•Purified bacterial 2-substituted (1–3)-β-d-glucan activated human macrophages.•This activation was indicative of an anti-inflammatory response.•The activation differed markedly from that caused by ...bacterial lipopolysaccharide.•This glucan from natural and recombinant strains had the same biological activity.
β-glucans produced by eukaryotic cells and by microorganisms are known to modulate immune responses by affecting macrophage activation. Here, we have investigated the effect of purified 2-substituted (1–3)-β-d-glucan, produced by either Pediococcus parvulus 2.6 or Lactococcus lactis NZ9000pNGTF, on the effector functions of human PMA-differentiated THP-1 cells and M1 pro-inflammatory monocyte-derived macrophages. The results reveal that this kind of β-D-glucan activates macrophages and has an anti-inflammatory effect.
Dendritic cells (DCs) promote tolerance or immunity depending on their maturation state, which is enhanced or accelerated upon MEK-ERK signaling pathway inhibition. We have determined the ...contribution of MEK-ERK activation to the profile of gene expression of human immature monocyte-derived dendritic cells (MDDCs) and peripheral blood myeloid DCs. ERK inhibition altered the expression of genes that mediate Chemokine (C-C motif) ligand 19 (CCL19)–directed migration (CCR7) and low-density lipoprotein (LDL) binding (CD36, SCARB1, OLR1, CXCL16) by immature DCs. In addition, ERK upregulated CCL2 expression while impairing the expression of DC maturation markers (RUNX3, ITGB7, IDO1). MEK-ERK–regulated genes exhibited an overrepresentation of cognate sequences for the aryl hydrocarbon receptor (AhR) transcription factor, whose transcriptional and DNA-binding activities increased in MDDCs upon exposure to the MEK1/2 inhibitor U0126. Therefore, the MEK-ERK signaling pathway regulates antigen capture, lymph node homing, and acquisition of maturation-associated genes, and its contribution to the maintenance of the immature state of MDDCs and myeloid DCs is partly dependent on the activity of AhR. Since pharmacologic modulation of the MEK-ERK signaling pathway has been proposed as a potential therapeutic strategy for cancer, our findings indicate that ERK inhibitors might influence antitumor responses through regulation of critical DC effector functions.
•Aryl hydrocarbon receptor (AhR) mediates the ERK-dependent maintenance of the immature state of monocyte-derived dendritic cells (MDDCs).•MEK-ERK regulates antigen capture, lymph node homing, and the acquisition of maturation-associated genes in MDDCs.
Abstract Microenvironmental conditions in infected, inflamed or damaged tissues are characterized by low levels of oxygen (hypoxia) and nutrients. Myeloid cells (mostly macrophages and neutrophils) ...account for 95% of the cells newly recruited into inflammatory sites, and exert their effector functions under these restrictive conditions. In the case of macrophages, adaptation to the surrounding tissue environment is underlined by their huge metabolic and functional plasticity, which allows them to critically participate in the maintenance of tissue homeostasis and the initiation and resolution of inflammatory processes under hypoxic conditions. Therefore, alterations in oxygen availability directly affect the macrophage functional state (polarization), a phenomenon that has been already illustrated in pathologies like cancer, atherosclerosis and obesity. This review summarizes recent advances on the molecular basis of macrophage sensing and response to changes in oxygen pressure, emphasizing the link among the hypoxia-induced signalling pathways, macrophage polarization and inflammatory pathologies.
Macrophages display a ample plethora of effector functions whose acquisition is promoted by the surrounding cytokine and cellular environment. Depending on the stimulus, macrophages become ...specialized ("polarized") for either pathogen elimination, tissue repair and wound healing or immunosuppression. This "polarization" versatility allows macrophages to critically contribute to tissue homeostasis, as they promote initiation and resolution of inflammatory responses. As a consequence, deregulation of the tissue macrophage polarization balance is an etiological agent of chronic inflammation, autoimmune diseases, cancer and even obesity and insulin resistance. In the present review we describe current concepts on the molecular basis and the patho-physiological implications of macrophage polarization, and describe its modulation by serotonin (5-HT), a neurotransmitter that regulates inflammation and tissue repair via a large set of receptors (5-HTR1-7). 5-HT modulates the phenotypic and functional polarization of macrophages, and contributes to the maintenance of an anti-inflammatory state mainly via 5-HTR2B and 5-HTR7, whose activation has a great impact on macrophage gene expression profile. The identification of 5-HTR2B and 5-HTR7 as functionally-relevant polarization markers suggests their therapeutic value in inflammatory pathologies as well as their potential involvement in linking the immune and nervous systems.
Tumor-associated macrophages (TAM) are important components of the multiple myeloma (MM) microenvironment that support malignant plasma cell survival and resistance to therapy. It has been proposed ...that macrophages (MØ) retain the capacity to change in response to stimuli that can restore their antitumor functions. Here, we investigated several approaches to reprogram MØ as a novel therapeutic strategy in MM. First, we found tumor-limiting and tumor-supporting capabilities for monocyte-derived M1-like MØ and M2-like MØ, respectively, when mixed with MM cells, both in vitro and in vivo. Multicolor confocal microscopy revealed that MM-associated MØ displayed a predominant M2-like phenotype in the bone marrow of MM patient samples, and a high expression of the pro-M2 cytokine macrophage migration inhibitory factor (MIF). To reprogram the protumoral M2-like MØ present in MM toward antitumoral M1-like MØ, we tested the pro-M1 cytokine granulocyte–macrophage colony-stimulating factor (GM-CSF) plus blockade of the M2 cytokines macrophage colony-stimulating factor or MIF. The combination of GM-CSF plus the MIF inhibitor 4-iodo-6-phenyl-pyrimidine achieved the best reprogramming responses toward an M1 profile, at both gene and protein expression levels, as well as remarkable tumoricidal effects. Furthermore, this combined treatment elicited MØ-dependent therapeutic responses in MM xenograft mouse models, which were linked to upregulation of M1 and reciprocal downregulation of M2 MØ markers. Our results reveal the therapeutic potential of reprogramming MØ in the context of MM.
•We report strategies to reprogram macrophages as a novel approach to treat MM mouse models using pro-M1 and blocking M2 signals.•MIF is upregulated in the bone marrow microenvironment of MM patients and plays an autocrine role in protumoral MØ polarization.
Lung surfactant protein A (SP-A) plays an important function in modulating inflammation in the lung. However, the exact role of SP-A and the mechanism by which SP-A affects IFN-γ-induced activation ...of alveolar macrophages (aMϕs) remains unknown. To address these questions, we studied the effect of human SP-A on rat and human aMϕs stimulated with IFN-γ, LPS, and combinations thereof and measured the induction of proinflammatory mediators as well as SP-A's ability to bind to IFN-γ or IFN-γR1. We found that SP-A inhibited (IFN-γ + LPS)-induced TNF-α, iNOS, and CXCL10 production by rat aMϕs. When rat macrophages were stimulated with LPS and IFN-γ separately, SP-A inhibited both LPS-induced signaling and IFN-γ-elicited STAT1 phosphorylation. SP-A also decreased TNF-α and CXCL10 secretion by ex vivo-cultured human aMϕs and M-CSF-derived macrophages stimulated by either LPS or IFN-γ or both. Hence, SP-A inhibited upregulation of IFN-γ-inducible genes (CXCL10, RARRES3, and ETV7) as well as STAT1 phosphorylation in human M-CSF-derived macrophages. In addition, we found that SP-A bound to human IFN-γ (KD = 11 ± 0.5 nM) in a Ca(2+)-dependent manner and prevented IFN-γ interaction with IFN-γR1 on human aMϕs. We conclude that SP-A inhibition of (IFN-γ + LPS) stimulation is due to SP-A attenuation of both inflammatory agents and that the binding of SP-A to IFN-γ abrogates IFN-γ effects on human macrophages, suppressing their classical activation and subsequent inflammatory response.
CLEC5A and CD163L1: new markers of human macrophage pro‐ and anti‐inflammatory polarization in healthy and pathological tissues.
Macrophages (Mφ) can be differentiated and polarized in vitro from ...human CD14+ monocytes under the influence of GM‐CSF (GM‐Mφ) and M‐CSF (M‐Mφ). GM‐Mφs are proinflammatory and M‐Mφs have an anti‐inflammatory phenotype. We found selective expression of the lectin C‐type lectin domain family 5 member A (CLEC5A) transcripts in GM‐Mφs and the scavenger receptor CD163 molecule‐like 1 (CD163L1) in M‐Mφs by microarray assay. In vitro, CD163L1 expression was induced by IL‐10 and M‐CSF and CLEC5A by inflammatory cytokines and cell adherence. In secondary lymphoid organs, their respective expression was restricted to CD68+/CD163+ Mφs that preferentially produced either TNF (CLEC5A+) or IL‐10 (CD163L1+). Mφs from healthy liver and colon tissue were mostly CD163L1+, and CLEC5A+ cells were scarce. In contrast, CLEC5A+ Mφs were abundant in the intestinal lamina propria from patients with inflammatory bowel disease (IBD), with higher numbers of CLEC5A+CD163L1+ found compared with those in secondary lymphoid organs. CLEC5A+ cells were CD14+CD209−CD11b+CD11c+TNF+IL‐10+, and single positive CD163L1+ cells were CD14−CD209+CD11b−CD11c−TNF−IL‐10+ in healthy donors and had lost the ability to produce IL‐10 and to express CD209 in those with IBD. In melanomas, CLEC5A+ tumor‐associated Mφs (TAMs) were not detected in 42% of the cases evaluated, but CD163L1+ TAMs were found in 100%. Similar to IBD, CD163L1+ TAMs expressed high levels of CD209 and produced significant amounts of IL‐10, and CLEC5A+ TAMs were CD14hi and produced enhanced levels of TNF in metastases. Overall, these results suggest that CD163L1 expression is associated with tissue‐resident Mφs with an anti‐inflammatory or anergic phenotype and that CLEC5A+ Mφs exhibit TNF‐producing ability and might display a proinflammatory effect.
Human LSECtin (liver and lymph node sinusoidal endothelial cell C‐type lectin, CLEC4G) is a C‐type lectin encoded within the L‐SIGN/DC‐SIGN/CD23 gene cluster. LSECtin acts as a pathogen attachment ...factor for Ebolavirus and the SARS coronavirus, and its expression can be induced by interleukin‐4 on monocytes and macrophages. Although reported as a liver and lymph node sinusoidal endothelial cell‐specific molecule, LSECtin could be detected in the MUTZ‐3 dendritic‐like cell line at the messenger RNA (mRNA) and protein level, and immunohistochemistry analysis on human liver revealed its presence in Kupffer cells coexpressing the myeloid marker CD68. The expression of LSECtin in myeloid cells was further corroborated through the analysis of the proximal regulatory region of the human LSECtin gene, whose activity was maximal in LSECtin+ myeloid cells, and which contains a highly conserved PU.1‐binding site. PU.1 transactivated the LSECtin regulatory region in collaboration with hematopoietic‐restricted transcription factors (Myb, RUNX3), and was found to bind constitutively to the LSECtin proximal promoter. Moreover, knockdown of PU.1 through the use of small interfering RNA led to a decrease in LSECtin mRNA levels in THP‐1 and monocyte‐derived dendritic cells, thus confirming the involvement of PU.1 in the myeloid expression of the lectin. Conclusion: LSECtin is expressed by liver myeloid cells, and its expression is dependent on the PU.1 transcription factor. (HEPATOLOGY 2009;49:287–296.)
Dendritic cell-specific ICAM-3 grabbing nonintegrin (DC-SIGN) is a monocyte-derived dendritic cell (MDDC)-specific lectin which participates in dendritic cell (DC) migration and DC-T lymphocyte ...interactions at the initiation of immune responses and enhances trans-infection of T cells through its HIV gp120-binding ability. The generation of a DC-SIGN-specific mAb has allowed us to determine that the acquisition of DC-SIGN expression during the monocyte-DC differentiation pathway is primarily induced by IL-4, and that GM-CSF cooperates with IL-4 to generate a high level of DC-SIGN mRNA and cell surface expression on immature MDDC. IL-4 was capable of inducing DC-SIGN expression on monocytes without affecting the expression of other MDDC differentiation markers. By contrast, IFN-alpha, IFN-gamma, and TGF-beta were identified as negative regulators of DC-SIGN expression, as they prevented the IL-4-dependent induction of DC-SIGN mRNA on monocytes, and a similar inhibitory effect was exerted by dexamethasone, an inhibitor of the monocyte-MDDC differentiation pathway. The relevance of the inhibitory action of dexamethasone, IFN, and TGF-beta on DC-SIGN expression was emphasized by their ability to inhibit the DC-SIGN-dependent HIV-1 binding to differentiating MDDC. These results demonstrate that DC-SIGN, considered as a MDDC differentiation marker, is a molecule specifically expressed on IL-4-treated monocytes, and whose expression is subjected to a tight regulation by numerous cytokines and growth factors. This feature might help in the development of strategies to modulate the DC-SIGN-dependent cell surface attachment of HIV for therapeutic purposes.
Upon inflammation, monocyte-derived macrophages (MΦ) infiltrate blood vessels to regulate several processes involved in vascular pathophysiology. However, little is known about the mediators ...involved. Macrophage polarization is crucial for a fast and efficient initial response (GM-MΦ) and a good resolution (M-MΦ) of the inflammatory process. The functional activity of polarized MΦ is exerted mainly through their secretome, which can target other cell types, including endothelial cells. Endoglin (CD105) is a cell surface receptor expressed by endothelial cells and MΦ that is markedly upregulated in inflammation and critically involved in angiogenesis. In addition, a soluble form of endoglin with anti-angiogenic activity has been described in inflammation-associated pathologies. The aim of this work was to identify components of the MΦ secretome involved in the shedding of soluble endoglin. We find that the GM-MΦ secretome contains metalloprotease 12 (MMP-12), a GM-MΦ specific marker that may account for the anti-angiogenic activity of the GM-MΦ secretome. Cell surface endoglin is present in both GM-MΦ and M-MΦ, but soluble endoglin is only detected in GM-MΦ culture supernatants. Moreover, MMP-12 is responsible for the shedding of soluble endoglin in vitro and in vivo by targeting membrane-bound endoglin in both MΦ and endothelial cells. These data demonstrate a direct correlation between GM-MΦ polarization, MMP-12, and soluble endoglin expression and function. By targeting endothelial cells, MMP-12 may represent a novel mediator involved in vascular homeostasis.
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