int-777 and Disease-Models--Animal

Parkinson's disease (PD) is one of the most common chronic progressive neurodegenerative diseases that affects both motor and non-motor functions. Bile acids modulate the immune system by targeting brain receptors. INT-777, a 6α-ethyl-23(S)-methyl derivative of cholic acid (S-EMCA), acts as an agonist for Takeda G protein-coupled receptor-5 (TGR5) and has neuroprotective properties. However, the effects of INT-777 on PD have not yet been investigated. In a subchronic PD model, mice treated with 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) developed motor deficits and cognitive impairment that were ameliorated after intranasal administration of INT-777. INT-777 prevented MPTP-induced neurodegeneration and microglia activation in the substantia nigra pars compacta, hippocampus, and cortical layer V. Based on bioinformatics and wet lab data, INT-777 inhibited microglia activation by suppressing the release of tumor necrosis factor alpha (TNF-α) in the hippocampus, along with secondary chemokines (C-C motif ligand 3 (CCL3) and CCL6) in these three brain regions. INT-777 inhibited TNF-α production by repairing mitochondrial damage, which was associated with nuclear factor-erythroid 2-related factor-2 (NRF2) activation and p62/LC3B-mediated autophagy. INT-777 reversed the downregulation of heme oxygenase-1 (HO1), NAD(P)H quinone oxidoreductase-1 (NQO1) and accumulation of p62 in microglia treated with 1-methyl-4-phenylpyridinium (MPP+). However, TGR5 knockdown in microglia abolished INT-777's inhibition of TNF-α release, resulting in neuronal death. Therefore, PD cognitive impairment is associated with hippocampal TNF-α elevation as a result of mitochondrial damage in microglia. Our data reveal the potential role of TGR5 in modulating inflammation-mediated neurodegeneration in PD, and provides new insights for bile acid metabolites as promising disease-modifying drugs for PD.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; 1-Methyl-4-phenylpyridinium; Animals; Cholic Acids; Disease Models, Animal; Dopaminergic Neurons; Mice; Mice, Inbred C57BL; Microglia; Mitochondrial Dynamics; Parkinson Disease, Secondary; Tumor Necrosis Factor-alpha

Hepatic ischemia/reperfusion injury (IRI) still remains an unavoidable problem in hepatectomy. The inflammatory response plays an important role in its pathogenesis. The plasma membrane-bound G protein-coupled bile acid receptor (TGR5), as one of G protein-coupled receptor (GPCR) families, has been proved to serve a protective role in several liver diseases. However, the exact function of TGR5 in modulating IRI remains obscure. We injected wild mice with a small interfering RNA of TGR5 (si-TGR5) or TGR5 agonist (INT-777) and established liver partial warm ischemia/reperfusion model. The results showed that knockdown of TGR5 significantly aggravated hepatic tissue injury, but treatment with INT-777 could reverse it, as evidenced by serum ALT and AST tests, liver histological injury, cytokines expressions, liver immunohistochemical analysis, and TUNEL staining. The apoptosis-associated proteins were evaluated after reperfusion. Moreover, we used primary bone marrow-derived macrophages (BMDMs) to establish hypoxia/reoxygenation (H/R) model to verify the anti-inflammation effect of TGR5. In in vivo experiments, we used TGR5-siRNA and TGR5 agonist (INT-777) to determine that TGR5 significantly attenuated liver damage after IRI through activating the Keap1-Nrf2 pathway. In addition, we found that overexpression of INT-777-activated TGR5 could reduce oxidative stress and inflammatory response in H/R-induced BMDMs through regulation of Keap1-Nef2 pathway during in vitro experiment. Importantly, these results were completely reversed in si-TGR5 BMDMs. In conclusion, the results indicated that TGR5 could effectively alleviated inflammation response via accelerating the activation of Keap1-Nrf2 signaling pathway during hepatic IRI, which may be meaningful in reducing related inflammatory molecules and adjusting inherent immunity.

Topics: Animals; Apoptosis; Cell Hypoxia; Cells, Cultured; Cholic Acids; Cytokines; Disease Models, Animal; Kelch-Like ECH-Associated Protein 1; Liver; Liver Diseases; Macrophages; Male; Mice, Inbred C57BL; NF-E2-Related Factor 2; Oxidative Stress; Receptors, G-Protein-Coupled; Reperfusion Injury; RNA Interference; Signal Transduction; Warm Ischemia

Chronic HBV infection is a major health problem worldwide. Currently, the first-line treatment for HBV is nucleos(t)ide analogs or interferons; however, efficient therapeutic approaches that enable cure are lacking. Therefore, anti-HBV agents with mechanisms distinct from those of current drugs are needed. Sodium taurocholate cotransporting polypeptide (NTCP) was previously identified as an HBV receptor that is inhibited by several compounds. Farnesoid X receptor (FXR) activation also inhibits NTCP function.. In this study, we investigated the inhibitory effect of bile acid (BA) derivatives-namely obeticholic acid (OCA), 6α-ethyl-24-nor-5β-cholane-3α,7α,23-triol-23 sulfate sodium salt (INT-767; a dual agonist of FXR and Takeda G protein-coupled receptor [TGR5]), and 6α-ethyl-23(S)-methyl-cholic acid (INT-777; a TGR5 agonist)-3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylisoxazole (GW4064; a FXR agonist), cyclosporin A, and irbesartan. OCA and INT-777 suppressed HBV infection in HepG2-human NTCP-C4 cells. Interestingly, INT-767 showed potent inhibition by attaching to HBV particles rather than binding to NTCP. As an entry inhibitor, INT-767 was stronger than various natural BAs. Furthermore, in chimeric mice with humanized liver, INT-767 markedly delayed the initial rise of HBsAg, HBeAg, and HBV DNA and reduced covalently closed circular DNA. The strong inhibitory effect of INT-767 may be due to the cumulative effect of its ability to inhibit the entry of HBV and to stimulate FXR downstream signaling, which affects the postentry step.. Our results suggest that BA derivatives, particularly INT-767, are prospective candidate anti-HBV agents. Clarifying the underlying mechanisms of BA derivatives would facilitate the development of anti-HBV agents.

Topics: Animals; Antiviral Agents; Bile Acids and Salts; Chenodeoxycholic Acid; Cholic Acids; Disease Models, Animal; Hep G2 Cells; Hepatitis B virus; Hepatitis B, Chronic; Humans; Male; Mice; Organic Anion Transporters, Sodium-Dependent; Prospective Studies; Receptors, Cytoplasmic and Nuclear; Receptors, G-Protein-Coupled; Symporters; Transplantation Chimera; Virus Internalization

Alopecia areata is an autoimmune hair loss disease with infiltration of pro-inflammatory cells into hair follicles. The role of Tgr5 in dermatitis has attracted considerable attention. The present study aimed to investigate the effect of Tgr5 in the development of Alopecia areata.. The study utilized a comparison control group design with four groups of wild-type group, wild-type+INT777 group, Tgr5-/- group, and Tgr5-/-+INT777 group. The mice were treated with INT777 (30 mg/kg/day) or the carrier solution (DMSO) intraperitoneally for 7 weeks, and the back skin was collected and analyzed by histology and immunohistochemistry staining. The lumbar vertebrae 4 has also been analyzed by DXA and Micro-CT.. Tgr5-/- mice displayed the decreasingly significant in hair area and length, skin thickness, and the ratio of anagen and telogen, collagen, and mast cell number and loss the bone mass than WT group. After treating with INT777, the appearance of alopecia areata and bone microstructure has improved. Immunohistochemistry and qPCR analysis showed that activation of Tgr5 can down-regulate the express of JAK1, STAT3, IL-6, TNF-α, and VEGF.. These findings indicate that activation of Tgr5 mediated amelioration of alopecia areata and osteoporosis by down-regulated JAK1-STAT3 signaling pathway.

Topics: Alopecia Areata; Animals; Anti-Inflammatory Agents; Bone Density; Cholic Acids; Disease Models, Animal; Hair Follicle; Interleukin-6; Janus Kinase 1; Lumbar Vertebrae; Male; Mice, Inbred C57BL; Mice, Knockout; Osteoporosis; Receptors, G-Protein-Coupled; Signal Transduction; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha; Vascular Endothelial Growth Factor A

Renewal and patterning of the intestinal epithelium is coordinated by intestinal stem cells (ISCs); dietary and metabolic factors provide signals to the niche that control ISC activity. Bile acids (BAs), metabolites in the gut, signal nutrient availability by activating the G protein-coupled bile acid receptor 1 (GPBAR1, also called TGR5). TGR5 is expressed in the intestinal epithelium, but it is not clear how its activation affects ISCs and regeneration of the intestinal epithelium. We studied the role of BAs and TGR5 in intestinal renewal, and regulation of ISC function in mice and intestinal organoids.. We derived intestinal organoids from wild-type mice and Tgr5. BAs and TGR5 agonists promoted growth of intestinal organoids. Administration of cholecystokinin to mice resulted in acute release of BAs into the intestinal lumen and increased proliferation of the intestinal epithelium. BAs and Tgr5 expression in ISCs were required for homeostatic intestinal epithelial renewal and fate specification, and for regeneration after colitis induction. Tgr5. BAs promote regeneration of the intestinal epithelium via activation of TGR5 in ISCs, resulting in activation of SRC and YAP and activation of their target genes. Release of endogenous BAs in the intestinal lumen is sufficient to promote ISC renewal and drives regeneration in response to injury.

Topics: Adaptor Proteins, Signal Transducing; Adult Stem Cells; Animals; Bile Acids and Salts; Cell Cycle Proteins; Cell Self Renewal; Cells, Cultured; Cholic Acids; Colitis; Dextran Sulfate; Disease Models, Animal; Epithelial Cells; Humans; Intestinal Mucosa; Male; Mice; Mice, Knockout; Organoids; Primary Cell Culture; Receptors, G-Protein-Coupled; Regeneration; Signal Transduction; src-Family Kinases; YAP-Signaling Proteins

Neuroinflammation plays an important role in the pathophysiology of Alzheimer's disease (AD) and memory impairment. Herein, we evaluated the neuroprotective effects of 6-ethyl-23(S)-methyl-cholic acid (INT-777), a specific G-protein coupled bile acid receptor 1 (TGR5) agonist, in the LPS-treated mouse model of acute neurotoxicity. Single intracerebroventricular (i.c.v.) injection of LPS remarkably induced mouse behavioral impairments in Morris water maze, novel object recognition, and Y-maze avoidance tests, which were ameliorated by INT-777 (1.5 or 3.0 μg/mouse, i.c.v.) treatment. Importantly, INT-777 treatment reversed LPS-induced TGR5 down-regulation, suppressed the increase of nuclear NF-κB p65, and mitigated neuroinflammation, evidenced by lower proinflammatory cytokines, less activation of microglia, and increased the ratio of p-CREB/CREB or mBDNF/proBDNF in the hippocampus and frontal cortex. In addition, INT-777 treatment also suppressed neuronal apoptosis, as indicated by the reduction of TUNEL-positive cells, decreased activation of caspase-3, increased the ratio of Bcl-2/Bax, and ameliorated synaptic dysfunction as evidenced by the upregulation of PSD95 and synaptophysin in the hippocampus and frontal cortex. Taken together, this study showed the potential neuroprotective effects of INT-777 against LPS-induced cognitive impairment, neuroinflammation, apoptosis, and synaptic dysfunction in mice.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; bcl-2-Associated X Protein; Calcium-Binding Proteins; Cholic Acids; Cognition Disorders; Cytokines; Disease Models, Animal; Drug Interactions; Encephalitis; Exploratory Behavior; Lipopolysaccharides; Male; Maze Learning; Mice; Mice, Inbred ICR; Microfilament Proteins; Nerve Tissue Proteins; Recognition, Psychology; Synapses

Bile acids receptor TGR5 and its agonist INT-777, which has been found to be involved in the NLRP3 inflammasome pathway, play an important role in inflammatory diseases. However, the role of INT-777 in acute pancreatitis (AP) has not been reported. In this present study, we found that TGR5 was expressed in pancreatic tissue and increased after AP onset induced by caerulein and further evaluated the impact of INT-777 on the severity of AP. The results showed that INT-777 could reduce the severity of AP in mice, which was manifested as decreased pancreatic tissue damage as well as the decrease of serum enzymes (amylase and lipase), pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) and the expression of necrosis related proteins (RIP3 and p-MLKL). Furthermore, we found that INT-777 reduced the reactive oxygen species (ROS) production in pancreatic acinar cells and inhibited the activation of NLRP3 inflammasome pathway. In conclusion, our data showed that INT-777 could protect pancreatic acinar cell against necrosis and reduce the severity of AP, which may be mediated by inhibiting ROS/NLRP3 inflammasome pathway.

Topics: Acinar Cells; Animals; Ceruletide; Cholic Acids; Disease Models, Animal; Inflammasomes; Male; Mice; Mice, Inbred ICR; Necrosis; NLR Family, Pyrin Domain-Containing 3 Protein; Pancreas, Exocrine; Pancreatitis; Protective Agents; Reactive Oxygen Species; Receptors, G-Protein-Coupled

Administration of cholic acid, or its synthetic derivative, 6-alpha-ethyl-23(S)-methylcholic acid (INT-777), activates the membrane GPCR, TGR5, influences whole body metabolism, reduces atherosclerosis, and benefits the cardiovascular physiology in mice. Direct effects of TGR5 agonists, and the role for TGR5, on myocardial cell biology and stress response are unknown.. Mice were fed chow supplemented with 0.5% cholic acid (CA) or 0.025% INT-777, a specific TGR5 agonist, or regular chow for 3 weeks. Anthropometric, biochemical, physiologic (electrocardiography and echocardiography), and molecular analysis was performed at baseline. CA and INT-777 fed mice were challenged with acute exercise-induced stress, acute catecholamine-induced stress, and hemodynamic stress induced by transverse aortic constriction (TAC) for a period of 8 weeks. In separate experiments, mice born with constitutive deletion of TGR5 in cardiomyocytes (CM-TGR5. Administration of CA and INT-777 supplemented diets upregulated TGR5 expression and activated Akt, PKA, and ERK1/2 in the heart. CA and INT-777 fed mice showed improved exercise tolerance, improved sensitivity to catecholamine and attenuation in pathologic remodeling of the heart under hemodynamic stress. In contrast, CM-TGR5. Bile acids, specifically TGR5 agonists, induce cytoprotective changes in the heart and improve myocardial response to physiologic, inotropic, and hemodynamic stress in mice. TGR5 plays a critical role in myocardial adaptability, and TGR5 activation may represent a potentially attractive treatment option in heart failure.

Topics: Adaptation, Physiological; Animals; Cardiotonic Agents; Cells, Cultured; Cholic Acids; Cytoprotection; Disease Models, Animal; Exercise Tolerance; Heart Failure; Male; Mice, Inbred C57BL; Mice, Knockout; Myocardial Contraction; Myocardium; Receptors, G-Protein-Coupled; Signal Transduction

Glucose and lipid metabolism disorders and chronic inflammation in the kidney tissues are largely responsible for causative pathological mechanism of renal fibrosis in diabetic nephropathy (DN). As our previous findings confirmed that, sphingosine 1-phosphate (S1P)/sphingosine 1-phosphate receptor 2 (S1P2) signaling activation promoted renal fibrosis in diabetes. Numerous studies have demonstrated that the G protein-coupled bile acid receptor TGR5 exhibits effective regulation of glucose and lipid metabolism and anti-inflammatory effects. TGR5 is highly expressed in kidney tissues, whether it attenuates the inflammation and renal fibrosis by inhibiting the S1P/S1P2 signaling pathway would be a new insight into the molecular mechanism of DN. Here we investigated the effects of TGR5 on diabetic renal fibrosis, and the underlying mechanism would be also discussed. We found that TGR5 activation significantly decreased the expression of intercellular adhesion molecule-1 (ICAM-1) and transforming growth factor-beta 1 (TGF-β1), as well as fibronectin (FN) induced by high glucose in glomerular mesangial cells (GMCs), which were pathological features of DN. S1P2 overexpression induced by high glucose was diminished after activation of TGR5, and AP-1 activity, including the phosphorylation of c-Jun/c-Fos and AP-1 transcription activity, was attenuated. As a G protein-coupled receptor, S1P2 interacted with TGR5 in GMCs. Furthermore, INT-777 lowered S1P2 expression and promoted S1P2 internalization. Taken together, TGR5 activation reduced ICAM-1, TGF-β1 and FN expressions induced by high glucose in GMCs, the mechanism might be through suppressing S1P/S1P2 signaling, thus ameliorating diabetic nephropathy.

Topics: Animals; Cells, Cultured; Cholic Acids; Diabetic Nephropathies; Disease Models, Animal; Fibronectins; Fibrosis; Glucose; Intercellular Adhesion Molecule-1; Lysophospholipids; Mesangial Cells; Mice, Inbred C57BL; Phosphorylation; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Receptors, Lysosphingolipid; RNA Interference; Signal Transduction; Sphingosine; Sphingosine-1-Phosphate Receptors; Transcription Factor AP-1; Transfection; Transforming Growth Factor beta1

Chronic cholangiopathies have limited therapeutic options and represent an important indication for liver transplantation. The nuclear farnesoid X receptor (FXR) and the membrane G protein-coupled receptor, TGR5, regulate bile acid (BA) homeostasis and inflammation. Therefore, we hypothesized that activation of FXR and/or TGR5 could ameliorate liver injury in Mdr2(-/-) (Abcb4(-/-)) mice, a model of chronic cholangiopathy. Hepatic inflammation, fibrosis, as well as bile secretion and key genes of BA homeostasis were addressed in Mdr2(-/-) mice fed either a chow diet or a diet supplemented with the FXR agonist, INT-747, the TGR5 agonist, INT-777, or the dual FXR/TGR5 agonist, INT-767 (0.03% w/w). Only the dual FXR/TGR5 agonist, INT-767, significantly improved serum liver enzymes, hepatic inflammation, and biliary fibrosis in Mdr2(-/-) mice, whereas INT-747 and INT-777 had no hepatoprotective effects. In line with this, INT-767 significantly induced bile flow and biliary HCO 3- output, as well as gene expression of carbonic anhydrase 14, an important enzyme able to enhance HCO 3- transport, in an Fxr-dependent manner. In addition, INT-767 dramatically reduced bile acid synthesis via the induction of ileal Fgf15 and hepatic Shp gene expression, thus resulting in significantly reduced biliary bile acid output in Mdr2(-/-) mice.. This study shows that FXR activation improves liver injury in a mouse model of chronic cholangiopathy by reduction of biliary BA output and promotion of HCO 3--rich bile secretion.

Topics: Adenosine Triphosphatases; Analysis of Variance; Animals; Anion Transport Proteins; ATP Binding Cassette Transporter, Subfamily B; ATP-Binding Cassette Sub-Family B Member 4; Bile Acids and Salts; Biliary Tract Diseases; Cholic Acids; Disease Models, Animal; Liver Diseases; Male; Mice; Mice, Inbred C57BL; Random Allocation; Receptors, Cytoplasmic and Nuclear; Receptors, G-Protein-Coupled; Statistics, Nonparametric