our results showed that GA obviously increased the cell surface expression of CD80, CD86, and MHCII molecules. Meanwhile, GA upregulated the expression of CCR7 and the production of TNF-α, IL-12, IL-6, and NO (the markers of classically activated (M1) macrophages), whereas it downregulated the expression of MR, Ym1, and Arg1 (the markers of alternatively activated (M2) macrophage). The functional tests showed that GA dramatically enhanced the uptake of FITC-dextran and E. coli K88 by BMDMs and decreased the intracellular survival of E. coli K88 and S. typhimurium. Moreover, we demonstrated that JNK and NF-κB activation are required for GA-induced NO and M1-related cytokines production, while ERK1/2 pathway exhibits a regulatory effect via induction of IL-10. Together, these findings indicated that GA promoted polarization of M1 macrophages and enhanced its phagocytosis and bactericidal capacity. The results expanded our knowledge about the role of GA in macrophage polarization.

  16. Loss of MCP-1 alters macrophage polarization and reduces NFκB activation in the foreign body response

    PubMed Central

    Moore, Laura Beth; Sawyer, Andrew J.; Charokopos, Antonios; Skokos, Eleni A.; Kyriakides, Themis R.

    2014-01-01

    Implantation of biomaterials elicits a foreign body response characterized by fusion of macrophages to form foreign body giant cells and fibrotic encapsulation. Studies of macrophage polarization in this response have suggested that alternative (M2) activation is associated with more favorable outcomes. Here we investigated this process in vivo by implanting mixed cellulose ester filters or PDMS disks in the peritoneal cavity of WT and MCP-1 KO mice. We analyzed classical (M1) and alternative (M2) gene expression via Q-PCR, immunohistochemistry, and ELISA in both non-adherent cells isolated by lavage and implant-adherent cells. Our results show that macrophages undergo unique activation that displays features of both M1 and M2 polarization including induction of TNF, which induces the expression and nuclear translocation of p50 and RelA determined by immunofluorescence and western blot. Both processes were compromised in fusion-deficient MCP-1 KO macrophages in vitro and in vivo. Furthermore, inclusion of BAY 11-7028, an inhibitor of NFκB activation, reduced nuclear translocation of RelA and fusion in WT macrophages. Our studies suggest that peritoneal implants elicit a unique macrophage polarization phenotype leading to induction of TNF and activation of the NFκB pathway. PMID:25242651

  17. Loss of monocyte chemoattractant protein-1 alters macrophage polarization and reduces NFκB activation in the foreign body response.

    PubMed

    Moore, Laura Beth; Sawyer, Andrew J; Charokopos, Antonios; Skokos, Eleni A; Kyriakides, Themis R

    2015-01-01

    Implantation of biomaterials elicits a foreign body response characterized by fusion of macrophages to form foreign body giant cells and fibrotic encapsulation. Studies of the macrophage polarization involved in this response have suggested that alternative (M2) activation is associated with more favorable outcomes. Here we investigated this process in vivo by implanting mixed cellulose ester filters or polydimethylsiloxane disks in the peritoneal cavity of wild-type (WT) and monocyte chemoattractant protein-1 (MCP-1) knockout mice. We analyzed classical (M1) and alternative (M2) gene expression via quantitative polymerase chain reaction, immunohistochemistry and enzyme-linked immunosorbent assay in both non-adherent cells isolated by lavage and implant-adherent cells. Our results show that macrophages undergo unique activation that displays features of both M1 and M2 polarization including induction of tumor necrosis factor α (TNF), which induces the expression and nuclear translocation of p50 and RelA determined by immunofluorescence and Western blot. Both processes were compromised in fusion-deficient MCP-1 KO macrophages in vitro and in vivo. Furthermore, inclusion of BAY 11-7028, an inhibitor of NFκB activation, reduced nuclear translocation of RelA and fusion in WT macrophages. Our studies suggest that peritoneal implants elicit a unique macrophage polarization phenotype leading to induction of TNF and activation of the NFκB pathway.

  18. Novel phosphate-activated macrophages prevent ectopic calcification by increasing extracellular ATP and pyrophosphate

    PubMed Central

    Villa-Bellosta, Ricardo; Hamczyk, Magda R.; Andrés, Vicente

    2017-01-01

    Purpose Phosphorus is an essential nutrient involved in many pathobiological processes. Less than 1% of phosphorus is found in extracellular fluids as inorganic phosphate ion (Pi) in solution. High serum Pi level promotes ectopic calcification in many tissues, including blood vessels. Here, we studied the effect of elevated Pi concentration on macrophage polarization and calcification. Macrophages, present in virtually all tissues, play key roles in health and disease and display remarkable plasticity, being able to change their physiology in response to environmental cues. Methods and results High-throughput transcriptomic analysis and functional studies demonstrated that Pi induces unpolarized macrophages to adopt a phenotype closely resembling that of alternatively-activated M2 macrophages, as revealed by arginine hydrolysis and energetic and antioxidant profiles. Pi-induced macrophages showed an anti-calcifying action mediated by increased availability of extracellular ATP and pyrophosphate. Conclusion We conclude that the ability of Pi-activated macrophages to prevent calcium-phosphate deposition is a compensatory mechanism protecting tissues from hyperphosphatemia-induced pathologic calcification. PMID:28362852

  19. Targeting macrophage anti-tumor activity to suppress melanoma progression

    PubMed Central

    Yang, Luhong; Liu, Chengfang; Zhang, Qi; Zhang, Linjing

    2017-01-01

    By phagocytosing cancer cells and their cellular debris, macrophages play a critical role in nonspecific defense (innate immunity) and, as antigen presenters, they help initiate specific defense mechanisms (adaptive immunity). Malignant melanoma is a lethal disease due to its aggressive capacity for metastasis and resistance to therapy. For decades, considerable effort has gone into development of an effective immunotherapy for treatment of metastatic melanoma. In this review, we focus on the anti-tumor activities of macrophages in melanoma and their potential as therapeutic targets in melanoma. Although macrophages can be re-educated through intercellular signaling to promote tumor survival owing to their plasticity, we expect that targeting the anti-tumor activity of macrophages remains a promising strategy for melanoma inhibition. The combination of tumoricidal macrophage activation and other treatments such as surgery, chemotherapy, and radiotherapy, may provide an effective and comprehensive anti-melanoma strategy. PMID:28060744

  20. Sodium-activated macrophages: the salt mine expands.

    PubMed

    Lucca, Liliana E; Hafler, David A

    2015-08-01

    High sodium consumption has been raising interest as a putative environmental factor linking Western lifestyle to the growing epidemic of autoimmune and inflammatory diseases. Now Zhang and colleagues show that high sodium drives macrophage to acquire a new proinflammatory effector phenotype with a distinct signature, paving the path to assess the role of salt-activated macrophages in human disease.

  1. EGFR regulates macrophage activation and function in bacterial infection.

    PubMed

    Hardbower, Dana M; Singh, Kshipra; Asim, Mohammad; Verriere, Thomas G; Olivares-Villagómez, Danyvid; Barry, Daniel P; Allaman, Margaret M; Washington, M Kay; Peek, Richard M; Piazuelo, M Blanca; Wilson, Keith T

    2016-09-01

    EGFR signaling regulates macrophage function, but its role in bacterial infection has not been investigated. Here, we assessed the role of macrophage EGFR signaling during infection with Helicobacter pylori, a bacterial pathogen that causes persistent inflammation and gastric cancer. EGFR was phosphorylated in murine and human macrophages during H. pylori infection. In human gastric tissues, elevated levels of phosphorylated EGFR were observed throughout the histologic cascade from gastritis to carcinoma. Deleting Egfr in myeloid cells attenuated gastritis and increased H. pylori burden in infected mice. EGFR deficiency also led to a global defect in macrophage activation that was associated with decreased cytokine, chemokine, and NO production. We observed similar alterations in macrophage activation and disease phenotype in the Citrobacter rodentium model of murine infectious colitis. Mechanistically, EGFR signaling activated NF-κB and MAPK1/3 pathways to induce cytokine production and macrophage activation. Although deletion of Egfr had no effect on DC function, EGFR-deficient macrophages displayed impaired Th1 and Th17 adaptive immune responses to H. pylori, which contributed to decreased chronic inflammation in infected mice. Together, these results indicate that EGFR signaling is central to macrophage function in response to enteric bacterial pathogens and is a potential therapeutic target for infection-induced inflammation and associated carcinogenesis.

  2. Binding of SycH chaperone to YscM1 and YscM2 activates effector yop expression in Yersinia enterocolitica.

    PubMed

    Cambronne, Eric D; Sorg, Joseph A; Schneewind, Olaf

    2004-02-01

    Yersinia enterocolitica transports YscM1 and YscM2 via the type III pathway, a mechanism that is required for the establishment of bacterial infections. Prior to host cell contact, YscM1 and YscM2 exert posttranscriptional regulation to inhibit expression of effector yop genes, which encode virulence factors that travel the type III pathway into the cytoplasm of macrophages. Relief from repression has been predicted to occur via the type III secretion of YscM1 and YscM2 into the extracellular medium, resulting in the depletion of regulatory molecules from the bacterial cytoplasm. Using digitonin fractionation and fluorescence microscopy of FlAsH-labeled polypeptides in Yersinia-infected cells, we have localized YscM1 and YscM2 within the host cell cytoplasm. Type III injection of YscM1 and YscM2 required the SycH chaperone. Expression of C-terminal fusions of YscM1 and YscM2 to the neomycin phosphotransferase reporter revealed sequences required for regulatory activity and for secretion in the absence of SycH. Coexpression of SycH and glutathione S-transferase (GST)-YscM1 or GST-YscM2, hybrid GST variants that cannot be transported by the type III apparatus, also relieved repression of Yop synthesis. GST-SycH bound to YscM1 and YscM2 and activated effector yop expression without initiation of the bound regulatory molecules into the type III pathway. Further, regulation of yop expression by YscM1, YscM2, and SycH is shown to act independently of factors that regulate secretion, and gel filtration chromotography revealed populations of YscM1 and YscM2 that are not bound to SycH under conditions where Yop synthesis is repressed. Taken together, these results suggest that YscM1- and YscM2-mediated repression may be relieved through binding to the cytoplasmic chaperone SycH prior to their type III injection into host cells.

  3. Binding of SycH Chaperone to YscM1 and YscM2 Activates Effector yop Expression in Yersinia enterocolitica

    PubMed Central

    Cambronne, Eric D.; Sorg, Joseph A.; Schneewind, Olaf

    2004-01-01

    Yersinia enterocolitica transports YscM1 and YscM2 via the type III pathway, a mechanism that is required for the establishment of bacterial infections. Prior to host cell contact, YscM1 and YscM2 exert posttranscriptional regulation to inhibit expression of effector yop genes, which encode virulence factors that travel the type III pathway into the cytoplasm of macrophages. Relief from repression has been predicted to occur via the type III secretion of YscM1 and YscM2 into the extracellular medium, resulting in the depletion of regulatory molecules from the bacterial cytoplasm. Using digitonin fractionation and fluorescence microscopy of FlAsH-labeled polypeptides in Yersinia-infected cells, we have localized YscM1 and YscM2 within the host cell cytoplasm. Type III injection of YscM1 and YscM2 required the SycH chaperone. Expression of C-terminal fusions of YscM1 and YscM2 to the neomycin phosphotransferase reporter revealed sequences required for regulatory activity and for secretion in the absence of SycH. Coexpression of SycH and glutathione S-transferase (GST)-YscM1 or GST-YscM2, hybrid GST variants that cannot be transported by the type III apparatus, also relieved repression of Yop synthesis. GST-SycH bound to YscM1 and YscM2 and activated effector yop expression without initiation of the bound regulatory molecules into the type III pathway. Further, regulation of yop expression by YscM1, YscM2, and SycH is shown to act independently of factors that regulate secretion, and gel filtration chromotography revealed populations of YscM1 and YscM2 that are not bound to SycH under conditions where Yop synthesis is repressed. Taken together, these results suggest that YscM1- and YscM2-mediated repression may be relieved through binding to the cytoplasmic chaperone SycH prior to their type III injection into host cells. PMID:14729710

  4. Macrophage Activation Syndrome in Paediatric Rheumatic Diseases.

    PubMed

    Islam, M I; Talukder, M K; Islam, M M; Laila, K; Rahman, S A

    2017-04-01

    Macrophage activation syndrome (MAS) is a potentially fatal complication of rheumatic disorders, which commonly occurs in systemic juvenile idiopathic arthritis (sJIA).This study was carried out with the aims of describing the clinical features, laboratory findings and outcomes of MAS associated with paediatric rheumatic diseases in the Department of Paediatrics, Bangabandhu Sheikh Mujib Medical University (BSMMU) and compare these results with previous studies on MAS. This retrospective study was conducted in the paediatric rheumatology wing of the Department of Paediatrics, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh. Clinical and laboratory profile of all the diagnosed cases of MAS were analyzed from the medical records from January 2010 to July 2015. Among 10 MAS patients, 6 were female and 4 were male. Seven patients of systemic JIA, two patients of SLE and one patient with Kawasaki Disease developed MAS in their course of primary disease. Mean duration of primary disease prior to development of MAS was 2.9 years and mean age of onset was 9.1 years. High continued fever and new onset hepatosplenomegaly were the hallmark of the clinical presentation. White blood cell count and platelet count came down from the mean of 16.2 to 10.2×10⁹/L and 254 to 90×10⁹/L. Mean erythrocyte sedimentation rate was dropped from 56 to 29 mm/hr. Six patients had abnormal liver enzyme level (ALT) and 5 had evidence of coagulopathy (prolonged prothrombin time and APTT) at the onset of disease. Hyperferritinnemia were found in all the patients. Bone marrow study was done in 5 patients but features of hamophagocytosis were found only in 2 patients. All patients received intravenous steroid and 3 patients who did not respond to steroid received additional cyclosporine. Mortality rate was 30% in this series. Macrophage activation syndrome is a fatal complication of paediatric rheumatic diseases among which s-JIA was predominant. Early diagnosis and

  5. Alternative activation modifies macrophage resistance to Mycobacterium bovis.

    PubMed

    Castillo-Velázquez, Uziel; Aranday-Cortés, Elihú; Gutiérrez-Pabello, José A

    2011-07-05

    The aim of this study was to evaluate the influence of macrophage alternative activation in the intracellular pathogen natural disease resistance phenotype of the host. Macrophage monolayers from resistant (R) (3) or susceptible (S) (3) cattle donors were treated with 10 ng/ml of bovine recombinant IL-4 (rbIL-4), and infected with virulent and avirulent Mycobacterium bovis (MOI 10:1). Bactericidal assays were performed to assess the bacterial phagocytic index and intracellular survival. Total RNA was reverse transcribed and used to analyze the relative changes in gene expression of IL-10, IL-12, IL-18 IL-1β, TNF-α, MCP-1, MCP-2, IL-6, MIP-1, MIP-3, iNOS, ARGII and SLAM by real time PCR. Cell supernatants were collected and nitric oxide and arginase production was assessed. Apoptosis induction was measured by TUNEL. IL-4 treatment increased the phagocytic index in both R and S macrophages; however intracellular survival was augmented mainly in S macrophages. Alternative activation decreased gene expression of pro-inflammatory cytokines, nitric oxide production and DNA fragmentation mainly in R macrophages. On the other hand, arginase production was not different between R and S macrophages. Alternative activation modifies the macrophage response against M. bovis. IL-4 treatment minimized the functional differences that exist between R and S macrophages. Copyright © 2011. Published by Elsevier B.V.

  6. Retinal Pigment Epithelial Cells Suppress Phagolysosome Activation in Macrophages

    PubMed Central

    Wang, Eric; Choe, Yoona; Ng, Tat Fong; Taylor, Andrew W.

    2017-01-01

    Purpose The eye is an immune-privileged microenvironment that has adapted several mechanisms of immune regulation to prevent inflammation. One of these potential mechanisms is retinal pigment epithelial cells (RPE) altering phagocytosis in macrophages. Methods The conditioned media of RPE eyecups from eyes of healthy mice and mice with experimental autoimmune uveitis (EAU) were used to treat primary macrophage phagocytizing pHrodo bacterial bioparticles. In addition, the neuropeptides were depleted from the conditioned media of healthy RPE eyecups and used to treat phagocytizing macrophages. The conditioned media from healthy and EAU RPE eyecups were assayed for IL-6, and IL-6 was added to the healthy conditioned media, and neutralized in the EAU conditioned media. The macrophages were treated with the conditioned media and assayed for fluorescence. The macrophages were imaged, and the fluorescence intensity, relative to active phagolysosomes, was measured. Also, the macrophages were assayed using fluorescent viability dye staining. Results The conditioned media from healthy, but not from EAU RPE eyecups suppressed phagolysosome activation. Depletion of the neuropeptides alpha-melanocyte–stimulating hormone and neuropeptide Y from the healthy RPE eyecup conditioned media resulted in macrophage death. In the EAU RPE eyecup conditioned media was 0.96 ± 0.18 ng/mL of IL-6, and when neutralized the conditioned media suppressed phagolysosome activation. Conclusions The healthy RPE through soluble molecules, including alpha-melanocyte–stimulating hormone and neuropeptide Y, suppresses the activation of the phagolysosome in macrophages. In EAU, the IL-6 produced by the RPE promotes the activation of phagolysosomes in macrophages. These results demonstrate that under healthy conditions, RPE promotes an altered pathway of phagocytized material in macrophages with implications on antigen processing and clearance. PMID:28241314

  7. Cardiac M2 muscarinic cholinoceptor activation by human chagasic autoantibodies: association with bradycardia

    PubMed Central

    Goin, J; Borda, E; Auger, S; Storino, R; Sterin-Borda, L

    1999-01-01

    OBJECTIVE—To assess whether exposure of cardiac muscarinic acetylcholine receptors (mAChR) to activating chagasic antimyocardial immunoglobulins results in bradycardia and other dysautonomic symptoms associated with the regulation of heart rate.
METHODS—Trypanosoma cruzi infected patients with bradycardia and other abnormalities in tests of the autonomic nervous system were studied and compared with normal subjects. Antipeptide antibodies in serum were demonstrated by an enzyme linked immunosorbent assay using a synthetic 24-mer-peptide corresponding antigenically to the second extracellular loop of the human heart M2 mAChR. The functional effect of affinity purified antipeptide IgG from chagasic patients on spontaneous beating frequency and cAMP production of isolated normal rat atria was studied.
RESULTS—There was a strong association between the finding of antipeptide antibodies in chagasic patients and the presence of basal bradycardia and an altered Valsalva manoeuvre (basal bradycardia: χ2 = 37.5, p < 0.00001; Valsalva manoeuvre: χ2 = 70.0, p < 0.00001). The antipeptide autoantibodies also showed agonist activity, decreasing the rate of contraction and cAMP production. The effects on rat atria resembled the effects of the authentic agonist and those of the total polyclonal chagasic IgG, being selectively blunted by atropine and AF-DX 116, and neutralised by the synthetic peptide corresponding in amino acid sequence to the second extracellular loop of the human M2 mAChR.
CONCLUSIONS—There is an association between circulating antipeptide autoantibodies in chagasic patients and the presence of bradycardia and other dysautonomic symptoms. Thus these autoantibodies are a marker of autoimmune cardiac autonomic dysfunction. The results support the hypothesis that autoimmune mechanisms play a role in the pathogenesis of chagasic cardioneuromyopathy.


Keywords: heart rate; bradycardia; autoantibodies; chagasic cardiomyopathy PMID

  8. IgG4 can induce an M2-like phenotype in human monocyte-derived macrophages through FcγRI.

    PubMed

    Swisher, Jennifer F A; Haddad, Devin A; McGrath, Anna G; Boekhoudt, Gunther H; Feldman, Gerald M

    2014-01-01

    Antibodies evoke cellular responses through the binding of their Fc region to Fc receptors, most of which contain immunoreceptor tyrosine-based activation motif domains and are thus considered "activating." However, there is a growing appreciation of these receptors for their ability to deliver an inhibitory signal as well. We previously described one such phenomenon whereby interferon (IFN)γ signaling is inhibited by immune complex signaling through FcγRI. To understand the implications of this in the context of therapeutic antibodies, we assessed individual IgG subclasses to determine their ability to deliver this anti-inflammatory signal in monocyte-derived macrophages. Like IgG1, we found that IgG4 is fully capable of inhibiting IFNγ-mediated events. In addition, F(ab')2 fragments that interfere with FcγRI signaling reversed this effect. For mAbs developed with either an IgG1 or an IgG4 constant region for indications where inflammation is undesirable, further examination of a potential Fc-dependent contribution to their mechanism of action is warranted.

  9. Honokiol attenuates Diet-Induced Nonalcoholic Steatohepatitis by regulating macrophage polarization through activating PPARγ.

    PubMed

    Zhong, Xueqing; Liu, Hailin

    2017-07-02

    Nonalcoholic steatohepatitis may develop into hepatic cirrhosis. This study aimed to investigate whether honokiol could prevent NASH induced by high cholesterol and high fat (CL) diet in mice and the possible mechanism involved. Mice were fed with CL diet for 12 weeks to establish a NASH model, honokiol (0.02% w/w in diet) was added to evaluate its effect on NASH. Murine peritoneal macrophages, RAW264.7 and ANA-1 cells were used to explore the possible mechanisms of honokiol on macrophage polarization. Mice developed NASH after fed with CL diet for 12 weeks. Honokiol supplementation alleviated insulin resistance, hepatic steatosis, inflammation and fibrosis induced by CL diet. Immunohistochemistry showed honokiol induced more M2 macrophages in livers compared with CL diet alone. Honokiol decreased M1 marker genes (TNFα, MCP-1), increased M2 marker genes (YM-1, IL-10, IL-4R, IL-13) expression in mice liver compared with CL diet. Moreover, treatment with honokiol lowered ALT and AST in serum and preserved liver from lipid peroxidation, evidenced by lowered hepatic MDA level. Honokiol has antioxidant function, as honokiol upregulated hepatic GSH and SOD level and downregulated hepatic CYP2E1 protein level. Hepatic PPARγ and its target genes were upregulated by honokiol. Furthermore, honokiol (10 μM) treatment in mouse peritoneal cells, RAW264.7 cells and ANA-1 cells led to M2 macrophage polarization, whereas a PPARγ antagonist, GW9662 abolished this effect of honokiol. Honokiol can attenuate CL diet induced NASH and the mechanism in which possibly is polarizing macrophages to M2 phenotype via PPARγ activation. This article is protected by copyright. All rights reserved.

  10. Toxoplasma gondii Chitinase Induces Macrophage Activation

    PubMed Central

    Almeida, Fausto; Sardinha-Silva, Aline; da Silva, Thiago Aparecido; Pessoni, André Moreira; Pinzan, Camila Figueiredo; Alegre-Maller, Ana Claudia Paiva; Cecílio, Nerry Tatiana; Moretti, Nilmar Silvio; Damásio, André Ricardo Lima; Pedersoli, Wellington Ramos; Mineo, José Roberto; Silva, Roberto Nascimento; Roque-Barreira, Maria Cristina

    2015-01-01

    Toxoplasma gondii is an obligate intracellular protozoan parasite found worldwide that is able to chronically infect almost all vertebrate species, especially birds and mammalians. Chitinases are essential to various biological processes, and some pathogens rely on chitinases for successful parasitization. Here, we purified and characterized a chitinase from T. gondii. The enzyme, provisionally named Tg_chitinase, has a molecular mass of 13.7 kDa and exhibits a Km of 0.34 mM and a Vmax of 2.64. The optimal environmental conditions for enzymatic function were at pH 4.0 and 50°C. Tg_chitinase was immunolocalized in the cytoplasm of highly virulent T. gondii RH strain tachyzoites, mainly at the apical extremity. Tg_chitinase induced macrophage activation as manifested by the production of high levels of pro-inflammatory cytokines, a pathogenic hallmark of T. gondii infection. In conclusion, to our knowledge, we describe for the first time a chitinase of T. gondii tachyzoites and provide evidence that this enzyme might influence the pathogenesis of T. gondii infection. PMID:26659253

  11. Effects of lipopolysaccharide on the catabolic activity of macrophages

    SciTech Connect

    Cluff, C.; Ziegler, H.K.

    1986-03-05

    The ability of macropha