Conjugated Bilirubin Differentially Regulates CD4+ T Effector Cells and T Regulatory Cell Function through Outside-In and Inside-Out Mechanisms: The Effects of HAV Cell Surface Receptor and Intracellular Signaling.

We recently reported an immune-modulatory role of conjugated bilirubin (CB) in hepatitis A virus (HAV) infection. During this infection the immune response relies on CD4+ T lymphocytes (TLs) and it may be affected by the interaction of HAV with its cellular receptor (HAVCR1/TIM-1) on T cell surface. How CB might affect T cell function during HAV infection remains to be elucidated. Herein, in vitro stimulation of CD4+ TLs from healthy donors with CB resulted in a decrease in the degree of intracellular tyrosine phosphorylation and an increase in the activity of T regulatory cells (Tregs) expressing HAVCR1/TIM-1. A comparison between CD4+ TLs from healthy donors and HAV-infected patients revealed changes in the TCR signaling pathway relative to changes in CB levels. The proportion of CD4+CD25+ TLs increased in patients with low CB serum levels and an increase in the percentage of Tregs expressing HAVCR1/TIM-1 was found in HAV-infected patients relative to controls. A low frequency of 157insMTTTVP insertion in the viral receptor gene HAVCR1/TIM-1 was found in patients and controls. Our data revealed that, during HAV infection, CB differentially regulates CD4+ TLs and Tregs functions by modulating intracellular pathways and by inducing changes in the proportion of Tregs expressing HAVCR1/TIM-1.

T-helper 17-related cytokines and IgE antibodies during hepatitis A virus infection in children.

We determined the serum IgE levels and T-helper (Th)17-related cytokines during distinct hepatitis A virus (HAV)-induced clinical courses in children. A significantly higher concentration of macrophage inflammatory protein 3α, interleukin (IL)-17E and IL-17F in HAV-infected children with intermediate liver injury compared with those with minor liver damage was found. A reduction in the IgE levels in those patients who showed the highest levels of IL-17F in the group of intermediate liver injury was found. The data suggested that the Th17-related profile is associated with the severity of HAV infection and might play a role on the modulation achieved by HAV during allergies.

Conjugated bilirubin affects cytokine profiles in hepatitis A virus infection by modulating function of signal transducer and activator of transcription factors.

Hepatitis A virus (HAV) infection is the major cause of acute liver failure in paediatric patients. The clinical spectrum of infection is variable, and liver injury is determined by altered hepatic enzyme function and bilirubin concentration. We recently reported differences in cytokine profiles between distinct HAV-induced clinical courses, and bilirubin has been recognized as a potential immune-modulator. However, how bilirubin may affect cytokine profiles underlying the variability in the course of infection has not been determined. Herein, we used a transcription factor (TF) binding site identification approach to retrospectively analyse cytokine expression in HAV-infected children and to predict the entire set of TFs associated with the expression of specific cytokine profiles. The results suggested that modulation of the activity of signal transducers and activators of transcription proteins (STATs) may play a central role during HAV infection. This led us to compare the degree of STAT phosphorylation in peripheral blood lymphoid cells (PBLCs) from paediatric patients with distinct levels of conjugated bilirubin (CB). Low CB levels in sera were associated with increased STAT-1 and STAT-5 phosphorylation. A positive correlation was observed between the serum interleukin-6 (IL-6) content and CB values, whereas higher levels of CB correlated with reduced serum IL-8 values and with a reduction in the proportion of PBLCs positive for STAT-5 phosphorylation. When CB was used to stimulate patients' PBLCs in vitro, the levels of IL-6 and tumour necrosis factor-α were increased. The data showed that bilirubin plays a role in STAT function and affects cytokine profile expression during HAV infection.

Inter-facility characterization of bacteria in seafood processing plants: Exploring potential reservoirs of spoilage organisms and the resistome.

A study was conducted in fish processing facilities to investigate the microbial composition, microbial metabolic potential, and distribution of antibiotic resistance genes. Whole metagenomic sequencing was used to analyze microbial communities from different processing rooms, operators and fish products. Taxonomic analyses identified the genera Pseudomonas and Psychrobacter as the most prevalent bacteria. A Principal Component Analysis revealed a distinct separation between fish product and environmental samples, as well as differences between fish product samples from companies processing either Gadidae or Salmonidae fish. Some particular bacterial genera and species were associated with specific processing rooms and operators. Metabolic analysis of metagenome assembled genomes demonstrated variations in microbiota metabolic profiles of microbiota across rooms and fish products. The study also examined the presence of antibiotic-resistance genes in fish processing environments, contributing to the understanding of microbial dynamics, metabolic potential, and implications for fish spoilage.

Grapefruit Juice Flavanones Modulate the Expression of Genes Regulating Inflammation, Cell Interactions and Vascular Function in Peripheral Blood Mononuclear Cells of Postmenopausal Women.

Grapefruit is a rich source of flavanones, phytochemicals suggested excreting vasculoprotective effects. We previously showed that flavanones in grapefruit juice (GFJ) reduced postmenopausal women's pulse-wave velocity (PWV), a measure of arterial stiffness. However, mechanisms of flavanone action in humans are largely unknown. This study aimed to decipher molecular mechanisms of flavanones by multi-omics analysis in PBMCs of volunteers consuming GFJ and flavanone-free control drink for 6 months. Modulated genes and microRNAs (miRNAs) were identified using microarrays. Bioinformatics analyses assessed their functions, interactions and correlations with previously observed changes in PWV. GFJ modified gene and miRNA expressions. Integrated analysis of modulated genes and miRNA-target genes suggests regulation of inflammation, immune response, cell interaction and mobility. Bioinformatics identified putative mediators of the observed nutrigenomic effect (STAT3, NF-κB) and molecular docking demonstrated potential binding of flavanone metabolites to transcription factors and cell-signaling proteins. We also observed 34 significant correlations between changes in gene expression and PWV. Moreover, global gene expression was negatively correlated with gene expression profiles in arterial stiffness and hypertension. This study revealed molecular mechanisms underlying vasculoprotective effects of flavanones, including interactions with transcription factors and gene and miRNA expression changes that inversely correlate with gene expression profiles associated with cardiovascular risk factors. Clinical Trial Registration: [ClinicalTrials.gov], identifier [NCT01272167].

Structurally related (-)-epicatechin metabolites and gut microbiota derived metabolites exert genomic modifications via VEGF signaling pathways in brain microvascular endothelial cells under lipotoxic conditions: Integrated multi-omic study.

Dysfunction of blood-brain barrier formed by endothelial cells of cerebral blood vessels, plays a key role in development of neurodegenerative disorders. Epicatechin exerts vasculo-protective effects through genomic modifications, however molecular mechanisms of action, particularly on brain endothelial cells, are largely unknow. This study aimed to use a multi-omic approach (transcriptomics of mRNA, miRNAs and lncRNAs, and proteomics), to provide novel in-depth insights into molecular mechanisms of how metabolites affect brain endothelial cells under lipid-stressed (as a model of BBB dysfunction) at physiological concentrations. We showed that metabolites can simultaneously modulate expression of protein-coding, non-coding genes and proteins. Integrative analysis revealed interactions between different types of RNAs and form functional groups of genes involved in regulation of processing like VEGF-related functions, cell signaling, cell adhesion and permeability. Molecular modeling of genomics data predicted that metabolites decrease endothelial cell permeability, increased by lipotoxic stress. Correlation analysis between genomic modifications observed and genomic signature of patients with vascular dementia and Alzheimer's diseases showed opposite gene expression changes. Taken together, this study describes for the first time a multi-omic mechanism of action by which (-)-epicatechin metabolites could preserve brain vascular endothelial cell integrity and reduce the risk of neurodegenerative diseases. SIGNIFICANCE: Dysfunction of the blood-brain barrier (BBB), characterized by dysfunction of endothelial cells of cerebral blood vessels, result in an increase in permeability and neuroinflammation which constitute a key factor in the development neurodegenerative disorders. Even though it is suggested that polyphenols can prevent or delay the development of these disorders, their impact on brain endothelial cells and underlying mechanisms of actions are unknow. This study aimed to use a multi-omic approach including analysis of expression of mRNA, microRNA, long non-coding RNAs, and proteins to provide novel global in-depth insights into molecular mechanisms of how (-)-epicatechin metabolites affect brain microvascular endothelial cells under lipid-stressed (as a model of BBB dysfunction) at physiological relevant conditions. The results provide basis of knowledge on the capacity of polyphenols to prevent brain endothelial dysfunction and consequently neurodegenerative disorders.

Anthocyanin-rich extract from black beans exerts anti-diabetic effects in rats through a multi-genomic mode of action in adipose tissue.

Black beans (BB) are an important source of a range of plant bioactive compounds including polyphenols, particularly anthocyanins. Several studies support that consumption of BB is associated with health benefits, including prevention of type 2 diabetes mellitus (T2DM). However, molecular mechanisms underlying the potential health properties of BB on adipose tissue (AT) are still largely unknown. The purpose of this study was to investigate multi-genomic effects of BB intake and identify regulatory networks potentially mediating T2DM on AT. Male Wistar diabetic rats consumed an anthocyanin-rich black bean extract for 5 weeks. Global gene expression from AT, protein coding and non-coding RNA profiles were determined using RNAseq. Biological function analyses were performed using a variety of bioinformatic tools. The evaluation of global gene expression profiles exhibited significant change following BB consumption with 406 significantly differentially expressed genes, 33 miRNA and 39 lncRNA and 3 snRNA. Functional analyses indicated that these genes play an important role in regulation of PI3K signaling, NIN/NF-kB signaling, insulin secretion, and endoplasmic reticulum (ER) organization. Interestingly, transcription factors such as GATA2, or POU2AF1 demonstrated to modulate their activity by BB extract by direct interaction with polyphenol metabolites, or by interactions with cell signaling proteins, like PKB, AKT or PI3K, that could control transcription factor activity and as a result impact on adipogenesis regulation. Therefore, the constant consumption of an anthocyanin-rich black bean extract may have anti-diabetic protective effects by modulating gene expression, resulting in a promising alternative for T2DM patients.

Flavanol Consumption in Healthy Men Preserves Integrity of Immunological-Endothelial Barrier Cell Functions: Nutri(epi)genomic Analysis.

SCOPE: While cocoa flavanol (CF) consumption improves cardiovascular risk biomarkers, molecular mechanisms underlying their protective effects are not understood. OBJECTIVE: To investigate nutri(epi)genomic effects of CF and identify regulatory networks potential mediating vascular health benefits. METHODS AND RESULTS: Twenty healthy middle-aged men consume CF (bi-daily 450 mg) or control drinks for 1 month. Microarray analysis identifies 2235 differentially expressed genes (DEG) involved in processes regulating immune response, cell adhesion, or cytoskeleton organization. Distinct patterns of DEG correlate with CF-related changes in endothelial function, arterial stiffness, and blood pressure. DEG profile negatively correlates with expression profiles of cardiovascular disease patients. CF modulated DNA methylation profile of genes implicates in cell adhesion, actin cytoskeleton organization, or cell signaling. In silico docking analyses indicate that CF metabolites have the potential of binding to cell signaling proteins and transcription factors. Incubation of plasma obtained after CF consumption decrease monocyte to endothelial adhesion and dose-dependently increase nitric oxide-dependent chemotaxis of circulating angiogenic cells further validating the biological functions of CF metabolites. CONCLUSION: In healthy humans, CF consumption may mediate vascular protective effects by modulating gene expression and DNA methylation towards a cardiovascular protective effect, in agreement with clinical results, by preserving integrity of immunological-endothelial barrier functions.

Systematic analysis of nutrigenomic effects of polyphenols related to cardiometabolic health in humans - Evidence from untargeted mRNA and miRNA studies.

Cardiovascular and metabolic disorders present major causes of mortality in the ageing population. Polyphenols present in human diets possess cardiometabolic protective properties, however their underlying molecular mechanisms in humans are still not well identified. Even though preclinical and in vitro studies advocate that these bioactives can modulate gene expression, most studies were performed using targeted approaches. With the objective to decipher the molecular mechanisms underlying polyphenols cardiometabolic preventive properties in humans, we performed integrative multi-omic bioinformatic analyses of published studies which reported improvements of cardiometabolic risk factors following polyphenol intake, together with genomic analyses performed using untargeted approach. We identified 5 studies within our criteria and nearly 5000 differentially expressed genes, both mRNAs and miRNAs, in peripheral blood cells. Integrative bioinformatic analyses (e.g. pathway and gene network analyses, identification of transcription factors, correlation of gene expression profiles with those associated with diseases and drug intake) revealed that these genes are involved in the processes such as cell adhesion and mobility, immune system, metabolism, or cell signaling. We also identified 27 miRNAs known to regulate processes such as cell cytoskeleton, chemotaxis, cell signaling, or cell metabolism. Gene expression profiles negatively correlated with expression profiles of cardiovascular disease patients, while a positive correlation was observed with gene expression profiles following intake of drugs against cardiometabolic disorders. These analyses further advocate for health protective effects of these bioactives against age-associated diseases. In conclusion, polyphenols can exert multi-genomic modifications in humans and use of untargeted methods coupled with bioinformatic analyses represent the best approach to decipher molecular mechanisms underlying healthy-ageing effects of these bioactives.

Molecular and structural characterization of expansins modulated by fungal endophytes in the Antarctic Colobanthus quitensis (Kunth) Bartl. Exposed to drought stress.

Expansins are proteins involved in cell wall metabolism that play an important role in plant growth, development, fruit ripening and abiotic stress tolerance. In the present study, we analyzed putative expansins that respond to drought stress. Five expansin genes were identified in cDNA libraries isolated from Colobanthus quitensis gown either with or without endophytic fungi under hydric stress. A differential transcript abundance was observed by qPCR analysis upon drought stress. To compare these expansin genes, and to suggest a possible mechanism of action at the molecular level, the structural model of the deduced proteins was obtained by comparative modeling methodology. The structures showed two domains and an open groove on the surface of the proteins was observed in the five structural models. The proteins were evaluated in terms of their protein-ligand interactions using four different ligands. The results suggested differences in their mode of protein-ligand interaction, in particular concerning the residues involved in the protein-ligand interaction. The presented evidence supports the participation of some members of the expansin multiprotein family in the response to drought stress in C. quitensis and suggest that the response is modulated by endophytic fungi.

Modeling the Th17 and Tregs Paradigm: Implications for Cancer Immunotherapy.

CD4 + T cell differentiation is governed by gene regulatory and metabolic networks, with both networks being highly interconnected and able to adapt to external stimuli. Th17 and Tregs differentiation networks play a critical role in cancer, and their balance is affected by the tumor microenvironment (TME). Factors from the TME mediate recruitment and expansion of Th17 cells, but these cells can act with pro or anti-tumor immunity. Tregs cells are also involved in tumor development and progression by inhibiting antitumor immunity and promoting immunoevasion. Due to the complexity of the underlying molecular pathways, the modeling of biological systems has emerged as a promising solution for better understanding both CD4 + T cell differentiation and cancer cell behavior. In this review, we present a context-dependent vision of CD4 + T cell transcriptomic and metabolic network adaptability. We then discuss CD4 + T cell knowledge-based models to extract the regulatory elements of Th17 and Tregs differentiation in multiple CD4 + T cell levels. We highlight the importance of complementing these models with data from omics technologies such as transcriptomics and metabolomics, in order to better delineate existing Th17 and Tregs bifurcation mechanisms. We were able to recompilate promising regulatory components and mechanisms of Th17 and Tregs differentiation under normal conditions, which we then connected with biological evidence in the context of the TME to better understand CD4 + T cell behavior in cancer. From the integration of mechanistic models with omics data, the transcriptomic and metabolomic reprograming of Th17 and Tregs cells can be predicted in new models with potential clinical applications, with special relevance to cancer immunotherapy.

Interplay between SMAD2 and STAT5A is a critical determinant of IL-17A/IL-17F differential expression.

Interleukins (IL)-17A and F are critical cytokines in anti-microbial immunity but also contribute to auto-immune pathologies. Recent evidence suggests that they may be differentially produced by T-helper (Th) cells, but the underlying mechanisms remain unknown. To address this question, we built a regulatory graph integrating all reported upstream regulators of IL-17A and F, completed by ChIP-seq data analyses. The resulting regulatory graph encompasses 82 components and 136 regulatory links. The graph was then supplemented by logical rules calibrated with original flow cytometry data using naive CD4+ T cells, in conditions inducing IL-17A or IL-17F. The model displays specific stable states corresponding to virtual phenotypes explaining IL-17A and IL-17F differential regulation across eight cytokine stimulatory conditions. Our model analysis points to the transcription factors NFAT2A, STAT5A and SMAD2 as key regulators of the differential expression of IL-17A and IL-17F, with STAT5A controlling IL-17F expression, and an interplay of NFAT2A, STAT5A and SMAD2 controlling IL-17A expression. We experimentally observed that the production of IL-17A was correlated with an increase of SMAD2 transcription, and the expression of IL-17F correlated with an increase of BLIMP-1 transcription, together with an increase of STAT5A expression (mRNA), as predicted by our model. Interestingly, RORγt presumably plays a more determinant role in IL-17A expression as compared to IL-17F expression. In conclusion, we propose the first mechanistic model accounting for the differential expression of IL-17A and F in Th cells, providing a basis to design novel therapeutic interventions in auto-immune and inflammatory diseases.

Integrated Multi-Omic Analyses of the Genomic Modifications by Gut Microbiome-Derived Metabolites of Epicatechin, 5-(4'-Hydroxyphenyl)-γ-Valerolactone, in TNFalpha-Stimulated Primary Human Brain Microvascular Endothelial Cells.

Cerebral blood vessels are lined with endothelial cells and form the blood-brain barrier. Their dysfunction constitutes a crucial event in the physiopathology of neurodegenerative disorders and cognitive impairment. Epicatechin can improve cognitive functions and lower the risk for Alzheimer's disease or stroke. However, molecular mechanisms of epicatechin on brain vascular endothelium are still unexplored. The objective of this study was to investigate the biological effects of gut microbiome-derived metabolites of epicatechin, 5-(4'-Hydroxyphenyl)-γ-valerolactone-3'-sulfate and 5-(4'-Hydroxyphenyl)-γ-valerolactone-3'-O-glucuronide, in TNF-α-stimulated human brain microvascular endothelial cells at low (nM) concentrations by evaluating their multi-omic modification (expression of mRNA, microRNA, long non-coding RNAs, and proteins). We observed that metabolites are biologically active and can simultaneously modulate the expression of protein-coding and non-coding genes as well as proteins. Integrative bioinformatics analysis of obtained data revealed complex networks of genomics modifications by acting at different levels of regulation. Metabolites modulate cellular pathways including cell adhesion, cytoskeleton organization, focal adhesion, signaling pathways, pathways regulating endothelial permeability, and interaction with immune cells. This study demonstrates multimodal mechanisms of action by which epicatechin metabolites could preserve brain vascular endothelial cell integrity, presenting mechanisms of action underlying epicatechin neuroprotective properties.

Molecular Determinants of the Cardiometabolic Improvements of Dietary Flavanols Identified by an Integrative Analysis of Nutrigenomic Data from a Systematic Review of Animal Studies.

SCOPE: Flavanols are important polyphenols of the human diet with extensive demonstrations of their beneficial effects on cardiometabolic health. They contribute to preserve health acting on a large range of cellular processes. The underlying mechanisms of action of flavanols are not fully understood but involve a nutrigenomic regulation. METHODS AND RESULTS: To further capture how the intake of dietary flavanols results in the modulation of gene expression, nutrigenomics data in response to dietary flavanols obtained from animal models of cardiometabolic diseases have been collected and submitted to a bioinformatics analysis. This systematic analysis shows that dietary flavanols modulate a large range of genes mainly involved in endocrine function, fatty acid metabolism, and inflammation. Several regulators of the gene expression have been predicted and include transcription factors, miRNAs and epigenetic factors. CONCLUSION: This review highlights the complex and multilevel action of dietary flavanols contributing to their strong potential to preserve cardiometabolic health. The identification of the potential molecular mediators and of the flavanol metabolites driving the nutrigenomic response in the target organs is still a pending question which the answer will contribute to optimize the beneficial health effects of dietary bioactives.

Systematic Bioinformatic Analyses of Nutrigenomic Modifications by Polyphenols Associated with Cardiometabolic Health in Humans-Evidence from Targeted Nutrigenomic Studies.

Cardiometabolic disorders are among the leading causes of mortality in the human population. Dietary polyphenols exert beneficial effects on cardiometabolic health in humans. Molecular mechanisms, however, are not completely understood. Aiming to conduct in-depth integrative bioinformatic analyses to elucidate molecular mechanisms underlying the protective effects of polyphenols on cardiometabolic health, we first conducted a systematic literature search to identify human intervention studies with polyphenols that demonstrate improvement of cardiometabolic risk factors in parallel with significant nutrigenomic effects. Applying the predefined inclusion criteria, we identified 58 differentially expressed genes at mRNA level and 5 miRNAs, analyzed in peripheral blood cells with RT-PCR methods. Subsequent integrative bioinformatic analyses demonstrated that polyphenols modulate genes that are mainly involved in the processes such as inflammation, lipid metabolism, and endothelial function. We also identified 37 transcription factors that are involved in the regulation of polyphenol modulated genes, including RELA/NFKB1, STAT1, JUN, or SIRT1. Integrative bioinformatic analysis of mRNA and miRNA-target pathways demonstrated several common enriched pathways that include MAPK signaling pathway, TNF signaling pathway, PI3K-Akt signaling pathway, focal adhesion, or PPAR signaling pathway. These bioinformatic analyses represent a valuable source of information for the identification of molecular mechanisms underlying the beneficial health effects of polyphenols and potential target genes for future nutrigenetic studies.

An Integrated Analysis of miRNA and Gene Expression Changes in Response to an Obesogenic Diet to Explore the Impact of Transgenerational Supplementation with Omega 3 Fatty Acids.

Insulin resistance decreases the ability of insulin to inhibit hepatic gluconeogenesis, a key step in the development of metabolic syndrome. Metabolic alterations, fat accumulation, and fibrosis in the liver are closely related and contribute to the progression of comorbidities, such as hypertension, type 2 diabetes, or cancer. Omega 3 (n-3) polyunsaturated fatty acids, such as eicosapentaenoic acid (EPA), were identified as potent positive regulators of insulin sensitivity in vitro and in animal models. In the current study, we explored the effects of a transgenerational supplementation with EPA in mice exposed to an obesogenic diet on the regulation of microRNAs (miRNAs) and gene expression in the liver using high-throughput techniques. We implemented a comprehensive molecular systems biology approach, combining statistical tools, such as MicroRNA Master Regulator Analysis pipeline and Boolean modeling to integrate these biochemical processes. We demonstrated that EPA mediated molecular adaptations, leading to the inhibition of miR-34a-5p, a negative regulator of Irs2 as a master regulatory event leading to the inhibition of gluconeogenesis by insulin during the fasting-feeding transition. Omics data integration provided greater biological insight and a better understanding of the relationships between biological variables. Such an approach may be useful for deriving innovative data-driven hypotheses and for the discovery of molecular-biochemical mechanistic links.

Molecular Insights into FaEG1, a Strawberry Endoglucanase Enzyme Expressed during Strawberry Fruit Ripening.

The endo-ß-1,4-glucanases (EGs) that belong to the glycosyl hydrolase family 9 (GH9) have roles in cell wall synthesis, remodeling and degradation. Previous studies have suggested that EGs may play a key role in the ripening of different fruits including strawberries. In this study, we used reverse-transcription quantitative polymerase chain reaction (RT-qPCR) assays to determine the transcript accumulation of an endo-ß-1,4-glucanase (FaEG1) during fruit development in two different strawberry 'Camarosa' and 'Monterey' with contrasting softening ratios. Phylogenetic analyses suggest that FaEG1 belongs to the α group of the GH9 family with other proteins previously described with roles in elongation, abscission and ripening. Comparative modeling was used to obtain the FaEG1 structure. The model displays a α-barrel-type structure that is typical of the GH9 enzyme family, and comprises 12 α-helices, 2 310 helices and 6 ß-sheets. The catalytic residues were oriented to the solvent in the middle of an open groove. Protein-ligand interactions were explored with cellulose and two xyloglucans as ligands; the results suggest that the FaEG1-cellulose and FaEG1-XXXGXXXG (the most abundant xyloglucan in strawberries) complexes were more stable complexes than XXFGXXFG. The cell wall degradation was observed by scanning electron microscopy (SEM). The data are congruent with the probable role of the FaEG1 protein in the dissembly of the cellulose-hemicellulose fraction during the ripening of strawberry fruit.

Conjugated Bilirubin Upregulates TIM-3 Expression on CD4+CD25+ T Cells: Anti-Inflammatory Implications for Hepatitis A Virus Infection.

Bilirubin (BR), a metabolite with increased concentrations in plasma during viral hepatitis, has been recognized as a potential immune-modulator. We recently reported that conjugated BR (CB) augments regulatory T cell (Treg) suppressor activity during acute hepatitis A virus (HAV) infection. However, the mechanisms related to the effects of CB on Treg function in the course of hepatotropic viral diseases have not been elucidated. T cell immunoglobulin domain and mucin domain 3 (TIM-3), via its interactions with galectin-9 (GAL-9), is a receptor associated with enhanced Treg function. Thus, TIM-3 expression may be related to the crosstalk between CB and Tregs during HAV infection. Herein, in vitro treatment with high concentrations of CB upregulated TIM-3 expression on Tregs from healthy donors. CB treatment in vitro did not induce de novo Treg generation, and in vitro stimulation with TGF-ß, which shows increased secretion during HAV infection, resulted in a trend toward increased TIM-3 expression on Tregs and CD4+ T lymphocytes (TLs) from healthy donors. Interestingly, an upregulation of TIM-3 expression on CD4+CD25+ T cells and an increase in the proportion of CD4+ TLs expressing GAL-9 were found in HAV-infected patients with abnormal CB values relative to healthy controls. In addition, a statistically significantly reduction in IL-17F production was observed after treatment of CD4+ TLs from healthy donors with high doses of CB in vitro. In summary, our results suggest that CB might regulate Treg activity via a TIM-3-mediated mechanism, ultimately leading to an anti-inflammatory hepatoprotective effect.

Rethinking the immune properties of bilirubin in viral hepatitis: from bench to bedside.

Communication between the immune system and metabolic components can be exemplified by the process of heme catabolism. The immunomodulatory functions of the enzymes, substrates and active products related to catabolism of the heme group have been extensively studied. Bilirubin (BR), the final breakdown product of heme, is primarily considered to be a toxic waste product but has recently been considered to be an immunomodulatory metabolite. Through mechanisms that include intracellular signaling and transcriptional control, BR affects those immune cell functions that regulate cell proliferation, differentiation and apoptosis. During the pathogenesis of viral hepatitis, the heme degradation pathway is disrupted, resulting in changes to normal BR concentrations. These alterations have been previously studied mainly as a consequence of the infection. However, little is known about the potential immunomodulatory role played by BR in the development of infectious hepatocellular diseases. Differences in BR levels in the context of viral hepatitis are likely to provide important insights into the metabolite-mediated mechanisms controlling the immune responses underlying both the long-term persistence of hepatitis C virus (HCV) infection and the resolution of hepatitis A virus (HAV) infection during the acute phase. In this review, the cross-talk between heme catabolism and immune function is described in detail. Special emphasis is given to discoveries that hold promise for identifying immunologic features of metabolic products in the resolution of viral diseases.