ABSTRACT
Multidimensional single-cell analyses of T cells have fueled the debate about whether there is extensive plasticity or 'mixed' priming of helper T cell subsets in vivo. Here, we developed an experimental framework to probe the idea that the site of priming in the systemic immune compartment is a determinant of helper T cell-induced immunopathology in remote organs. By site-specific in vivo labeling of antigen-specific T cells in inguinal (i) or gut draining mesenteric (m) lymph nodes, we show that i-T cells and m-T cells isolated from the inflamed central nervous system (CNS) in a model of multiple sclerosis (MS) are distinct. i-T cells were Cxcr6+, and m-T cells expressed P2rx7. Notably, m-T cells infiltrated white matter, while i-T cells were also recruited to gray matter. Therefore, we propose that the definition of helper T cell subsets by their site of priming may guide an advanced understanding of helper T cell biology in health and disease.
Subject(s)
Autoimmunity , Brain/immunology , Cell Lineage , Encephalomyelitis, Autoimmune, Experimental/immunology , Intestines/immunology , Skin/immunology , T-Lymphocytes, Helper-Inducer/immunology , Adoptive Transfer , Animals , Autoimmunity/drug effects , Brain/drug effects , Brain/metabolism , Calcium Signaling , Cerebrospinal Fluid/immunology , Cerebrospinal Fluid/metabolism , Encephalomyelitis, Autoimmune, Experimental/drug therapy , Encephalomyelitis, Autoimmune, Experimental/genetics , Encephalomyelitis, Autoimmune, Experimental/metabolism , Fingolimod Hydrochloride/pharmacology , Gene Expression Profiling , Genes, T-Cell Receptor , HEK293 Cells , Humans , Immunosuppressive Agents/pharmacology , Intestines/drug effects , Intravital Microscopy , Mice, Inbred C57BL , Mice, Knockout , Microscopy, Fluorescence , Multiple Sclerosis, Relapsing-Remitting/genetics , Multiple Sclerosis, Relapsing-Remitting/immunology , Multiple Sclerosis, Relapsing-Remitting/metabolism , Phenotype , Prospective Studies , RNA-Seq , Receptors, CXCR6/genetics , Receptors, CXCR6/metabolism , Receptors, Purinergic P2X7/genetics , Receptors, Purinergic P2X7/metabolism , Single-Cell Analysis , Skin/drug effects , Skin/metabolism , T-Lymphocytes, Helper-Inducer/drug effects , T-Lymphocytes, Helper-Inducer/metabolism , T-Lymphocytes, Helper-Inducer/transplantation , TranscriptomeABSTRACT
Clostridium difficile infection (CDI) is facilitated by alteration of the microbiome following antibiotic administration. Antimicrobial therapy directed against the pathogen can treat CDI. Unfortunately, â¼20% of successfully treated patients will suffer recurrence. Bezlotoxumab, a human monoclonal antibody, binds to C. difficile toxin B (TcdB), reducing recurrence presumably by limiting epithelial damage and facilitating microbiome recovery.
Subject(s)
Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing/therapeutic use , Clostridioides difficile/physiology , Enterocolitis, Pseudomembranous/drug therapy , Broadly Neutralizing Antibodies , Gastrointestinal Microbiome , Humans , Intestines/drug effects , Secondary PreventionABSTRACT
The Th17 cell-lineage-defining cytokine IL-17A contributes to host defense and inflammatory disease by coordinating multicellular immune responses. The IL-17 receptor (IL-17RA) is expressed by diverse intestinal cell types, and therapies targeting IL-17A induce adverse intestinal events, suggesting additional tissue-specific functions. Here, we used multiple conditional deletion models to identify a role for IL-17A in secretory epithelial cell differentiation in the gut. Paneth, tuft, goblet, and enteroendocrine cell numbers were dependent on IL-17A-mediated induction of the transcription factor ATOH1 in Lgr5+ intestinal epithelial stem cells. Although dispensable at steady state, IL-17RA signaling in ATOH1+ cells was required to regenerate secretory cells following injury. Finally, IL-17A stimulation of human-derived intestinal organoids that were locked into a cystic immature state induced ATOH1 expression and rescued secretory cell differentiation. Our data suggest that the cross talk between immune cells and stem cells regulates secretory cell lineage commitment and the integrity of the mucosa.
Subject(s)
Basic Helix-Loop-Helix Transcription Factors/metabolism , Intestinal Mucosa/cytology , Receptors, G-Protein-Coupled/metabolism , Receptors, Interleukin-17/metabolism , Stem Cells/metabolism , Animals , Cell Communication , Cell Differentiation/drug effects , Cell Lineage/drug effects , Colitis/chemically induced , Colitis/metabolism , Colitis/pathology , Dextran Sulfate/adverse effects , Humans , Interleukin-17/metabolism , Interleukin-17/pharmacology , Intestinal Mucosa/metabolism , Intestines/drug effects , Intestines/metabolism , Intestines/pathology , Mice , Mice, Knockout , NF-kappa B/metabolism , Receptors, Interleukin-17/deficiency , SOX9 Transcription Factor/metabolism , Signal Transduction , Stem Cells/cytologyABSTRACT
Changes in the gut microbiome have pivotal roles in the pathogenesis of acute graft-versus-host disease (aGVHD) after allogenic haematopoietic cell transplantation (allo-HCT)1-6. However, effective methods for safely resolving gut dysbiosis have not yet been established. An expansion of the pathogen Enterococcus faecalis in the intestine, associated with dysbiosis, has been shown to be a risk factor for aGVHD7-10. Here we analyse the intestinal microbiome of patients with allo-HCT, and find that E. faecalis escapes elimination and proliferates in the intestine by forming biofilms, rather than by acquiring drug-resistance genes. We isolated cytolysin-positive highly pathogenic E. faecalis from faecal samples and identified an anti-E. faecalis enzyme derived from E. faecalis-specific bacteriophages by analysing bacterial whole-genome sequencing data. The antibacterial enzyme had lytic activity against the biofilm of E. faecalis in vitro and in vivo. Furthermore, in aGVHD-induced gnotobiotic mice that were colonized with E. faecalis or with patient faecal samples characterized by the domination of Enterococcus, levels of intestinal cytolysin-positive E. faecalis were decreased and survival was significantly increased in the group that was treated with the E. faecalis-specific enzyme, compared with controls. Thus, administration of a phage-derived antibacterial enzyme that is specific to biofilm-forming pathogenic E. faecalis-which is difficult to eliminate with existing antibiotics-might provide an approach to protect against aGVHD.
Subject(s)
Bacteriophages , Enterococcus faecalis , Gastrointestinal Microbiome , Graft vs Host Disease , Adult , Aged , Animals , Female , Humans , Male , Mice , Middle Aged , Young Adult , Bacteriophages/enzymology , Bacteriophages/genetics , Biofilms/drug effects , Biofilms/growth & development , Dysbiosis/complications , Dysbiosis/microbiology , Enterococcus faecalis/drug effects , Enterococcus faecalis/genetics , Enterococcus faecalis/growth & development , Enterococcus faecalis/metabolism , Enterococcus faecalis/virology , Feces/microbiology , Germ-Free Life , Graft vs Host Disease/complications , Graft vs Host Disease/microbiology , Graft vs Host Disease/prevention & control , Graft vs Host Disease/therapy , Hematopoietic Stem Cell Transplantation/adverse effects , In Vitro Techniques , Intestines/drug effects , Intestines/microbiology , Perforin/metabolism , Risk Factors , Transplantation, Homologous/adverse effects , Whole Genome Sequencing , Drug Resistance, Bacterial/drug effects , Anti-Bacterial Agents/pharmacologyABSTRACT
Intestinal fibrosis is considered an inevitable complication of Crohn's disease (CD) that results in symptoms of obstruction and stricture formation. Endoscopic or surgical treatment is required to treat the majority of patients. Progress in the management of stricturing CD is hampered by the lack of effective antifibrotic therapy; however, this situation is likely to change because of recent advances in other fibrotic diseases of the lung, liver, and skin. In this review, we summarize data from randomized controlled trials (RCTs) of antifibrotic therapies in these conditions. Multiple compounds have been tested for antifibrotic effects in other organs. According to their mechanisms, they were categorized into growth factor modulators, inflammation modulators, 5-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors, intracellular enzymes and kinases, renin-angiotensin system (RAS) modulators, and others. From our review of the results from the clinical trials and discussion of their implications in the gastrointestinal tract, we have identified several molecular candidates that could serve as potential therapies for intestinal fibrosis in CD.
Subject(s)
Constriction, Pathologic/drug therapy , Crohn Disease/drug therapy , Inflammation/drug therapy , Randomized Controlled Trials as Topic , Constriction, Pathologic/diagnosis , Crohn Disease/diagnosis , Fibrosis/drug therapy , Humans , Inflammation/pathology , Intestines/drug effects , Intestines/pathologyABSTRACT
Host microbial cross-talk is essential to maintain intestinal homeostasis. However, maladaptation of this response through microbial dysbiosis or defective host defense toward invasive intestinal bacteria can result in chronic inflammation. We have shown that macrophages differentiated in the presence of the bacterial metabolite butyrate display enhanced antimicrobial activity. Butyrate-induced antimicrobial activity was associated with a shift in macrophage metabolism, a reduction in mTOR kinase activity, increased LC3-associated host defense and anti-microbial peptide production in the absence of an increased inflammatory cytokine response. Butyrate drove this monocyte to macrophage differentiation program through histone deacetylase 3 (HDAC3) inhibition. Administration of butyrate induced antimicrobial activity in intestinal macrophages in vivo and increased resistance to enteropathogens. Our data suggest that (1) increased intestinal butyrate might represent a strategy to bolster host defense without tissue damaging inflammation and (2) that pharmacological HDAC3 inhibition might drive selective macrophage functions toward antimicrobial host defense.
Subject(s)
Anti-Infective Agents/pharmacology , Butyrates/pharmacology , Cell Differentiation/drug effects , Macrophages/drug effects , Monocytes/drug effects , Animals , Cell Differentiation/genetics , Cells, Cultured , Colon/drug effects , Colon/metabolism , Colon/microbiology , Cytokines/genetics , Cytokines/metabolism , Dysbiosis/microbiology , Gene Expression Regulation/drug effects , Humans , Intestines/drug effects , Intestines/microbiology , Macrophages/metabolism , Macrophages/microbiology , Mice, Inbred C57BL , Microbiota/drug effects , Microbiota/physiology , Monocytes/metabolism , Monocytes/microbiologyABSTRACT
Tobacco smoking is positively correlated with non-alcoholic fatty liver disease (NAFLD)1-5, but the underlying mechanism for this association is unclear. Here we report that nicotine accumulates in the intestine during tobacco smoking and activates intestinal AMPKα. We identify the gut bacterium Bacteroides xylanisolvens as an effective nicotine degrader. Colonization of B. xylanisolvens reduces intestinal nicotine concentrations in nicotine-exposed mice, and it improves nicotine-exacerbated NAFLD progression. Mechanistically, AMPKα promotes the phosphorylation of sphingomyelin phosphodiesterase 3 (SMPD3), stabilizing the latter and therefore increasing intestinal ceramide formation, which contributes to NAFLD progression to non-alcoholic steatohepatitis (NASH). Our results establish a role for intestinal nicotine accumulation in NAFLD progression and reveal an endogenous bacterium in the human intestine with the ability to metabolize nicotine. These findings suggest a possible route to reduce tobacco smoking-exacerbated NAFLD progression.
Subject(s)
Bacteria , Intestines , Nicotine , Non-alcoholic Fatty Liver Disease , Tobacco Smoking , Animals , Humans , Mice , Bacteria/drug effects , Bacteria/metabolism , Ceramides/biosynthesis , Nicotine/adverse effects , Nicotine/metabolism , Non-alcoholic Fatty Liver Disease/chemically induced , Non-alcoholic Fatty Liver Disease/etiology , Non-alcoholic Fatty Liver Disease/microbiology , Sphingomyelin Phosphodiesterase/metabolism , Tobacco Smoking/adverse effects , Tobacco Smoking/metabolism , Intestines/drug effects , Intestines/microbiology , AMP-Activated Protein Kinases/metabolism , Disease ProgressionABSTRACT
Enteric pathogens are exposed to a dynamic polymicrobial environment in the gastrointestinal tract1. This microbial community has been shown to be important during infection, but there are few examples illustrating how microbial interactions can influence the virulence of invading pathogens2. Here we show that expansion of a group of antibiotic-resistant, opportunistic pathogens in the gut-the enterococci-enhances the fitness and pathogenesis of Clostridioides difficile. Through a parallel process of nutrient restriction and cross-feeding, enterococci shape the metabolic environment in the gut and reprogramme C. difficile metabolism. Enterococci provide fermentable amino acids, including leucine and ornithine, which increase C. difficile fitness in the antibiotic-perturbed gut. Parallel depletion of arginine by enterococci through arginine catabolism provides a metabolic cue for C. difficile that facilitates increased virulence. We find evidence of microbial interaction between these two pathogenic organisms in multiple mouse models of infection and patients infected with C. difficile. These findings provide mechanistic insights into the role of pathogenic microbiota in the susceptibility to and the severity of C. difficile infection.
Subject(s)
Clostridioides difficile , Enterococcus , Microbial Interactions , Animals , Humans , Mice , Anti-Bacterial Agents/pharmacology , Arginine/deficiency , Arginine/metabolism , Clostridioides difficile/metabolism , Clostridioides difficile/pathogenicity , Clostridioides difficile/physiology , Disease Models, Animal , Drug Resistance, Bacterial , Enterococcus/drug effects , Enterococcus/metabolism , Enterococcus/pathogenicity , Enterococcus/physiology , Gastrointestinal Microbiome/drug effects , Intestines/drug effects , Intestines/metabolism , Intestines/microbiology , Leucine/metabolism , Ornithine/metabolism , Virulence , Disease SusceptibilityABSTRACT
Dietary fibres can exert beneficial anti-inflammatory effects through microbially fermented short-chain fatty acid metabolites<sup>1,2</sup>, although the immunoregulatory roles of most fibre diets and their microbiota-derived metabolites remain poorly defined. Here, using microbial sequencing and untargeted metabolomics, we show that a diet of inulin fibre alters the composition of the mouse microbiota and the levels of microbiota-derived metabolites, notably bile acids. This metabolomic shift is associated with type 2 inflammation in the intestine and lungs, characterized by IL-33 production, activation of group 2 innate lymphoid cells and eosinophilia. Delivery of cholic acid mimics inulin-induced type 2 inflammation, whereas deletion of the bile acid receptor farnesoid X receptor diminishes the effects of inulin. The effects of inulin are microbiota dependent and were reproduced in mice colonized with human-derived microbiota. Furthermore, genetic deletion of a bile-acid-metabolizing enzyme in one bacterial species abolishes the ability of inulin to trigger type 2 inflammation. Finally, we demonstrate that inulin enhances allergen- and helminth-induced type 2 inflammation. Taken together, these data reveal that dietary inulin fibre triggers microbiota-derived cholic acid and type 2 inflammation at barrier surfaces with implications for understanding the pathophysiology of allergic inflammation, tissue protection and host defence.
Subject(s)
Bile Acids and Salts , Dietary Fiber , Gastrointestinal Microbiome , Inflammation , Inulin , Animals , Humans , Mice , Bile Acids and Salts/metabolism , Cholic Acid/pharmacology , Dietary Fiber/pharmacology , Gastrointestinal Microbiome/drug effects , Gastrointestinal Microbiome/physiology , Immunity, Innate , Inflammation/chemically induced , Inflammation/classification , Inflammation/pathology , Inulin/pharmacology , Lymphocytes/cytology , Lymphocytes/drug effects , Lymphocytes/immunology , Metabolomics , Lung/drug effects , Lung/pathology , Intestines/drug effects , Intestines/microbiology , Intestines/pathology , Interleukin-33/metabolism , Eosinophils/cytology , Eosinophils/drug effects , Eosinophils/immunologyABSTRACT
Genome-wide association studies have identified risk loci linked to inflammatory bowel disease (IBD)1-a complex chronic inflammatory disorder of the gastrointestinal tract. The increasing prevalence of IBD in industrialized countries and the augmented disease risk observed in migrants who move into areas of higher disease prevalence suggest that environmental factors are also important determinants of IBD susceptibility and severity2. However, the identification of environmental factors relevant to IBD and the mechanisms by which they influence disease has been hampered by the lack of platforms for their systematic investigation. Here we describe an integrated systems approach, combining publicly available databases, zebrafish chemical screens, machine learning and mouse preclinical models to identify environmental factors that control intestinal inflammation. This approach established that the herbicide propyzamide increases inflammation in the small and large intestine. Moreover, we show that an AHR-NF-κB-C/EBPß signalling axis operates in T cells and dendritic cells to promote intestinal inflammation, and is targeted by propyzamide. In conclusion, we developed a pipeline for the identification of environmental factors and mechanisms of pathogenesis in IBD and, potentially, other inflammatory diseases.
Subject(s)
Environment , Herbicides , Inflammation , Inflammatory Bowel Diseases , Intestines , Animals , Mice , Inflammation/chemically induced , Inflammation/etiology , Inflammation/immunology , Inflammation/pathology , Inflammatory Bowel Diseases/chemically induced , Inflammatory Bowel Diseases/etiology , Inflammatory Bowel Diseases/immunology , Inflammatory Bowel Diseases/pathology , Zebrafish , Machine Learning , Databases, Factual , Disease Models, Animal , Intestines/drug effects , Intestines/immunology , Intestines/metabolism , Intestines/pathology , NF-kappa B , CCAAT-Enhancer-Binding Protein-beta , Receptors, Aryl Hydrocarbon , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Dendritic Cells/drug effects , Dendritic Cells/immunology , Dendritic Cells/metabolism , Herbicides/adverse effectsABSTRACT
A novel group of biocidal compounds are the Crystal 3D (Cry) and Cytolytic (Cyt) proteins produced by Bacillus thuringiensis (Bt). Some Bt Cry proteins have a selective nematocidal activity, with Cry5B being the most studied. Cry5B kills nematode parasites by binding selectively to membrane glycosphingolipids, then forming pores in the cell membranes of the intestine leading to damage. Cry5B selectively targets multiple species of nematodes from different clades and has no effect against mammalian hosts. Levamisole is a cholinergic anthelmintic that acts by selectively opening L-subtype nicotinic acetylcholine receptor ion-channels (L-AChRs) that have been found on muscles of nematodes. A synergistic nematocidal interaction between levamisole and Cry5B at the whole-worm level has been described previously, but the location, mechanism and time-course of this synergism is not known. In this study we follow the timeline of the effects of levamisole and Cry5B on the Ca2+ levels in enterocyte cells in the intestine of Ascaris suum using fluorescence imaging. The peak Ca2+ responses to levamisole were observed after approximately 10 minutes while the peak responses to activated Cry5B were observed after approximately 80 minutes. When levamisole and Cry5B were applied simultaneously, we observed that the responses to Cry5B were bigger and occurred sooner than when it was applied by itself. It is proposed that the synergism is due to the cytoplasmic Ca2+ overload that is induced by the combination of levamisole opening Ca2+ permeable L-subtype nAChRs and the Ca2+ permeable Cry5B toxin pores produced in the enterocyte plasma membranes. The effect of levamisole potentiates and speeds the actions of Cry5B that gives rise to bigger Ca2+ overloads that accelerates cell-death of the enterocytes.
Subject(s)
Ascaris suum , Bacillus thuringiensis Toxins , Bacterial Proteins , Endotoxins , Hemolysin Proteins , Levamisole , Levamisole/pharmacology , Animals , Bacillus thuringiensis Toxins/pharmacology , Endotoxins/pharmacology , Endotoxins/metabolism , Hemolysin Proteins/pharmacology , Hemolysin Proteins/metabolism , Bacterial Proteins/metabolism , Ascaris suum/drug effects , Anthelmintics/pharmacology , Intestines/drug effects , Intestines/parasitology , Drug Synergism , Antinematodal Agents/pharmacology , Bacillus thuringiensis/drug effectsABSTRACT
Metformin, the world's most prescribed anti-diabetic drug, is also effective in preventing type 2 diabetes in people at high risk1,2. More than 60% of this effect is attributable to the ability of metformin to lower body weight in a sustained manner3. The molecular mechanisms by which metformin lowers body weight are unknown. Here we show-in two independent randomized controlled clinical trials-that metformin increases circulating levels of the peptide hormone growth/differentiation factor 15 (GDF15), which has been shown to reduce food intake and lower body weight through a brain-stem-restricted receptor. In wild-type mice, oral metformin increased circulating GDF15, with GDF15 expression increasing predominantly in the distal intestine and the kidney. Metformin prevented weight gain in response to a high-fat diet in wild-type mice but not in mice lacking GDF15 or its receptor GDNF family receptor α-like (GFRAL). In obese mice on a high-fat diet, the effects of metformin to reduce body weight were reversed by a GFRAL-antagonist antibody. Metformin had effects on both energy intake and energy expenditure that were dependent on GDF15, but retained its ability to lower circulating glucose levels in the absence of GDF15 activity. In summary, metformin elevates circulating levels of GDF15, which is necessary to obtain its beneficial effects on energy balance and body weight, major contributors to its action as a chemopreventive agent.
Subject(s)
Body Weight/drug effects , Energy Metabolism/drug effects , Growth Differentiation Factor 15/metabolism , Metformin/pharmacology , Administration, Oral , Adult , Aged , Animals , Blood Glucose/analysis , Blood Glucose/metabolism , Diet, High-Fat , Double-Blind Method , Energy Intake/drug effects , Enterocytes/cytology , Enterocytes/drug effects , Female , Glial Cell Line-Derived Neurotrophic Factor Receptors/antagonists & inhibitors , Glial Cell Line-Derived Neurotrophic Factor Receptors/deficiency , Glial Cell Line-Derived Neurotrophic Factor Receptors/genetics , Growth Differentiation Factor 15/blood , Growth Differentiation Factor 15/deficiency , Growth Differentiation Factor 15/genetics , Homeostasis/drug effects , Humans , Intestines/cytology , Intestines/drug effects , Male , Metformin/administration & dosage , Mice , Mice, Obese , Middle Aged , Weight Loss/drug effectsABSTRACT
The development of intestinal organoids from single adult intestinal stem cells in vitro recapitulates the regenerative capacity of the intestinal epithelium1,2. Here we unravel the mechanisms that orchestrate both organoid formation and the regeneration of intestinal tissue, using an image-based screen to assay an annotated library of compounds. We generate multivariate feature profiles for hundreds of thousands of organoids to quantitatively describe their phenotypic landscape. We then use these phenotypic fingerprints to infer regulatory genetic interactions, establishing a new approach to the mapping of genetic interactions in an emergent system. This allows us to identify genes that regulate cell-fate transitions and maintain the balance between regeneration and homeostasis, unravelling previously unknown roles for several pathways, among them retinoic acid signalling. We then characterize a crucial role for retinoic acid nuclear receptors in controlling exit from the regenerative state and driving enterocyte differentiation. By combining quantitative imaging with RNA sequencing, we show the role of endogenous retinoic acid metabolism in initiating transcriptional programs that guide the cell-fate transitions of intestinal epithelium, and we identify an inhibitor of the retinoid X receptor that improves intestinal regeneration in vivo.
Subject(s)
Organoids/cytology , Organoids/physiology , Phenotype , Regeneration/physiology , Animals , Cell Differentiation/drug effects , Cell Differentiation/genetics , Enterocytes/cytology , Enterocytes/drug effects , Homeostasis/drug effects , Intestinal Mucosa/drug effects , Intestinal Mucosa/metabolism , Intestines/cytology , Intestines/drug effects , Male , Mice , Mice, Inbred C57BL , Organoids/drug effects , Organoids/metabolism , Receptors, Retinoic Acid/antagonists & inhibitors , Receptors, Retinoic Acid/metabolism , Regeneration/drug effects , Sequence Analysis, RNA , Signal Transduction/drug effects , Transcription, Genetic/drug effects , Tretinoin/metabolism , Vitamin A/pharmacologyABSTRACT
Ischemia-reperfusion (IR) injury is primarily characterized by the restoration of blood flow perfusion and oxygen supply to ischemic tissue and organs, but it paradoxically leads to tissue injury aggravation. IR injury is a challenging pathophysiological process that is difficult to avoid clinically and frequently occurs during organ transplantation, surgery, shock resuscitation, and other processes. The major causes of IR injury include increased levels of free radicals, calcium overload, oxidative stress, and excessive inflammatory response. Ghrelin is a newly discovered brain-intestinal peptide with anti-inflammatory and antiapoptotic effects that improve blood supply. The role and mechanism of ghrelin in intestinal ischemia-reperfusion (IIR) injury remain unclear. We hypothesized that ghrelin could attenuate IIR-induced oxidative stress and apoptosis. To investigate this, we established IIR by using a non-invasive arterial clip to clamp the root of the superior mesenteric artery (SMA) in mice. Ghrelin was injected intraperitoneally at a dose of 50 µg/kg 20 min before IIR surgery, and [D-Lys3]-GHRP-6 was injected intraperitoneally at a dose of 12 nmol/kg 20 min before ghrelin injection. We mimicked the IIR process with hypoxia-reoxygenation (HR) in Caco-2 cells, which are similar to intestinal epithelial cells in structure and biochemistry. Our results showed that ghrelin inhibited IIR/HR-induced oxidative stress and apoptosis by activating GHSR-1α. Moreover, it was found that ghrelin activated the GHSR-1α/Sirt1/FOXO1 signaling pathway. We further inhibited Sirt1 and found that Sirt1 was critical for ghrelin-mediated mitigation of IIR/HR injury. Overall, our data suggest that pretreatment with ghrelin reduces oxidative stress and apoptosis to attenuate IIR/HR injury by binding with GHSR-1α to further activate Sirt1.
Subject(s)
Apoptosis , Forkhead Box Protein O1 , Ghrelin , Mice, Inbred C57BL , Oxidative Stress , Receptors, Ghrelin , Reperfusion Injury , Sirtuin 1 , Ghrelin/pharmacology , Ghrelin/metabolism , Reperfusion Injury/metabolism , Reperfusion Injury/drug therapy , Sirtuin 1/metabolism , Animals , Mice , Receptors, Ghrelin/metabolism , Humans , Male , Forkhead Box Protein O1/metabolism , Apoptosis/drug effects , Oxidative Stress/drug effects , Signal Transduction/drug effects , Intestines/drug effects , Caco-2 CellsABSTRACT
Although patients generally prefer oral drug delivery to injections, low permeability of the gastrointestinal tract makes this method impossible for most biomacromolecules. One potential solution is codelivery of macromolecules, including therapeutic proteins or nucleic acids, with intestinal permeation enhancers; however, enhancer use has been limited clinically by modest efficacy and toxicity concerns surrounding long-term administration. Here, we hypothesized that plant-based foods, which are well tolerated by the gastrointestinal tract, may contain compounds that enable oral macromolecular absorption without causing adverse effects. Upon testing more than 100 fruits, vegetables, and herbs, we identified strawberry and its red pigment, pelargonidin, as potent, well-tolerated enhancers of intestinal permeability. In mice, an oral capsule formulation comprising pelargonidin and a 1 U/kg dose of insulin reduced blood glucose levels for over 4 h, with bioactivity exceeding 100% relative to subcutaneous injection. Effects were reversible within 2 h and associated with actin and tight junction rearrangement. Furthermore, daily dosing of mice with pelargonidin for 1 mo resulted in no detectable side effects, including weight loss, tissue damage, or inflammatory responses. These data suggest that pelargonidin is an exceptionally effective enhancer of oral protein uptake that may be safe for routine pharmaceutical use.
Subject(s)
Anthocyanins , Fragaria , Intestinal Absorption , Intestines , Proteins , Administration, Oral , Animals , Anthocyanins/chemistry , Anthocyanins/pharmacology , Fragaria/chemistry , Insulin/administration & dosage , Insulin/pharmacokinetics , Intestinal Absorption/drug effects , Intestines/drug effects , Intestines/metabolism , Mice , Permeability , Proteins/administration & dosage , Proteins/pharmacokineticsABSTRACT
In patients with high-level radiation exposure, gastrointestinal injury is the main cause of death. Despite the severity of damage to the gastrointestinal tract, no specific therapeutic option is available. Tauroursodeoxycholic acid (TUDCA) is a conjugated form of ursodeoxycholic acid that suppresses endoplasmic reticulum (ER) stress and regulates various cell-signaling pathways. We investigated the effect of TUDCA premedication in alleviating intestinal damage and enhancing the survival of C57BL/6 mice administered a lethal dose (15Gy) of focal abdominal irradiation. TUDCA was administered to mice 1 h before radiation exposure, and reduced apoptosis of the jejunal crypts 12 h after irradiation. At later timepoint (3.5 days), irradiated mice manifested intestinal morphological changes that were detected via histological examination. TUDCA decreased the inflammatory cytokine levels and attenuated the decrease in serum citrulline levels after radiation exposure. Although radiation induced ER stress, TUDCA pretreatment decreased ER stress in the irradiated intestinal cells. The effect of TUDCA indicates the possibility of radiation therapy for cancer in tumor cells. TUDCA did not affect cell proliferation and apoptosis in the intestinal epithelium. TUDCA decreased the invasive ability of the CT26 metastatic colon cancer cell line. Reduced invasion after TUDCA treatment was associated with decreased matrix metalloproteinase (MMP)-7 and MMP-13 expression, which play important roles in invasion and metastasis. This study shows a potential role of TUDCA in protecting against radiation-induced intestinal damage and inhibiting tumor cell migration without any radiation and radiation therapy effect.
Subject(s)
Apoptosis , Endoplasmic Reticulum Stress , Mice, Inbred C57BL , Radiation-Protective Agents , Taurochenodeoxycholic Acid , Animals , Taurochenodeoxycholic Acid/pharmacology , Endoplasmic Reticulum Stress/drug effects , Endoplasmic Reticulum Stress/radiation effects , Apoptosis/drug effects , Apoptosis/radiation effects , Radiation-Protective Agents/pharmacology , Mice , Male , Intestines/radiation effects , Intestines/drug effects , Intestines/pathology , Disease Models, Animal , Intestinal Mucosa/drug effects , Intestinal Mucosa/radiation effects , Intestinal Mucosa/pathology , Intestinal Mucosa/metabolism , Radiation Injuries, Experimental/prevention & control , Radiation Injuries, Experimental/pathology , Radiation Injuries, Experimental/drug therapy , Radiation Injuries, Experimental/metabolism , Matrix Metalloproteinase 13/metabolism , Cell Proliferation/drug effects , Cell Proliferation/radiation effectsABSTRACT
Novel therapeutic strategies are needed to attenuate increased systemic and gut inflammation that contribute to morbidity and mortality in chronic HIV infection despite potent antiretroviral therapy (ART). The goal of this study is to use preclinical models of chronic treated HIV to determine whether the antioxidant and anti-inflammatory apoA-I mimetic peptides 6F and 4F attenuate systemic and gut inflammation in chronic HIV. We used two humanized murine models of HIV infection and gut explants from 10 uninfected and 10 HIV infected persons on potent ART, to determine the in vivo and ex vivo impact of apoA-I mimetics on systemic and intestinal inflammation in HIV. When compared to HIV infected humanized mice treated with ART alone, mice on oral apoA-I mimetic peptide 6F with ART had consistently reduced plasma and gut tissue cytokines (TNF-α, IL-6) and chemokines (CX3CL1) that are products of ADAM17 sheddase activity. Oral 6F attenuated gut protein levels of ADAM17 that were increased in HIV-1 infected mice on potent ART compared to uninfected mice. Adding oxidized lipoproteins and endotoxin (LPS) ex vivo to gut explants from HIV infected persons increased levels of ADAM17 in myeloid and intestinal cells, which increased TNF-α and CX3CL1. Both 4F and 6F attenuated these changes. Our preclinical data suggest that apoA-I mimetic peptides provide a novel therapeutic strategy that can target increased protein levels of ADAM17 and its sheddase activity that contribute to intestinal and systemic inflammation in treated HIV. The large repertoire of inflammatory mediators involved in ADAM17 sheddase activity places it as a pivotal orchestrator of several inflammatory pathways associated with morbidity in chronic treated HIV that make it an attractive therapeutic target.
Subject(s)
Apolipoprotein A-I , HIV Infections/pathology , Inflammation/pathology , Intestines/drug effects , Peptides/pharmacology , ADAM17 Protein/drug effects , Animals , Anti-HIV Agents/pharmacology , Humans , MiceABSTRACT
PURPOSE OF REVIEW: Over the past decade, trophic gastrointestinal hormonal factors have been included in the intestinal rehabilitation programs for short bowel syndrome (SBS). Up today the only trophic factor approved for clinical practice is the glucagon-like peptide-2 (GLP-2) analogue, teduglutide. A literature review on the last 2-year data on GLP-2 analogues for the treatment of SBS in adults has been performed. RECENT FINDINGS: Several reports on real-world data on the efficacy and safety of teduglutide treatment for SBS, some case-reports on the use of teduglutide in non-SBS conditions as well as phase 2 trials on new GL-2 analogues on patients with SBS have been retrieved. SUMMARY: Real-world data confirmed the teduglutide efficacy not only in weaning off IVS in accurately selected patients but also increased the alert on the risk of development of gastrointestinal polyps related to the drug; the impact of the therapy on patients' QoL deserves further studies and the cost-utility of the treatment is still uncertain. Some case reports highlighted the potential benefit of treatment with teduglutide in non-SBS gastrointestinal diseases, such as graft-versus-host disease, primary amyloidosis and refractory microscopic colitis. Phase 2 RCTs on safety and efficacy of two new long-acting GLP-2 analogues, glepaglutide and apraglutide, were published, and phase 3 RCTs have been completed.
Subject(s)
Gastrointestinal Agents , Glucagon-Like Peptide 2 , Peptides , Short Bowel Syndrome , Humans , Short Bowel Syndrome/drug therapy , Short Bowel Syndrome/rehabilitation , Glucagon-Like Peptide 2/therapeutic use , Peptides/therapeutic use , Gastrointestinal Agents/therapeutic use , Adaptation, Physiological/drug effects , Adult , Intestines/drug effects , Intestines/physiopathology , Quality of LifeABSTRACT
A 2D-intestinal epithelial Caco-2/RAW 264.7 macrophage co-culture model was developed to demonstrate the relative efficacy of different phenolic acids to mitigate changes in Caco-2 epithelial cell redox state initiated both directly by autoxidation products, H2O2, and indirectly through cell communication events originating from cytokine stimulated macrophage. An inducer cocktail (lipopolysaccharide + interferon gamma) was used to activate RAW 264.7 cells in the 2D- Caco-2/RAW co-culture and intracellular changes in Caco-2 cell redox signaling occurred in response to positive changes (p < 0.05) in inflammatory biomarkers derived in macrophage that included IL-6, TNF-α, nitric oxide and peroxynitrite, respectively. Phenolic acids varied in relative capacity to reduce NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) in cocktail inflamed induced macrophage. This response in addition to the relative predisposition of gallic acid (GA) to undergo autoxidation to generate H2O2 activity (p < 0.05), culminated in downstream cell signaling in Caco-2 nuclear factor erythroid 2-related factor (Nrf2) activity (increase 26.9 %), altered monolayer integrity (increase 33.7 %), and release of interleukin 8 (IL-8) (decrease 80.5 %) (p < 0.05). It can be concluded that the co-culture model described herein was useful to assess the importance of communication between cytokine stimulated macrophage and intestinal cells. Moreover, the relative unique efficacy of GA, compared to other phenolic acids tested to protect against activated macrophage induced changes related to intestinal dysfunction were particularly relevant to epithelial redox signaling, intestinal permeability and regulation of tight junction proteins. This study concludes that phenolic acids are not equal in the capacity to protect against intestinal cell dysfunction despite some indication of biological activity.
Subject(s)
Coculture Techniques , Gallic Acid , Tight Junction Proteins , Caco-2 Cells , Gallic Acid/pharmacology , Humans , Mice , Animals , RAW 264.7 Cells , Tight Junction Proteins/metabolism , Inflammation/metabolism , Oxidation-Reduction/drug effects , Hydrogen Peroxide/metabolism , Macrophages/metabolism , Macrophages/drug effects , Intestines/cytology , Intestines/drug effects , Intestinal Mucosa/metabolism , Intestinal Mucosa/drug effectsABSTRACT
BACKGROUND: α-Ketoglutarate (AKG) plays a pivotal role in mitigating inflammation and enhancing intestinal health. OBJECTIVES: This study aimed to investigate whether AKG could protect against lipopolysaccharide (LPS)-induced intestinal injury by alleviating disorders in mitochondria-associated endoplasmic reticulum (MAM) membranes, dysfunctional mitochondrial dynamics, and endoplasmic reticulum (ER) stress in a piglet model. METHODS: Twenty-four piglets were subjected to a 2 × 2 factorial design with dietary factors (basal diet or 1% AKG diet) and LPS treatment (LPS or saline). After 21 d of consuming either the basal diet or AKG diet, piglets received injections of LPS or saline. The experiment was divided into 4 treatment groups [control (CON) group: basal diet + saline; LPS group: basal diet +LPS; AKG group: AKG diet + saline; and AKG_LPS group: AKG + LPS], each consisting of 6 piglets. RESULTS: The results demonstrated that compared with the CON group, AKG enhanced jejunal morphology, antioxidant capacity, and the messenger RNA and protein expression of tight junction proteins. Moreover, it has shown a reduction in serum diamine oxidase activity and D-lactic acid content in piglets. In addition, fewer disorders in the ER-mitochondrial system were reflected by AKG, as evidenced by AKG regulating the expression of key molecules of mitochondrial dynamics (mitochondrial calcium uniporter, optic atrophy 1, fission 1, and dynamin-related protein 1), ER stress [activating transcription factor (ATF) 4, ATF 6, CCAAT/enhancer binding protein homologous protein, eukaryotic initiation factor 2α, glucose-regulated protein (GRP) 78, and protein kinase R-like ER kinase], and MAM membranes [mitofusin (Mfn)-1, Mfn-2, GRP 75, and voltage-dependent anion channel-1]. CONCLUSIONS: Dietary AKG can prevent mitochondrial dynamic dysfunction, ER stress, and MAM membrane disorder, ultimately alleviating LPS-induced intestinal damage in piglets.