Dataset Information

Antibody-Directed Glucocorticoid Targeting to CD163 in M2-type Macrophages Attenuates Fructose-Induced Liver Inflammatory Changes.

ABSTRACT: Increased consumption of high-caloric carbohydrates contributes substantially to endemic non-alcoholic fatty liver disease in humans, covering a histological spectrum from fatty liver to steatohepatitis. Hypercaloric intake and lipogenetic effects of fructose and endotoxin-driven activation of liver macrophages are suggested to be essential to disease progression. In the present study, we show that a low dose of an anti-CD163-IgG-dexamethasone conjugate targeting the hemoglobin scavenger receptor CD163 in Kupffer cells and other M2-type macrophages has a profound effect on liver inflammatory changes in rats on a high-fructose diet. The diet induced severe non-alcoholic steatohepatitis (NASH)-like changes within a few weeks but the antibody-drug conjugate strongly reduced inflammation, hepatocyte ballooning, fibrosis, and glycogen deposition. Non-conjugated dexamethasone or dexamethasone conjugated to a control IgG did not have this effect but instead exacerbated liver lipid accumulation. The low-dose anti-CD163-IgG-dexamethasone conjugate displayed no apparent systemic side effects. In conclusion, macrophage targeting by antibody-directed anti-inflammatory low-dose glucocorticoid therapy seems to be a promising approach for safe treatment of fructose-induced liver inflammation.

SUBMITTER: Svendsen P

PROVIDER: S-EPMC5363319 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature

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Antibody-Directed Glucocorticoid Targeting to CD163 in M2-type Macrophages Attenuates Fructose-Induced Liver Inflammatory Changes.

Svendsen Pia P Graversen Jonas H JH Etzerodt Anders A Hager Henrik H Røge Rasmus R Grønbæk Henning H Christensen Erik I EI Møller Holger J HJ Vilstrup Hendrik H Moestrup Søren K SK

Molecular therapy. Methods & clinical development 20161224

Increased consumption of high-caloric carbohydrates contributes substantially to endemic non-alcoholic fatty liver disease in humans, covering a histological spectrum from fatty liver to steatohepatitis. Hypercaloric intake and lipogenetic effects of fructose and endotoxin-driven activation of liver macrophages are suggested to be essential to disease progression. In the present study, we show that a low dose of an anti-CD163-IgG-dexamethasone conjugate targeting the hemoglobin scavenger recepto ...[more]

PMID: 28344991

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Project description: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)-? 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-?, IL-1?, 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 and practical approach for inflammatory conditions that could lead to persistent pain, i.e. major surgeries, burns, rheumatoid arthritis, etc.

Project description:Inflammatory breast cancer (IBC) is the most aggressive and lethal form of breast cancer, characterized by a high infiltration of tumor-associated macrophages and poor prognosis. To identify new biomarkers and to elucidate the molecular mechanisms underlying IBC pathogenesis, we investigated the expression pattern of heparanase (HPSE) and its activator cathepsin L (CTSL). First, we quantitated the HPSE and CTSL mRNA levels in a cohort of breast cancer patients after curative surgery (20 IBC and 20-non-IBC). We discovered that both HPSE and CTSL mRNA levels were significantly induced in IBC tissue vis-à-vis non-IBC patients (p?<0?.05 and p?<0?.001, respectively). According to the molecular subtypes, HPSE mRNA levels were significantly higher in carcinoma tissues of triple negative (TN)-IBC as compared to TN-non-IBC (p?<0?.05). Mechanistically, we discovered that pharmacological inhibition of HPSE activity resulted in a significant reduction of invasiveness in the IBC SUM149 cell line. Moreover, siRNA-mediated HPSE knockdown significantly downregulated the expression of the metastasis-related gene MMP2 and the cancer stem cell marker CD44. We also found that IBC tumors revealed robust heparanase immune-reactivity and CD163+ M2-type tumor-associated macrophages, with a positive correlation of both markers. Moreover, the secretome of axillary tributaries blood IBC CD14+ monocytes and the cytokine IL-10 significantly upregulated HPSE mRNA and protein expression in SUM149 cells. Intriguingly, massively elevated IL-10 mRNA expression with a trend of positive correlation with HPSE mRNA expression was detected in carcinoma tissue of IBC. Our findings highlight a possible role played by CD14+ monocytes and CD163+ M2-type tumor-associated macrophages in regulating HPSE expression possibly via IL-10. Overall, we suggest that heparanase, cathepsin L and CD14+ monocytes-derived IL-10 may play an important role in the pathogenesis of IBC and their targeting could have therapeutic implications.

Project description:In vivo studies of implanted acellular biological scaffolds in experimental animals have shown constructive remodeling mediated by anti-inflammatory macrophages. Little is known about the human macrophage response to such biomaterials, or the nature of the signaling mechanisms that govern the macrophage phenotype in this environment. The cellular events at the interface of a tissue and implanted decellularized biomaterial were examined by establishing a novel ex vivo tissue culture model in which surgically excised human urinary tract tissue was combined with porcine acellular bladder matrix (PABM). Evaluation of the tissue-biomaterial interface showed a time-dependent infiltration of the biomaterial by CD68(+) CD80(-) macrophages. The migration of CD68(+) cells from the tissue to the interface was accompanied by maturation to a CD163(hi) phenotype, suggesting that factor(s) associated with the biomaterial or the wound edge was/were responsible for the active recruitment and polarization of local macrophages. Glucocorticoid receptor (GR) and peroxisome proliferator activated receptor gamma (PPAR?) signaling was investigated as candidate pathways for integrating inflammatory responses; both showed intense nuclear labeling in interface macrophages. GR and PPAR? activation polarized peripheral blood-derived macrophages from a default M1 (CD80(+)) toward an M2 (CD163(+)) phenotype, but PPAR? signaling predominated, as its antagonism blocked any GR-mediated effect. Seeding on PABM was effective at polarizing peripheral blood-derived macrophages from a default CD80(+) phenotype on glass to a CD80(-) phenotype, with intense nuclear localization of PPAR?. These results endorse in vivo observations that the infiltration of decellularized biological scaffolds, exemplified here by PABM, is pioneered by macrophages. Thus, it appears that natural factors present in PABM are involved in the active recruitment and polarization of macrophages to a CD163(+) phenotype, with activation of PPAR? identified as the candidate pathway. The harnessing of these natural matrix-associated factors may be useful in enhancing the integration of synthetic and other natural biomaterials by polarizing macrophage activation toward an M2 regulatory phenotype.

Project description:BackgroundToll-like receptors (TLRs) and macrophages play an important role in rheumatoid arthritis (RA). Currently, it is not clear whether inflammatory M1 or anti-inflammatory M2 predominate among the resident macrophages in the synovium. In the present study, we set out to investigate the impact of TLR stimulation on monocyte-derived M1 and M2 macrophage function and phenotype by mimicking the exposure to abundant TLR agonists as occurs in the context of RA. The response of macrophage subsets to TLR2 and TLR4 activation was evaluated on cluster of differentiation (CD) marker profile; cytokine secretion; gene expression; and NF-?B, interferon regulatory factors 3 and 7 (IRF3/7), and mitogen-activated protein kinase (MAPK) activation.MethodsHuman monocytes were isolated from peripheral blood of healthy individuals and patients with RA and differentiated into M1-like and M2-like macrophages by granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF), respectively. Cells were either (1) stimulated with TLR ligands Pam3 or lipopolysaccharide (LPS) or (2) classically activated via interferon (IFN)-?/LPS. Cytokine production was measured by enzyme-linked immunosorbent assay, and gene expression was measured by qPCR. Cells were stained for CD markers and analyzed by fluorescence-activated cell sorting. NF-?B, IRF3/7, and MAPKs were detected by Western blotting.ResultsMonocyte-derived macrophages of healthy donors (HD) or patients with RA displayed comparable subset-specific phenotypes upon exposure to TLR agonists. CD14 and CD163 marker expression on M2 macrophages did not change upon TLR2 and TLR4 engagement. By contrast, M2 gene markers HMOX1, FOLR2, and SLC40A1 were decreased. Importantly, M2 macrophages derived from HD or patients with RA showed both a decreased ratio of interleukin (IL)-10/IL-6 and IL-10/IL-8 upon stimulation with TLR2 ligand Pam3 compared with TLR4 ligand LPS. Gene expression of TLR2 was increased, whereas TLR4 expression was decreased, by TLR ligand stimulation. MAPKs p38, extracellular signal-regulated kinase 1/2, and c-Jun N-terminal kinase were activated more strongly in M2 than in M1 macrophages by Pam3 or LPS.ConclusionsWe show that the anti-inflammatory activity of M2 macrophages is reduced in the presence of abundant TLR2 ligands without significant changes in cell surface markers. Thus, the classical M1/M2 paradigm based on cellular markers does not apply to macrophage functions in inflammatory conditions such as RA.

Project description:BackgroundThe anti-inflammatory properties of the cannabinoid 2 receptor (CB2R) in injury and inflammatory diseases have been widely substantiated. Specifically, the anti-inflammatory effect of CB2R may be achieved by regulating macrophage polarisation. Several research findings suggested that the activation of CB2R could attenuate inflammation by reducing pro-inflammatory M1 macrophage polarisation and promoting anti-inflammatory M2 polarisation. However, considering CB2R inhibits fibrosis and M2 promotes fibrosis, that the activation of CB2R may lead to an increase in M2 macrophages seems contradictory. Therefore, we hypothesised that the activation of CB2R to attenuate inflammation is not achieved by up-regulating M2 macrophages.MethodsWe established an incised wound model using mouse skin and used this to evaluate the effect of CB2R agonists (JWH133 or GP1a) and an antagonist (AM630) on wound healing. At various post-injury intervals, we used western blot analysis, immunofluorescence staining, enzyme-linked immunosorbent assay and quantitative reverse transcription polymerase chain reaction assays to determine CB2R protein expression, M1/M2 macrophage infiltration, and the protein and gene expression of M1/M2-associated markers and cytokines in skin lesions.ResultsActivation of CB2R significantly reduced M1 macrophage infiltration and slightly increased M2 macrophage infiltration. Similarly, gene expression and protein levels of M1-associated markers and cytokines (interleukin [IL]-6, IL-12, CD86 and inducible nitric oxide synthase) were significantly down-regulated after CB2R agonist administration; in contrast, markers and cytokines were increased in the CB2R antagonist-treated group. Conversely, the administration of agonists slightly increased gene expression and protein levels of M2-associated markers and cytokines (IL-4, IL-10, CD206 and arginase-1 [Arg-1]); however, a statistical significance at most time points post-injury was not noted.ConclusionIn summary, our findings suggested that during incised skin wound healing in mice, increased levels of CB2R may affect inflammation by regulating M1 rather than M2 macrophage subtype polarisation. These results offer a novel understanding of the molecular mechanisms involved in the inhibition of inflammation by CBR2 that may lead to new treatments for cutaneous inflammation.

Dataset Information

Antibody-Directed Glucocorticoid Targeting to CD163 in M2-type Macrophages Attenuates Fructose-Induced Liver Inflammatory Changes.

Publications

Antibody-Directed Glucocorticoid Targeting to CD163 in M2-type Macrophages Attenuates Fructose-Induced Liver Inflammatory Changes.

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