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The activation of immune cells in response to stimuli present in their microenvironment is regulated by their metabolic profile. Unlike the signal transduction events, which overlap to a huge degree in diverse cellular processes, the metabolome of a cell reflects a more precise picture of cell physiology and function. Different factors governing the cellular metabolome include receptor signaling, macro and micronutrients, normoxic and hypoxic conditions, energy needs, and biomass demand. Macrophages have enormous plasticity and can perform diverse functions depending upon their phenotypic state. This review presents recent updates on the cellular metabolome and molecular patterns associated with M1 and M2 macrophages, also termed "classically activated macrophages" and "alternatively activated macrophages," respectively. M1 macrophages are proinflammatory in nature and predominantly Th1-specific immune responses induce their polarization. On the contrary, M2 macrophages are anti-inflammatory in nature and primarily participate in Th2-specific responses. Interestingly, the same macrophage cell can adapt to the M1 or M2 phenotype depending upon the clues from its microenvironment. We elaborate on the various tissue niche-specific factors, which govern macrophage metabolism and heterogeneity. Furthermore, the current review provides an in-depth account of deregulated macrophage metabolism associated with pathological disorders such as cancer, obesity, and atherosclerosis. We further highlight significant differences in various metabolic pathways governing the cellular bioenergetics and their impact on macrophage effector functions and associated disorders.
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Inflamação/patologia , Macrófagos/citologia , Humanos , Inflamação/metabolismo , Macrófagos/metabolismo , Transdução de SinaisRESUMO
Classically activated macrophages (CAMs) play a crucial protective role in the host by killing the invading pathogens. However, excessive activation of CAMs causes chronic inflammation leading to host tissue damage. Thus, control of macrophage activity is necessary to prevent chronic inflammation. To date, regulation of CAMs in the development of chronic inflammatory diseases has not been elucidated. In this study, we investigated the effect of a pan-caspase inhibitor, zVAD-fmk, in cell death in lipopolysaccharide (LPS)-activated macrophages, CAMs. Necrostatin-1, an inhibitor of necroptosis, inhibited zVAD-fmk-induced cell death in CAMs. The expression of mixed lineage kinase domain-like protein (MLKL) involved in the necroptosis pathway was up-regulated by LPS in CAMs. zVAD-fmk enhanced the phosphorylation of MLKL in CAMs. Moreover, inhibition of activation of mitogen activated protein kinase p38 and generation of reactive oxygen species (ROS) reduced zVAD-fmk-induced cell death in CAMs. Inhibition of ROS generation decreased the activation of MLKL and p38 in zVAD-fmk-treated CAMs. These results, taken together, indicate that zVAD-fmk-induced cell death occurred by necroptosis through ROS-mediated activation of MLKL and p38 in CAMs. Elucidation of the molecular mechanism underlying zVAD-fmk-induced necroptosis in CAMs might help in better understanding its significance in chronic inflammatory diseases.
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Clorometilcetonas de Aminoácidos/farmacologia , Inibidores de Caspase/farmacologia , Macrófagos/efeitos dos fármacos , Necroptose/efeitos dos fármacos , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Morte Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Fosforilação/efeitos dos fármacosRESUMO
IL-1 family cytokines play a dual role in the gut, with different family members contributing either protective or pathogenic effects. IL-36γ is an IL-1 family cytokine involved in polarizing type-1 immune responses. However, its function in the gut, including in colorectal cancer pathogenesis, is not well appreciated. In a murine model of colon carcinoma, IL-36γ controls tertiary lymphoid structure formation and promotes a type-1 immune response concurrently with a decrease in expression of immune checkpoint molecules in the tumor microenvironment. Here, we demonstrate that IL-36γ plays a similar role in driving a pro-inflammatory phenotype in human colorectal cancer. We analyzed a cohort of 33 primary colorectal carcinoma tumors using imaging, flow cytometry, and transcriptomics to determine the pattern and role of IL-36γ expression in this disease. In the colorectal tumor microenvironment, we observed IL-36γ to be predominantly expressed by M1 macrophages and cells of the vasculature, including smooth muscle cells and high endothelial venules. This pattern of IL-36γ expression is associated with a CD4+ central memory T cell infiltrate and an increased density of B cells in tertiary lymphoid structures, as well as with markers of fibrosis. Conversely, expression of the antagonist to IL-36 signaling, IL-1F5, was associated with intratumoral expression of checkpoint molecules, including PD-1, PD-L1, and CTLA4, which can suppress the immune response. These data support a role for IL-36γ in the physiologic immune response to colorectal cancer by sustaining inflammation within the tumor microenvironment.
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Neoplasias Colorretais/imunologia , Inflamação/imunologia , Interleucina-1/imunologia , Estruturas Linfoides Terciárias/imunologia , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Interleucina-1/metabolismo , Masculino , Pessoa de Meia-Idade , Microambiente Tumoral/genética , Microambiente Tumoral/imunologiaRESUMO
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-1ß, 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.
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Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Diferenciação Celular , Quimiocina CCL17/biossíntese , Quimiocina CCL17/metabolismo , Quimiocina CCL5/biossíntese , Quimiocina CCL5/metabolismo , Quimiocina CXCL10/biossíntese , Quimiocina CXCL10/metabolismo , Quimiocinas CC/biossíntese , Quimiocinas CC/metabolismo , Endocitose/efeitos dos fármacos , Endocitose/imunologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Humanos , Imunofenotipagem , Interferon gama/farmacologia , Interleucina-13/farmacologia , Interleucina-1beta/biossíntese , Interleucina-1beta/metabolismo , Interleucina-4/farmacologia , Interleucina-8/biossíntese , Interleucina-8/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/citologia , Macrófagos/imunologia , Monócitos/citologia , Monócitos/imunologia , Cultura Primária de Células , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Leishmania infection of phagocytic cells, such as macrophages, induces the differentiation of infected cells into different phenotypes according to their surrounding microenvironments. The classical activation of macrophages involves metabolic reprogramming, in which several metabolites such as succinate, fumarate and itaconate are accumulated. The immunoregulatory functions of itaconate in the context of Leishmania infection were investigated in this paper. Ex vivo bone marrow-derived macrophages were differentiated into classically activated macrophages through IFNG activation and infection with Leishmania infantum. A high-throughput real-time qPCR experiment was designed for the analyses of 223 genes involved in immune response and metabolism. The transcriptional profile of classically activated macrophages revealed the enrichment of the IFNG response pathways and the upregulation of genes such as Cxcl9, Irf1, Acod1, Il12b, Il12rb1, Nos2 or Stat1. In vitro pre-stimulation with itaconate induced a loss of the parasite control and the upregulation of genes related to local acute inflammatory response. Our results reveal that itaconate accumulation dampened classically activated macrophage antiparasitic activity, and this is reflected by the differential expression of the Il12b, Icosl and Mki67 genes. The possibility of inducing parasite-killing responses in the host through metabolic reprograming is an interesting approach for the treatment of Leishmania infections that will undoubtedly attract increasing attention in the coming years.
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Macrophage activation refers to the enhanced functionality of macrophages in response to endogenous or exogenous stimuli. Due to the existence of limitless stimuli and a multitude of receptors on macrophage surfaces, the nature of activation (or acquired functioning) can be specific to the encountering stimulus. This article describes a macrophage-activation screening platform in a 96-well format. The methodology involves the generation of bone marrow-derived macrophages, their activation into two extreme activation states, and screening of activated macrophages for expression of bonafide protein biomarkers. A high-throughput and stringent assay to determine macrophage activation markers developed in this article can be adapted for biomarker determination in pathological conditions and toxicant/drug safety screening.
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Tumor-associated macrophages are an important part of the tumor microenvironment. The presence of certain populations of macrophages within tumor tissue may be associated with either better or worse disease prognosis. The study of these cells is currently receiving a great deal of attention, with the most important topics of investigation raised being: the typification of subpopulations of tumor-associated macrophages; identification of the prognostic significance of population density and distribution of macrophages in the tumor microenvironment; ways to influence macrophage activity, migration and differentiation within the tumor. The answers to these questions can improve the efficiency of immunoterapy for malignancies. The presented article briefly reviews recent findings on tumor-associated macrophages in solid malignancies.
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Neoplasias , Macrófagos Associados a Tumor , Humanos , Macrófagos/patologia , Prognóstico , Microambiente TumoralRESUMO
Macrophages comprise the front line of defense against various pathogens. Classically activated macrophages (M1), induced by IFN-γ and LPS, highly express inflammatory cytokines and contribute to inflammatory processes. By contrast, alternatively activated macrophages (M2) are induced by IL-4 and IL-13, produce IL-10, and display anti-inflammatory activity. Adenylate kinase 4 (Ak4), an enzyme that transfers phosphate group among ATP/GTP, AMP, and ADP, is a key modulator of ATP and maintains the homeostasis of cellular nucleotides which is essential for cell functions. However, its role in regulating the function of macrophages is not fully understood. Here we report that Ak4 expression is induced in M1 but not M2 macrophages. Suppressing the expression of Ak4 in M1 macrophages with shRNA or siRNA enhances ATP production and decreases ROS production, bactericidal ability and glycolysis in M1 cells. Moreover, Ak4 regulates the expression of inflammation genes, including Il1b, Il6, Tnfa, Nos2, Nox2, and Hif1a, in M1 macrophages. We further demonstrate that Ak4 inhibits the activation of AMPK and forms a positive feedback loop with Hif1α to promote the expression of inflammation-related genes in M1 cells. Furthermore, RNA-seq analysis demonstrates that Ak4 also regulates other biological processes in addition to the expression of inflammation-related genes in M1 cells. Interestingly, Ak4 does not regulate M1/M2 polarization. Taken together, our study uncovers a potential mechanism linking energy consumption and inflammation in macrophages.
Assuntos
Proteínas Quinases Ativadas por AMP/fisiologia , Adenilato Quinase/fisiologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/fisiologia , Inflamação/etiologia , Macrófagos/fisiologia , Trifosfato de Adenosina/metabolismo , Animais , Polaridade Celular , Células Cultivadas , Feminino , Glicólise , Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismoRESUMO
Metabolic modulation of macrophage activation has emerged as a promising strategy lately in immunotherapeutics. However, macrophages have a broad spectrum of functions and thus, understanding the exact metabolic changes that drive a particular immune response, is of major importance. In our previous work, we have reported a key role of nitric oxide (NOâ) in two(2)-signal activated macrophages [M(2-signals)]. Further characterization using metabolic analysis in intact cells, showed that the basal and maximal respiration levels of M(2-signals) were comparable, with cells being unresponsive to the injections-inducd mitochondrial stress. Here, we show that excessive NOâ secretion by the M(2-signals) macrophages, interferes with the oxygen (O2) consumption measurements on cells using the seahorse metabolic analyzer. This is attributed mainly to the consumption of ambient oxygen by NOâ to form NO2- and/or NO3- but also to the reduction of O2 to superoxide anion (O2â-) by stray electrons from the electron transport chain, leading to the formation of peroxynitrite (ONOO-). We found that reactive species-donors in the absence of cells, produce comparable oxygen consumption rates (OCR) with M(2-signals) macrophages. Furthermore, inhibition of NOâ production, partly recovered the respiration of activated macrophages, while external addition of NOâ in non-activated macrophages downregulated their OCR levels. Our findings are crucial for the accurate metabolic characterization of cells, especially in cases where reactive nitrogen or oxygen species are produced in excess.
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Macrophages are the primary cell type in the immune system and are activated in response to diverse external stimuli. They can exert inflammatory or anti-inflammatory responses depending on the type of stimuli in the system, and as such rely on a complex network of diverse signaling pathways to function. Non-coding RNAs have recently been established as an important class of regulators linked to the activation of macrophages. Although the roles of many microRNAs have been described, the majority of the other types of regulatory non-coding RNAs, including long non-coding RNAs (lncRNAs), remain undefined. In this study, we performed a comprehensive analysis of several large RNA sequencing datasets to identify common lncRNAs differentially expressed after M1 or M2 macrophage induction. We identified the lncRNAs that underwent significant changes in each dataset and established a list of commonly altered lncRNAs. We expect that our data will act as a valuable resource for future studies designed to uncover the roles of these lncRNAs in determining macrophage fates.
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Redes Reguladoras de Genes/genética , Ativação de Macrófagos/genética , Macrófagos/fisiologia , RNA Longo não Codificante/genética , Animais , Linhagem Celular , Perfilação da Expressão Gênica/métodos , Humanos , Camundongos , Células RAW 264.7RESUMO
Purpose: To investigate the role of G-protein-coupled bile acid receptor-1, Gpbar1 (TGR5) in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease.Methods: The mRNA level of TGR5, iNOS, Arg1, CD16, and CD206 in macrophages was assayed by real-time PCR. ELISA was used to detect the production of cytokines in cell culture supernatants. The frequencies of CD4+IFN-γ+ and CD4+ IL-17+ T cells were tested by flow cytometry.Results: A decreased expression of TGR5 in M1 macrophages was observed in active VKH patients as compared with normal controls. TGR5 stimulation of M1 macrophages with INT-777 caused a shift of the inflammatory M1 toward the anti-inflammatory M2 macrophage subtype. TGR5 activation of macrophages co-cultured with CD4+ T cells inhibited Th1 and Th17 polarization, as well as the release of IFN-γ and IL-17 in the culture supernatant.Conclusion: Our results show that a decreased TGR5 expression might contribute to the pathogenesis of VKH disease.
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Regulação da Expressão Gênica , Leucócitos Mononucleares/metabolismo , RNA Mensageiro/genética , Receptores Acoplados a Proteínas G/genética , Síndrome Uveomeningoencefálica/genética , Adulto , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Humanos , Leucócitos Mononucleares/patologia , Masculino , Receptores Acoplados a Proteínas G/biossíntese , Síndrome Uveomeningoencefálica/metabolismo , Síndrome Uveomeningoencefálica/patologiaRESUMO
Glutathione (GSH) transferase (GST) is an essential enzyme in cestodes for the detoxification of xenobiotics. In Taenia solium, two GSTs (Ts25GST and Ts26GST kDa) were isolated as a fraction (SGSTF) by GSH-Sepharose-4B. Both are located on the tegument. Immunization assays with SGSTF reduced up to 90% of the parasitic load in a murine model of cysticercosis. It prompted us to investigate how SGSTF induces this protective immune response. To test it, we exposed peritoneal macrophages to SGSTF for 24 h; such exposure favored the production of IL-12, TNF, and IL-10 as well as the expression of nitric oxide synthase 2 inducible (Nos2) and CD86, but did not induce the expression of chitinase-like 3 (Chil3). Confocal microscopy showed that the macrophages internalize the SGSTF which co-localized after 1 h with MHC-II in their plasma membranes. Macrophages exposed to SGSTF and co-cultured with anti-CD3 pre-activated T CD4+ cells, enhanced the proliferation of CD4+ cells, induced high interferon-γ (IFN-γ) secretion, and elevated the expression of CD25 and CD69, molecules associated with cell activation. Similar assay using T CD4+ cells from DO11.10 mice and ovalbumin (OVA) peptide+SGSTF as stimuli, showed enhanced cell proliferation and OVA-specific IFN-γ secretion. These data are in-line with those indicating that the P1, P5, and P6 peptides of Schistosoma japonicum 28GST highly promote T-cell proliferation and Th1 response in vitro We found that such peptides are also present on Ts25GST and Ts26GST. It suggests that SGSTF activates peritoneal macrophages to a classically activated-like phenotype, and that these macrophages induce the differentiation of T CD4+ cells toward a Th1-type response.
Assuntos
Glutationa Transferase/farmacologia , Macrófagos Peritoneais/imunologia , Taenia solium/enzimologia , Células Th1/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Epitopos , Feminino , Glutationa Transferase/farmacocinética , Interações Hospedeiro-Parasita , Interferon gama/metabolismo , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Camundongos Endogâmicos BALB C , Taenia solium/patogenicidade , Teníase/imunologia , Células Th1/parasitologiaRESUMO
BACKGROUND: Inflammatory bowel disease (IBD) is characterized by chronic and non-specific inflammation of the intestinal mucosa and mainly includes ulcerative colitis and Crohn's disease. AIM: To explore the beneficial effect of ToxoROP16I/III-induced M2 phynotype macrophages in homeostasis of IBDs through downregulation of M1 inflammatory cells. METHODS: RAW264.7 macrophages stimulated by lipopolysaccharide (LPS) (M1 cells) were co-cultured with Caco-2 cells as an inflammatory model of IBD in vitro. The expression of ToxoROP16I/III was observed in RAW264.7 macrophages that were transfected with pEGFP-rop16 I/III. The phenotypes of M2 and M1 macrophage cells were assessed by quantitative real-time reverse transcriptase polymerase chain reaction and the expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, transforming growth factor (TGF)-ß1, IL-10, inducible nitric oxide synthase (iNOS), and arginase-1 (Arg-1) was detected. The expression of iNOS, Arg-1, signal transducer and activator of transcription 3 (Stat3), p-Stat3, Stat6, p-Stat6, programmed death ligand-2 (PD-L2), caspase-3, -8, and -9 was analyzed by Western blotting, and Griess assays were performed to detect nitric oxide (NO). TNF-α, IL-1ß, IL-6, TGF-ß1, and IL-10 expression in the supernatants was detected by enzyme-linked immunosorbent assay, and Caco-2 cell apoptosis was determined by flow cytometry after mixing M1 cells with M2 cells in a Caco-2 cell co-culture system. RESULTS: M1 cells exhibited significantly increased production of iNOS, NO, TNF-α, IL-1ß, and IL-6, while ToxoROP16I/III induced macrophage bias to M2 cells in vitro, showing increased expression of Arg-1, IL-10 and TGF-ß1 and elevated production of p-Stat3 and p-Stat6. The mixed M1 and M2 cell culture induced by ToxoROP16I/III exhibited decreased production of NO and iNOS and upregulated expression of Arg-1 and PD-L2. Accordingly, Caco-2 cells became apoptotic, and apoptosis-associated proteins such as caspase-3, -8 and -9 were dampened during co-culture of M1 and M2 cells. Flow cytometry analysis showed that co-culture of M1 cells with Caco-2 cells facilitated the apoptosis of Caco-2 cells, but co-culture of M1 and M2 cells alleviated Caco-2 cell apoptosis. CONCLUSION: ToxoROP16I/III-induced M2 macrophages inhibited apoptosis of Caco-2 cells caused by M1 macrophages. This finding may help gain a better understanding of the underlying mechanism and represent a promising therapeutic strategy for IBDs.
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Doenças Inflamatórias Intestinais/fisiopatologia , Doenças Inflamatórias Intestinais/terapia , Mucosa Intestinal/fisiopatologia , Macrófagos/metabolismo , Proteínas Tirosina Quinases/metabolismo , Proteínas de Protozoários/metabolismo , Toxoplasma/metabolismo , Animais , Apoptose , Células CACO-2 , Técnicas de Cocultura , Citocinas/metabolismo , Regulação para Baixo , Homeostase , Humanos , Imunoterapia , Inflamação , Lentivirus , Lipopolissacarídeos , Macrófagos/efeitos dos fármacos , Camundongos , Fenótipo , Células RAW 264.7RESUMO
Macrophage activation is a process that is highly relevant in both infectious and chronic diseases and so the study of macrophage activation is of high interest to investigators in many fields of biomedical research. Bone marrow-derived macrophages are a popular choice for studying macrophage activation in vitro, as these cells are relatively hardy and many macrophages can be produced from just one mouse. Here we describe the activation of mouse macrophages in vitro, including special considerations for cell culture, detachment, kinetics, and activation states.
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Células da Medula Óssea/citologia , Técnicas de Cultura de Células/métodos , Ativação de Macrófagos , Macrófagos/citologia , Animais , Cinética , Macrófagos/metabolismo , CamundongosRESUMO
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-1ß, 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.
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Ciclofilina A/farmacologia , Macrófagos/efeitos dos fármacos , Adulto , Idoso , Animais , Artrite Experimental/induzido quimicamente , Artrite Experimental/patologia , Colágeno/efeitos adversos , Feminino , Humanos , Inflamação , Masculino , Camundongos , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Ativação Transcricional/efeitos dos fármacosRESUMO
Cellular interactions are critical during development, tissue fitness and epithelial tumor development. The expression levels of specific genes confer to tumoral cells a survival advantage versus the normal neighboring cells. As a consequence, cells surrounding tumors are eliminated and engulfed by macrophages. We propose a novel scenario in which circulating cells facing a tumor can reproduce these cellular interactions. In vitro cultured macrophages from murine bone marrow were used to investigate this hypothesis. M1 macrophages in tumoral medium upregulated markers of a suboptimal condition, such as Sparc and TyrRS, and undergo apoptosis. However, M2 macrophages display higher Myc expression levels and proliferate at the expense of M1. Resulting M1 apoptotic debris is engulfed by M2 in a Sparc- and TyrRS-dependent manner. These findings suggest that tumor-dependent macrophage elimination could deplete immune response against tumors. This possibility could be relevant for macrophage based anti-tumoral strategies.
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BACKGROUND: The role of the macrophages in cystic fibrosis (CF) lung disease has been poorly studied. We hypothesized that alternatively activated M2 macrophages are abnormal in CF lung disease. METHODS: Blood samples were collected from adults (n=13) children (n=27) with CF on admission for acute pulmonary exacerbation and when clinically stable. Monocytes were differentiated into macrophages and polarized into classical (M1) and alternatively-activated (M2) phenotypes, function determined ex-vivo and compared with healthy controls. RESULTS: In the absence of functional cystic fibrosis trans-membrane conductance regulator (CFTR), either naturally in patients with CF or induced with CFTR inhibitors, monocyte-derived macrophages do not respond to IL-13/IL-4, fail to polarize into M2s associated with a post-transcriptional failure to produce and express IL-13Rα1 on the macrophage surface Polarization to the M1 phenotype was unaffected. CONCLUSIONS: CFTR-dependent imbalance of macrophage phenotypes and functions could contribute to the exaggerated inflammatory response seen in CF lung disease.
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Fibrose Cística , Subunidade alfa1 de Receptor de Interleucina-13/imunologia , Pulmão , Ativação de Macrófagos/imunologia , Adulto , Criança , Fibrose Cística/imunologia , Fibrose Cística/patologia , Fibrose Cística/fisiopatologia , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Feminino , Humanos , Inflamação/imunologia , Interleucina-13/imunologia , Interleucina-4/imunologia , Pulmão/imunologia , Pulmão/patologia , Pulmão/fisiopatologia , Macrófagos/imunologia , Macrófagos/patologia , MasculinoRESUMO
Chromogranin-A (CHGA) is a prohormone secreted by neuroendocrine cells and is a precursor of several bioactive peptides, which are implicated in different and distinctive biological and immune functions. Chromofungin (CHR: CHGA47-66) is a short peptide with antimicrobial effects and encodes from CHGA exon-IV. Inflammatory bowel disease (IBD) is characterized by alterations in the activation of pro-inflammatory pathways, pro-inflammatory macrophages (M1), and nuclear transcription factor kappa B (NF-κB) signaling leading to the perpetuation of the inflammatory process. Here, we investigated the activity of CHR (CHGA Exon-IV) in persons with active ulcerative colitis (UC) and the underlying mechanisms in dextran sulfate sodium (DSS)-colitis in regard to macrophages activation and migration. Tissue mRNA expression of CHR (CHGA Exon-IV) was down regulated in active UC compared to healthy individuals and negatively correlated with pro-inflammatory macrophages (M1) cytokines, toll-like receptors (TLR)-4, and pNF-κB activity. In DSS colitis, CHR (CHGA Exon-IV) expression was reduced, and exogenous CHR treatment decreased the severity of colitis associated with a reduction of M1 macrophages markers and pNF-κB. In vitro, CHR treatment reduced macrophages migration, decreased pro-inflammatory cytokines production and pNF-κB. Targeting CHR may represent a promising new direction in research to define new therapeutic targets and biomarkers associated with IBD.
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Cromogranina A/metabolismo , Cromogranina A/farmacologia , Colite Ulcerativa/metabolismo , Inflamação/metabolismo , Macrófagos Peritoneais/efeitos dos fármacos , NF-kappa B/metabolismo , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Animais , Colite/induzido quimicamente , Sulfato de Dextrana/toxicidade , Regulação para Baixo , Regulação da Expressão Gênica , Humanos , Lipopolissacarídeos/toxicidade , Masculino , Camundongos Endogâmicos C57BL , NF-kappa B/genética , Transdução de SinaisRESUMO
A clear understanding of the interactions between classically activated macrophages (MÏ1) and γδT cells may improve current therapeutic approaches, including that of immunotherapy for treating certain types of cancer. The present study aimed to expand the current knowledge by showing the effect of culture supernatants of MÏ1 on the proliferation, cell surface marker expression and tumor suppression effects of γδT cells, and by exploring the potential mechanisms involved. In vitro, MÏ1 were cultured by GM-CSF and IFN-γ. The isopentenyl pyrophosphate method was used to amplify human peripheral blood γδT cells. The surface markers of macrophages and γδT cells were detected by flow cytometry. The proliferation of γδT cells induced by the culture supernatants of MÏ1 was investigated using the MTT assay. The lactate dehydrogenase method was used to detect the cytotoxicity of γδT cells on the SGC-7901 gastric cancer cell line. Ten days after cultivation, the percentage of γδT cells from the repertoire of naive cells, expanded from 4.21 to 91.27%. The percentage of cells expressing CD44 was 94%. The percentage of CD68 on cultured MÏ1 was increased from 17.7 to 73.2%. The culture supernatants of MÏ1 increased the proliferation of γδT cells compared with the control group (33.8% vs. 0, P<0.01). The culture supernatants of MÏ1 increased the cytotoxicity of γδT cells compared with the control group (70.18 vs. 47.25%, P<0.01). In conclusion, the supernatant of cultured MÏ1 promotes the proliferation of γδT cells and their cytotoxic effect on the SGC-7901 gastric cancer cell line.
RESUMO
Macrophages are abundant in the tumor microenvironment where they adopt a pro-tumor phenotype following alternative polarization induced by paracrine factors from cancer and stromal cells. In contrast, classically activated macrophages have tumoricidal activities, such that the polarization of tumor-associated macrophages has become a novel therapeutic target. Toll-like receptor 4 engagement promotes classical activation of macrophages, and recent literature suggests TLR4 agonism to prevent metastasis and promote survival in experimental metastasis models. A growing number of studies indicate that TLR4 can respond to opioids, including the opioid receptor-inactive morphine metabolite morphine-3-glucuronide (M3G). We measured the activation of TLR4 in a reporter cell line exogenously expressing TLR4 and TLR4 co-receptors, and confirmed that M3G weakly but significantly activates TLR4. We hypothesized that M3G would promote the expression of classical activation signature genes in macrophages in vitro. We exposed mouse and human macrophage cell lines to M3G or the TLR4 activator lipopolysaccharide (LPS), alone or in combination with interferon gamma (IFN-γ). The classical macrophage activation markers tested were iNOS, CD86, IL-6, or TNF-α in RAW 264.7 cells and IL-6, IL-12, IL-23, TNF-α, CXCL10, and CXCL11 in THP1 cells. Our results show that despite exhibiting TLR4-activation ability, M3G does not elicit the expression of classical activation markers in LPS-responsive macrophages.