Recent studies on intracellular metabolism in dendritic cells (DCs) and macrophages provide new insights on the functioning of these critical controllers of innate and adaptive immunity. Both cell types undergo profound metabolic reprogramming in response to environmental cues, such as hypoxia or nutrient alterations, but importantly also in response to danger signals and cytokines. Metabolites such as succinate and citrate have a direct impact on the functioning of macrophages. Immunogenicity and tolerogenicity of DCs is also determined by anabolic and catabolic processes, respectively. These findings provide new prospects for therapeutic manipulation in inflammatory diseases and cancer.

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Immunometabolism governs dendritic cell and macrophage function

The induction of CD11b(+)Gr-1(+) myeloid-derived suppressor cells (MDSCs) is an important immune-evading mechanism used by tumors. However, the exact nature and function of MDSCs remain elusive, especially because they constitute a heterogeneous population that has not yet been clearly defined. Here, we identified 2 distinct MDSC subfractions with clear morphologic, molecular, and functional differences. These fractions consisted of either mononuclear cells (MO-MDSCs), resembling inflammatory monocytes, or low-density polymorphonuclear cells (PMN-MDSCs), akin to immature neutrophils. Interestingly, both MO-MDSCs and PMN-MDSCs suppressed antigen-specific T-cell responses, albeit using distinct effector molecules and signaling pathways. Blocking IFN-gamma or disrupting STAT1 partially impaired suppression by MO-MDSCs, for which nitric oxide (NO) was one of the mediators. In contrast, while IFN-gamma was strictly required for the suppressor function of PMN-MDSCs, this did not rely on STAT1 signaling or NO production. Finally, MO-MDSCs were shown to be potential precursors of highly antiproliferative NO-producing mature macrophages. However, distinct tumors differentially regulated this inherent MO-MDSC differentiation program, indicating that this phenomenon was tumor driven. Overall, our data refine tumor-induced MDSC functions by uncovering mechanistically distinct MDSC subpopulations, potentially relevant for MDSC-targeted therapies.

Identification of discrete tumor-induced myeloid-derived suppressor cell subpopulations with distinct T-CELL suppressive activity

Cysteine cathepsins: From structure, function and regulation to new frontiers

Astrocytes are neural cells of ectodermal, neuroepithelial origin that provide for homeostasis and defense of the central nervous system (CNS). Astrocytes are highly heterogeneous in morphological appearance; they express a multitude of receptors, channels, and membrane transporters. This complement underlies their remarkable adaptive plasticity that defines the functional maintenance of the CNS in development and aging. Astrocytes are tightly integrated into neural networks and act within the context of neural tissue; astrocytes control homeostasis of the CNS at all levels of organization from molecular to the whole organ.

Physiology of astroglia

Despite widespread clinical use of antimalarial drugs such as hydroxychloroquine and chloroquine in the treatment of rheumatoid arthritis (RA), systemic lupus erythematosus (SLE) and other inflammatory rheumatic diseases, insights into the mechanism of action of these drugs are still emerging. Hydroxychloroquine and chloroquine are weak bases and have a characteristic ‘deep’ volume of distribution and a half-life of around 50 days. These drugs interfere with lysosomal activity and autophagy, interact with membrane stability and alter signalling pathways and transcriptional activity, which can result in inhibition of cytokine production and modulation of certain co-stimulatory molecules. These modes of action, together with the drug’s chemical properties, might explain the clinical efficacy and well-known adverse effects (such as retinopathy) of these drugs. The unknown dose–response relationships of these drugs and the lack of definitions of the minimum dose needed for clinical efficacy and what doses are toxic pose challenges to clinical practice. Further challenges include patient non-adherence and possible context-dependent variations in blood drug levels. Available mechanistic data give insights into the immunomodulatory potency of hydroxychloroquine and provide the rationale to search for more potent and/or selective inhibitors. Hydroxychloroquine and chloroquine are antimalarial drugs commonly used for the treatment of rheumatic diseases. Multiple mechanisms might explain the efficacy and adverse effects of these drugs, but further investigation could lead to the development of more specific and potent drugs.

Mechanisms of action of hydroxychloroquine and chloroquine: implications for rheumatology.

Endosomal TLRs play an important role in innate immune response as well as in autoimmune processes. In the therapy of systemic lupus erythematosus, antimalarial drugs chloroquine, hydroxychloroquine, and quinacrine have been used for a long time. Their suppression of endosomal TLR activation has been attributed to the inhibition of endosomal acidification, which is a prerequisite for the activation of these receptors. We discovered that chloroquine inhibits only activation of endosomal TLRs by nucleic acids, whereas it augments activation of TLR8 by a small synthetic compound, R848. We detected direct binding of antimalarials to nucleic acids by spectroscopic experiments and determined their cellular colocalization. Further analysis revealed that other nucleic acid-binding compounds, such as propidium iodide, also inhibited activation of endosomal TLRs and colocalized with nucleic acids to endosomes. We found that imidazoquinolines, which are TLR7/8 agonists, inhibit TLR9 and TLR3 even in the absence of TLR7 or TLR8, and their mechanism of inhibition is similar to the antimalarials. In contrast to bafilomycin, none of the tested antimalarials and imidazoquinolines inhibited endosomal proteolysis or increased the endosomal pH, confirming that inhibition of pH acidification is not the underlying cause of inhibition. We conclude that the direct binding of inhibitors to nucleic acids mask their TLR-binding epitope and may explain the efficiency of those compounds in the treatment of autoimmune diseases.

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Mechanism of Endosomal TLR Inhibition by Antimalarial Drugs and Imidazoquinolines

C-type lectin DC-SIGN: an adhesion, signalling and antigen-uptake molecule that guides dendritic cells in immunity.

A molecule with a relatively simple chemical structure, resveratrol has been found to interact with multiple molecular targets, many of them associated with inflammation and immunity. Indeed, it has been shown to act directly on central players of both innate and adaptive immunity, such as macrophages, lymphocytes, and dendritic cells. In addition, there is very little evidence suggesting significant deleterious side effects of resveratrol, further highlighting its potential future use as a therapeutic agent. This review provides an up-to-date discussion on recent advances regarding anti-inflammatory effects of resveratrol, mechanisms of action, and its potential for therapeutic use.

Anti-inflammatory effects of resveratrol and its potential use in therapy of immune-mediated diseases.

In immune-mediated diseases of the central nervous system, astrocytes exposed to interferon-γ (IFN-γ) can express major histocompatibility complex (MHC) class II molecules and antigens on their surface. MHC class II molecules are thought to be delivered to the cell surface by membrane-bound vesicles. However, the characteristics and dynamics of this vesicular traffic are unclear, particularly in reactive astrocytes, which overexpress intermediate filament (IF) proteins that may affect trafficking. The aim of this study was to determine the mobility of MHC class II vesicles in wild-type (WT) astrocytes and in astrocytes devoid of IFs. The identity of MHC class II compartments in WT and IF-deficient astrocytes 48 h after IFN-γ activation was determined immunocytochemically by using confocal microscopy. Time-lapse confocal imaging and Alexa Fluor546-dextran labeling of late endosomes/lysosomes in IFN-γ treated cells was used to characterize the motion of MHC class II vesicles. The mobility of vesicles was analyzed using ParticleTR software. Confocal imaging of primary cultures of WT and IF-deficient astrocytes revealed IFN-γ induced MHC class II expression in late endosomes/lysosomes, which were specifically labeled with Alexa Fluor546-conjugated dextran. Live imaging revealed faster movement of dextran-positive vesicles in IFN-γ-treated than in untreated astrocytes. Vesicle mobility was lower in IFN-γ-treated IF-deficient astrocytes than in WT astrocytes. Thus, the IFN-γ-induced increase in the mobility of MHC class II compartments is IF-dependent. Since reactivity of astrocytes is a hallmark of many CNS pathologies, it is likely that the up-regulation of IFs under such conditions allows a faster and therefore a more efficient delivery of MHC class II molecules to the cell surface. In vivo, such regulatory mechanisms may enable antigen-presenting reactive astrocytes to respond rapidly and in a controlled manner to CNS inflammation.

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IFN-γ-induced increase in the mobility of MHC class II compartments in astrocytes depends on intermediate filaments

Resveratrol is a polyphenol that acts on multiple molecular targets important for cell differentiation and activation. Dendritic cells (DCs) are a functionally diverse cell type and represent the most potent antigen-presenting cells of the immune system. In this study, we investigated resveratrol-induced effects on DCs during their differentiation and maturation. Our results show that resveratrol induces DC-associated tolerance, particularly when applied during DC differentiation. Costimulatory molecules CD40, CD80 and CD86 were down-regulated, as was the expression of major histocompatibility complex (MHC) class II molecules. Surface expression of inhibitory immunoglobulin-like transcript 3 (ILT3) and ILT4 molecules was induced, while human leucocyte antigen (HLA)-G expression was not affected. Resveratrol-treated DCs lost the ability to produce interleukin (IL)-12p70 after activation, but had an increased ability to produce IL-10. Such DCs were poor stimulators of allogeneic T cells and had lowered ability to induce CD4(+) T-cell migration. Furthermore, treated cells were able to generate allogeneic IL-10-secreting T cells, but were not competent in inducing FoxP3 expression These tolerogenic effects are probably associated with the effect of resveratrol on multiple molecular targets through which it interferes with DC differentiation and nuclear factor (NF)-kappaB translocation. Our data provide new insights into the molecular and functional mechanisms of the tolerogenic effects that resveratrol exerts on DCs.

Urban Švajger

Papers

Mechanism of Endosomal TLR Inhibition by Antimalarial Drugs and Imidazoquinolines

C-type lectin DC-SIGN: an adhesion, signalling and antigen-uptake molecule that guides dendritic cells in immunity.

Anti-inflammatory effects of resveratrol and its potential use in therapy of immune-mediated diseases.

IFN-γ-induced increase in the mobility of MHC class II compartments in astrocytes depends on intermediate filaments

Dendritic cells treated with resveratrol during differentiation from monocytes gain substantial tolerogenic properties upon activation.