A decrease in the abundance and biodiversity of intestinal bacteria within the Firmicutes phylum has been associated with inflammatory bowel disease (IBD). In particular, the anti-inflammatory ...bacterium Faecalibacterium prausnitzii, member of the Firmicutes phylum and one of the most abundant species in healthy human colon, is underrepresented in the microbiota of IBD patients. The aim of this study was to investigate the immunomodulatory properties of F. prausnitzii strain A2-165, the biofilm forming strain HTF-F and the extracellular polymeric matrix (EPM) isolated from strain HTF-F. For this purpose, the immunomodulatory properties of the F. prausnitzii strains and the EPM were studied in vitro using human monocyte-derived dendritic cells. Then, the capacity of the F. prausnitzii strains and the EPM of HTF-F to suppress inflammation was assessed in vivo in the mouse dextran sodium sulphate (DSS) colitis model. The F. prausnitzii strains and the EPM had anti-inflammatory effects on the clinical parameters measured in the DSS model but with different efficacy. The immunomodulatory effects of the EPM were mediated through the TLR2-dependent modulation of IL-12 and IL-10 cytokine production in antigen presenting cells, suggesting that it contributes to the anti-inflammatory potency of F. prausnitzii HTF-F. The results show that F. prausnitzii HTF-F and its EPM may have a therapeutic use in IBD.
Dietary protein sources can have profound effects on host-microbe interactions in the gut that are critically important for immune resilience. However more knowledge is needed to assess the impact of ...different protein sources on gut and animal health. Thirty-six wildtype male C57BL/6J mice of 35 d age (n = 6/group; mean ± SEM body weight 21.9 ± 0.25 g) were randomly assigned to groups fed for four weeks with semi synthetic diets prepared with one of the following protein sources containing (300 g/kg as fed basis): soybean meal (SBM), casein, partially delactosed whey powder, spray dried plasma protein, wheat gluten meal and yellow meal worm. At the end of the experiment, mice were sacrificed to collect ileal tissue to acquire gene expression data, and mammalian (mechanistic) target of rapamycin (mTOR) activity, ileal digesta to study changes in microbiota and serum to measure cytokines and chemokines. By genome-wide transcriptome analysis, we identified fourteen high level regulatory genes that are strongly affected in SBM-fed mice compared to the other experimental groups. They mostly related to the mTOR pathway. In addition, an increased (P < 0.05) concentration of granulocyte colony-stimulating factor was observed in serum of SBM-fed mice compared to other dietary groups. Moreover, by 16S rRNA sequencing, we observed that SBM-fed mice had higher (P < 0.05) abundances of Bacteroidales family S24-7, compared to the other dietary groups. We showed that measurements of genome-wide expression and microbiota composition in the mouse ileum reveal divergent responses to diets containing different protein sources, in particular for a diet based on SBM.
The immunomodulatory properties of non-digestible polysaccharides (NDPs) have been recognized in in vitro and in vivo studies. The latter mostly demonstrated altered frequencies and inflammatory ...status of immune cells as clinical parameters. Most of the NDP activity will be exerted in the intestine where they can directly interact with macrophages. The predominant macrophage phenotype in the intestine is M2-like, with M1-like macrophages arising during inflammation. Here, we investigated transcriptional and functional impact on these macrophage phenotypes by NDP-treatment (i.e. yeast-derived soluble β-glucan (yeast-βG), apple-derived RG-I (apple-RGI), shiitake-derived β-glucan (shiitake-βG) or wheat-derived arabinoxylan (wheat-AX)). Wheat-AX, and to a lesser extent shiitake-βG and apple-RGI but not yeast-βG, reduced endocytosis and antigen processing capacity of M1- and M2-like macrophages. Moreover, the NDPs, and most notably wheat-AX, strongly induced transcription and secretion of a unique set of cytokines and chemokines. Conditioned medium from wheat-AX-treated M2-like macrophages subsequently demonstrated strongly increased monocyte recruitment capacity. These findings are in line with clinically observed immunomodulatory aspects of NDPs making it tempting to speculate that clinical activity of some NDPs is mediated through enhanced chemoattraction and modifying activity of intestinal immune cells.
Lactobacillus mucosae DPC 6426 has previously demonstrated potentially cardio-protective properties, in the form of dyslipidaemia and hypercholesterolemia correction in an apolipoprotein-E deficient ...mouse model. This study aims to characterise the manner in which this microbe may modulate host bile pool composition and immune response, in the context of cardiovascular disease. Lactobacillus mucosae DPC 6426 was assessed for bile salt hydrolase activity and specificity. The microbe was compared against several other enteric strains of the same species, as well as a confirmed bile salt hydrolase-active strain, Lactobacillus reuteri APC 2587.
Quantitative bile salt hydrolase assays revealed that enzymatic extracts from Lactobacillus reuteri APC 2587 and Lactobacillus mucosae DPC 6426 demonstrate the greatest activity in vitro. Bile acid profiling of porcine and murine bile following incubation with Lactobacillus mucosae DPC 6426 confirmed a preference for hydrolysis of glyco-conjugated bile acids. In addition, the purified exopolysaccharide and secretome of Lactobacillus mucosae DPC 6426 were investigated for immunomodulatory capabilities using RAW264.7 macrophages. Gene expression data revealed that both fractions stimulated increases in interleukin-6 and interleukin-10 gene transcription in the murine macrophages, while the entire secretome was necessary to increase CD206 transcription. Moreover, the exopolysaccharide elicited a dose-dependent increase in nitric oxide and interleukin-10 production from RAW264.7 macrophages, concurrent with increased tumour necrosis factor-α secretion at all doses.
This study indicates that Lactobacillus mucosae DPC 6426 modulates both bile pool composition and immune system tone in a manner which may contribute significantly to the previously identified cardio-protective phenotype.
In higher vertebrates, mineralo- (aldosterone) and glucocorticoids (cortisol/corticosterone) exert their multiple actions via specific transcription factors, glucocorticoid (GR) and mineralocorticoid ...(MR) receptors. Teleostean fishes lack aldosterone and mineral regulatory processes seem under dominant control by cortisol. Despite the absence of the classical mineralocorticoid aldosterone, teleostean fishes do have an MR with cortisol and possibly 11-deoxycorticosterone (DOC) (as alternative for aldosterone) as predominant ligands. We studied corticoid receptors in common carp (Cyprinus carpio L). Through homology cloning and bioinformatic analysis, we found duplicated GR genes and a single MR gene. The GR genes likely result from a major genomic duplication event in the teleostean lineage; we propose that the gene for a second MR was lost. Transactivation studies show that the carp GRs and MR have comparable affinity for cortisol; the MR has significantly higher sensitivity to DOC, and this favours a role for DOC as MR ligand in fish physiology. mRNA of the GRs and the MR is expressed in forebrain (in pallial areas homologous to mammalian hippocampus), corticotrophin-releasing hormone (CRH) cells in the pre-optic nucleus (NPO) and pituitary pars distalis ACTH cells, three key neural/endocrine components of the stress axis. After exposure to prolonged and strong (not to mild acute) stressors, mRNA levels of both GRs and MR become down-regulated in the brain, but not in the NPO CRH cells or pituitary ACTH cells. Our data predicts a function in stress physiology for all CRs and suggest telencephalon as a first line cortisol target in stress.
Glucocorticoids (GCs) are commonly used to treat a variety of immune diseases. However, the efficacy of treatment is greatly influenced by an individual variation in sensitivity to GCs, which is ...caused by differences in the glucocorticoid receptor (GR). The variable receptor profile results from variations in the GR gene, or alternative splicing of the gene coded. We investigated the evolution of the GR gene by comparing genomic GR sequences of vertebrates. Exon length and amino acid sequence are conserved among all classes of vertebrates studied, which indicates strong evolutionary pressure on conservation of this gene. Interestingly, teleostean fishes have two different GR proteins. One of the duplicate fish GR genes has a nine-amino-acid insert in the DNA binding region that results from alternative splicing. The duplicate GR genes and products of alternative splicing in teleostean fishes are differentially expressed in vivo and show different transactivation capacity in vitro. The presence of two GR genes appears to be a result of divergence of receptors rather than of ligands. Teleostean fishes express different, evolutionarily related, functional GR proteins within a single organism. Hereby, teleostean fishes present a model that facilitates investigation of the molecular basis of cortisol resistance and different regulatory functions of cortisol.
► Soluble and cell-expressed LAIR-1 binds to transmembrane collagens expressed on tumor cells. ► Knock-down of prolyl 4-hydroxylase diminishes LAIR-1 binding to tumor cells. ► LAIR-1 is activated by ...transmembrane and extracellular collagens produced by tumor cells. ► Collagen XVII expression by target cells inhibits NK cell cytotoxicity.
Many tumor types over-express collagens, what correlates with enhanced metastatic capacity and unfavorable clinical outcome. This is generally explained by the importance of collagens in creating a microenvironment that supports tumor cell survival and enhances cell migration. Importantly, collagens act as ligands for the inhibitory receptor LAIR-1, which inhibits the function of multiple types of immune cells. Here we propose a new role for tumor expressed collagens and show that these structural proteins can be exploited by tumor cells to inhibit immune responses through an interaction with LAIR-1.
We show that both LAIR-1-Fc fusion proteins and LAIR-1 expressing cells bind to transmembrane collagens expressed by tumor cells. Interference with collagen expression by specific knock-down of prolyl 4-hydroxylase diminishes LAIR-1 binding to tumor cells, demonstrating the specificity of the interaction. Consistently, both transmembrane collagens and extracellular collagens produced by multiple tumor cell types can activate LAIR-1. Furthermore, overexpression of collagen XVII on target cells results in diminished NK cell cytotoxic activity. Thus tumor-expressed collagens can bind and trigger immune inhibitory signaling via LAIR-1, suggesting that collagens indeed may affect tumor immune evasion.
Muc2-deficient mice show no signs of ileal pathology but the mechanisms remained unknown.
Wild-type (WT), Muc2, and Muc2 mice were killed at 2, 4, and 8 weeks of age. Total RNA from ileum was used ...for full genome transcriptome analysis and qPCR. Microbiota composition was determined using a mouse intestinal chip (MITChip). Morphological and immunohistological studies were performed on segments of ileum.
The ileum was colonized by more diverse microbiota in young (week 4) WT than in Muc2 mice, and composition was influenced by genotype. Weaning was associated with major changes in the transcriptome of all mice, and the highest number of differentially expressed genes compared with adults, reflecting temporal changes in microbiota. Although the spatial compartmentalization of bacteria was compromised in Muc2 mice, gene set enrichment analysis revealed a downregulation of Toll-like receptor, immune, and chemokine signaling pathways compared to WT mice. The predicted effects of enhanced IL-22 signaling were identified in the Muc2 transcriptome as the upregulation of epithelial cell proliferation altered expression of mitosis and cell-cycle control pathways. This is consistent with increased villus length and number of Ki67 epithelial cells in Muc2 mice. Additionally, expression of the network of IL-22 regulated defense genes, including Fut2, Reg3β, Reg3γ, Relmb, and the Defensin Defb46 were increased in Muc2 mice.
These findings highlight a role for the IL-22-STAT3 pathway in maintaining ileal homeostasis when the mucus barrier is compromised and its potential as a target for novel therapeutic strategies in inflammatory bowel disease.
The stress hormone cortisol is deeply involved in immune regulation in all vertebrates. Common carp (
Cyprinus carpio L.) express four corticoid receptors that may modulate immune responses: three ...glucocorticoid receptors (GR); GR1, with two splice variants (GR1a and GR1b), GR2 and a single mineralocorticoid receptor (MR). All receptors are expressed as of 4 days post-fertilization and may thus play a critical role in development and functioning of the adult immune system. Immune tissues and cells predominantly express mRNA for GRs compared to mRNA for the MR. Three-dimensional protein structure modeling predicts, and transfection assays confirm that alternative splicing of GR1 does not influence the capacity to induce transcription of effector genes. When tested for cortisol activation, GR2 is the most sensitive corticoid receptor in carp, followed by the MR and GR1a and GR1b. Lipopolysacharide (LPS) treatment of head kidney phagocytes quickly induces GR1 expression and inhibits GR2 expression. Cortisol treatment
in vivo enhances GR1a and MR mRNA expression, but only mildly, and cortisol treatment
in vitro does not affect receptor expression of phagocytes. Cortisol has no direct effect on the LPS-induced receptor profile. Therefore, an immune rather than a stress stimulus regulates GR expression. Cortisol administered at stress levels to phagocytes
in vitro significantly inhibits LPS-induced expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin-12 (IL-12) (subunit p35) and of inducible nitric oxide synthase (
iNOS) expression. A physiologically differential function for GR1 and GR2 in the immune response of fish to infection is indicated.
Leptin is a key factor in the regulation of food intake and is an important factor in the pathophysiology of obesity. However, more than a decade after the discovery of leptin in mouse, information ...regarding leptin in any nonmammalian species is still scant. We report the identification of duplicate leptin genes in common carp (Cyprinus carpio). The unique gene structure, the conservation of both cysteines that form leptin’s single disulfide bridge, and stable clustering in phylogenetic analyses substantiate the unambiguous orthology of mammalian and carp leptins, despite low amino acid identity. The liver is a major yet not the only site of leptin expression. However, neither 6 d nor 6 wk of fasting nor subsequent refeeding affected hepatic leptin expression, although the carp predictably shifted from carbohydrate to lipid metabolism. Animals that were fed to satiation grew twice as fast as controls; however, they did not show increased leptin expression at the termination of the study. Hepatic leptin expression did, however, display an acute and transient postprandial increase that follows the postprandial plasma glucose peak. In summary, leptin mRNA expression in carp changes acutely after food intake, but involvement of leptin in the long-term regulation of food intake and energy metabolism was not evident from fasting for days or weeks or long-term feeding to satiation. These are the first data on the regulation of leptin expression in any nonmammalian species.
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