Takeda G-protein Receptor Research Articles

Galacto-Oligosaccharides Alleviate LPS-Induced Immune Imbalance in Small Intestine through Regulating Gut Microbe Composition and Bile Acid Pool.

Recent evidence suggests that the protective effect of gut microbiota on intestinal inflammation can be achieved through a microbe-bile acids (BAs) mechanism. Galacto-oligosaccharides (GOS) are a kind of prebiotic that alter gut microbiota composition. To verify whether GOS has a protective effect on intestinal inflammation through a microbe-BAs mechanism, this research was performed in a lipopolysaccharide (LPS) porcine model with the presence or absence of GOS. GOS prevented LPS-induced production of pro-inflammatory cytokines, the decrease of bacterial bile salt hydrolase-containing bacteria abundance, and the decrease of chendoxycholic acid (CDCA) level in piglets. Additionally, CDCA decreased LPS-induced production of pro-inflammatory cytokines, induced the expression of the takeda G-protein receptor 5 (TGR5), and its downstream cyclic adenosine monophosphate (cAMP) production in lamina propria-derived CD11b+ cells. The cAMP inhibitor eliminated the protective effect of CDCA on lamina propria-derived CD11b+ cells. These results suggested that GOS reduced the production of pro-inflammatory cytokines and inhibited NF-κB activation via microbe-BA-dependent TGR5-cAMP signaling in LPS-challenged piglets.

Oleanolic acid promotes skeletal muscle fiber type transformation by activating TGR5-mediated CaN signaling pathway

In recent years, the impact of bile acids and their representative G protein-coupled bile acid receptor 1 Takeda-G-protein-receptor-5 (TGR5) signaling pathway on muscle function and metabolic health has gained considerable interest. Increasing the content of slow muscle fibers has been recognized as an effective strategy to improve metabolic health. Oleanolic acid (OA) is a naturally occurring triterpenoid compound derived from plants, which can activate TGR5. The aim of this study was to investigate the effect of OA and TGR5 on muscle fiber types and further explore the underlying TGR5-dependent mechanisms. In this study, mice were divided into three groups and dietary supplementation with 0, 50, or 100 mg/kg OA. In addition, C2C12 cells were treated with OA at concentrations of 0, 5, 10, and 20 µM. Our studies revealed that OA promoted the conversion of fast to slow muscle fibers. In addition, it was found that OA activated the TGR5-mediated calcineurin (CaN)/nuclear factor of activated T cells cytoplasmic 1 (NFATc1) signaling pathway. Further mechanistic investigations demonstrated that inhibiting TGR5 and CaN abolished the effects of OA on muscle fiber types transformation. In conclusion, this study found that OA promotes the transformation of fast muscle fibers to slow muscle fibers through the TGR5-mediated CaN/NFATc1 signaling pathway.

Protective effects of scutellaria-coptis herb couple against non-alcoholic steatohepatitis via activating NRF2 and FXR pathways in vivo and in vitro

Ethnopharmacological relevanceScutellaria-coptis herb couple (SC) is a classic herbal pair used in many Traditional Chinese Medicine (TCM) formulations in the treatment of endocrine and metabolic deseases. Diabetes mellitus and non-alcoholic steatohepatitis (NASH) are both endocrine and metabolic diseases. Previous studies have shown that SC has anti-diabetic effects. However, the effect and mechanism of SC against NASH remains unclear. Aim of the studyThis study aimed to demonstrate the effect and mechanism of SC against NASH through the nuclear factor-erythroid 2-related factor 2 (Nrf2) and farnesoid X receptor (FXR) dual signaling pathways in vivo and in vitro. Materials and methodsThe high fat diet-fed rat model, and HepG2 and RAW264.7 cell models were used. Serum biochemical indexes and liver histopathological changes were examined. Metabolomics, transcriptomics, and flow cytometry were performed. RT-qPCR and western blot analysis were performed to provide expression of NRF2 and FXR pathway signal molecules during SC's anti-NASH treatment in vivo and in vitro. ResultsSC had anti-NASH effects in vivo with significantly improvement of serum NASH biochemical index and hepatopathological structure; meanwhile, SC significantly elevated the expression levels of FXR protein in liver and intestinal tissues, and cholesterol 7a-hydroxylase (CYP7A1) protein in liver. The mRNA expression levels of Takeda G protein receptor 5 (TGR5), CYP7A1, fibroblast growth factor receptor-4 (FGFR4), FXR, small heterodimer partner (SHP), fibroblast growth factor 15/19 (FGF15/19) and glucagon-like peptide-1 (GLP-1) were significantly elevated by SC. SC reduced the levels of NorCA, isoLCA and α-MCA in the feces of NAFLD rats. In vitro, SC-containing serum (SC-CS) was found to significantly reduce intracellular lipid deposition, inhibit ROS production, reduce intracellular Malondialdehyde (MDA) and IL-1β levels, and enhance the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Six differential genes closely related to oxidative stress and Nrf2 were identified by transcriptomic analysis. SC-CS up-regulated the expression of NRF2, and reduced the expression of TXNIP and Caspase-1 genes in RAW264.7 cells. In addition, SC-CS reduced the expression of Keap1 and NF-κB, and up-regulated the expression of Nrf2, heme oxygenase-1 (HO-1), quinone oxidoreductase 1 (NQO1), and SOD; SC-CS elevated the protein level of NRF2, and reduced the protein level of TXNIP in HepG2 cells. Conclusionsthe mechanisms of SC action against NASH was closely related to the simultaneous activations of both NRF2 and FXR signaling pathways. These findings provide a new insight into the anti-NASH application of SC in clinical settings and demonstrate the potential of SC in the treatment of NASH.

Peroxisome proliferator-activated receptors, farnesoid X receptor, and dual modulating drugs in hypertension.

Hypertension characterized by an elevated blood pressure is a cardiovascular disease that afflicts greater than one in every three adults worldwide. Nuclear receptors are large superfamily of DNA-binding transcription factors that target genes to regulate metabolic and cardiovascular function. Drugs have been developed for nuclear receptors such as peroxisome proliferator-activated receptors (PPARα and PPARγ) and farnesoid X receptor (FXR). PPARα, PPARγ, and FXR agonists are used clinically to treat lipid disorders and metabolic diseases. Evidence from clinical studies and animal hypertension models have demonstrated that PPARα, PPARγ, and FXR agonism can lower blood pressure and decrease end organ damage which could be useful for the treatment of hypertension in patients with metabolic diseases. Unfortunately, PPAR and FXR agonists have unwanted clinical side effects. There have been recent developments to limit side effects for PPAR and FXR agonists. Combining PPAR and FXR agonism with soluble epoxide hydrolase (sEH) inhibition or Takeda G protein receptor 5 (TGR5) agonism has been demonstrated in preclinical studies to have actions that would decrease clinical side effects. In addition, these dual modulating drugs have been demonstrated in preclinical studies to have blood pressure lowering, anti-fibrotic, and anti-inflammatory actions. There is now an opportunity to thoroughly test these novel dual modulators in animal models of hypertension associated with metabolic diseases. In particular, these newly developed dual modulating PPAR and FXR drugs could be beneficial for the treatment of metabolic diseases, organ fibrosis, and hypertension.

CYP3A deficiency alters bile acid homeostasis and leads to changes in hepatic susceptibility in rats

Cytochrome P450 3A (CYP3A) as an important enzyme metabolizes many drugs and a variety of endogenous substances. Bile acids (BA) regulate physiological function by activating BA receptors. In this study, CYP3A1/2 gene knockout (KO) and wild-type (WT) rats were used to investigate the regulatory effects of CYP3A on BA homeostasis and liver function. Compared with WT rats, BA concentrations in serum, liver and small intestine of CYP3A1/2 KO rats increased significantly, which was due to the decrease of catabolism and the increase of synthesis. In particular, the composition of serum BA (overall hydrophobicity) presented an age- and CYP3A-dependent manner. With the aging of WT rats, the serum BA became more hydrophobic, while this trend was delayed in CYP3A1/2 KO rats. Moreover, the level of serum total cholesterol, the precursor of BA synthesis, decreased by about 20% in CYP3A1/2 KO rats, which is due to the low synthesis but high biotransformation rate. The increase of BA pool further led to the change of transcription level of BA receptor in liver (pregnane X receptor) and small intestine (Takeda G-protein receptor 5), and affected the function and morphology of CYP3A1/2 KO rat liver. In conclusion, CYP3A is a key regulator of BA homeostasis in rats, especially in regulating BA pool size, composition and balance of anabolism, and prevents susceptibility to hepatotoxicity under BA overload.

Marine Bile Natural Products as Agonists of the TGR5 Receptor

Agonism of the G protein-coupled bile acid receptor "Takeda G-protein receptor 5" (TGR5) aids in attenuating cholesterol accumulation due to atherosclerotic progression. Although mammalian bile compounds can activate TGR5, they are generally weak agonists, and more effective compounds need to be identified. In this study, two marine bile compounds (5β-scymnol and its sulfate) were compared with mammalian bile compounds deoxycholic acid (DCA) and ursodeoxycholic acid (UDCA) using an in vitro model of TGR5 agonism. The response profiles of human embryonic kidney 293 cells (HEK293) transfected to overexpress TGR5 (HEK293-TGR5) and incubated with subcytotoxic concentrations of test compounds were compared to nontransfected HEK293 control cells using the specific calcium-binding fluorophore Fura-2AM to measure intracellular calcium [Ca2+]i release. Scymnol and scymnol sulfate caused a sustained increase in [Ca2+]i within TGR5 cells only, which was abolished by a specific inhibitor for Gαq protein (UBO-QIC). Sustained increases in [Ca2+]i were seen in both cell types with DCA exposure; this was unaffected by UBO-QIC, indicating that TGR5 activation was not involved. Exposure to UDCA did not alter [Ca2+]i, suggesting a lack of TGR5 bioactivity. These findings demonstrated that both scymnol and scymnol sulfate are novel agonists of TGR5 receptors, showing therapeutic potential for treating atherosclerosis.

Dietary Fiber Is Essential to Maintain Intestinal Size, L-Cell Secretion, and Intestinal Integrity in Mice.

Dietary fiber has been linked to improved gut health, yet the mechanisms behind this association remain poorly understood. One proposed mechanism is through its influence on the secretion of gut hormones, including glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2). We aimed to: 1) investigate the impact of a fiber deficient diet on the intestinal morphological homeostasis; 2) evaluate L-cell secretion; and 3) to ascertain the role of GLP-1, GLP-2 and Takeda G protein-receptor-5 (TGR5) signaling in the response using GLP-1 receptor, GLP-2 receptor and TGR5 knockout mice. Female C57BL/6JRj mice (n = 8) either received a standard chow diet or were switched to a crude fiber-deficient diet for a short (21 days) and long (112 days) study period. Subsequent identical experiments were performed in GLP-1 receptor, GLP-2 receptor and TGR5 knockout mice. The removal of fiber from the diet for 21 days resulted in a decrease in small intestinal weight (p < 0.01) and a corresponding decrease in intestinal crypt depth in the duodenum, jejunum and ileum (p < 0.001, p < 0.05, and p < 0.01, respectively). Additionally, colon weight was decreased (p < 0.01). These changes were associated with a decrease in extractable GLP-1, GLP-2 and PYY in the colon (p < 0.05, p < 0.01, and p < 0.01). However, we could not show that the fiber-dependent size decrease was dependent on GLP-1 receptor, GLP-2 receptor or TGR5 signaling. Intestinal permeability was increased following the removal of fiber for 112 days. In conclusion, our study highlights the importance of dietary fiber to maintain intestinal weight, colonic L-cell secretion and intestinal integrity.

Gut Dysbiosis and Abnormal Bile Acid Metabolism in Colitis-Associated Cancer.

Background Patients with prolonged inflammatory bowel disease (IBD) can develop into colorectal cancer (CRC), also called colitis-associated cancer (CAC). Studies have shown the association between gut dysbiosis, abnormal bile acid metabolism, and inflammation process. Here, we aimed to investigate these two factors in the CAC model. Methods C57BL/6 mice were randomly allocated to two groups: azoxymethane/dextran sodium sulfate (AOM/DSS) and control. The AOM/DSS group received AOM injection followed by DSS drinking water. Intestinal inflammation, mucosal barrier, and bile acid receptors were determined by real-time PCR and immunohistochemistry. Fecal microbiome and bile acids were detected via 16S rRNA sequencing and liquid chromatography-mass spectrometry. Results The AOM/DSS group exhibited severe mucosal barrier impairment, inflammatory response, and tumor formation. In the CAC model, the richness and biodiversity of gut microbiota were decreased, along with significant alteration of composition. The abundance of pathogens was increased, while the short-chain fatty acids producing bacteria were reduced. Interestingly, Clostridium XlV and Lactobacillus, which might be involved in the bile acid deconjugation, transformation, and desulfation, were significantly decreased. Accordingly, fecal bile acids were decreased, accompanied by reduced transformation of primary to secondary bile acids. Given bile acid receptors, the ileum farnesoid X receptor-fibroblast growth factor 15 (FXR-FGF15) axis was downregulated, while Takeda G-protein receptor 5 (TGR5) was overexpressed in colonic tumor tissues. Conclusion Gut dysbiosis might alter the metabolism of bile acids and promote CAC, which would provide a potential preventive strategy of CAC by regulating gut microbiota and bile acid metabolism.

TGR5 activation attenuates neuroinflammation via Pellino3 inhibition of caspase-8/NLRP3 after middle cerebral artery occlusion in rats

BackgroundNucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) plays an important role in mediating inflammatory responses during ischemic stroke. Bile acid receptor Takeda-G-protein-receptor-5 (TGR5) has been identified as an important component in regulating brain inflammatory responses. In this study, we investigated the mechanism of TGR5 in alleviating neuroinflammation after middle cerebral artery occlusion (MCAO).MethodsSprague-Dawley rats were subjected to MCAO and TGR5 agonist INT777 was administered intranasally 1 h after MCAO. Small interfering RNAs (siRNA) targeting TGR5 and Pellino3 were administered through intracerebroventricular injection 48 h before MCAO. Infarct volumes and neurologic scores were evaluated, and ELISA, flow cytometry, immunofluorescence staining, immunoblotting, and co-immunoprecipitation were used for the evaluations.ResultsEndogenous TGR5 and Pellino3 levels increased after MCAO. TGR5 activation by INT777 significantly decreased pro-inflammatory cytokine, cleaved caspase-8, and NLRP3 levels, thereby reducing brain infarctions; both short- and long-term neurobehavioral assessments showed improvements. Ischemic damage induced the interaction of TGR5 with Pellino3. Knockdown of either TGR5 or Pellino3 increased the accumulation of cleaved caspase-8 and NLRP3, aggravated cerebral impairments, and abolished the anti-inflammatory effects of INT777 after MCAO.ConclusionsTGR5 activation attenuated brain injury by inhibiting neuroinflammation after MCAO, which could be mediated by Pellino3 inhibition of caspase-8/NLRP3.

Intestinal Co-culture System to Study TGR5 Agonism and Gut Restriction.

The activation of the Takeda G-protein receptor 5 (TGR5, also known as the G protein-coupled bile acid receptor 1, GPBAR1) in enteroendocrine L-cells results in secretion of the anti-diabetic hormone Glucagon-Like Peptide 1 (GLP-1) into systemic circulation. Consequently, recent research has focused on identification and development of TGR5 agonists as type 2 diabetes therapeutics. However, the clinical application of TGR5 agonists has been hampered by side effects of these compounds that primarily result from their absorption into circulation. Here we describe an in vitro screening protocol to evaluate the TGR5 agonism, GLP-1 secretion, and gut-restricted properties of small molecules. The protocol involves differentiating gut epithelial and endocrine cells together in transwells to assess both the pharmacodynamics of TGR5 agonists and the toxicity of compounds to the intestinal monolayer. As a proof of concept, we demonstrate the use of the protocol in evaluating properties of naturally occurring bile acid metabolites that are potent TGR5 agonists. This protocol is adapted from Chaudhari et al. (2021).

Vitamin D receptor is overexpressed in the duodenum of patients with irritable bowel syndrome.

Irritable bowel syndrome (IBS) is one of the most common functional gastrointestinal disorders, and bile acids are thought to be associated with the pathogenesis of IBS. Bile acid receptors are expressed on intestinal epithelial cells. However, no study has assessed bile acid receptor proteins in IBS. Therefore, we examined the intestinal mucosal expression of bile acid receptors in patients with IBS. Intestinal biopsies were performed in patients with IBS and controls. Mast cells, vitamin D receptor (VDR), and somatostatin were stained with specific antibodies. Levels of VDR, farnesoid X receptor (FXR), takeda-G-protein-receptor-5 (TGR5), claudins, and transient-receptor-potential-cation-channel-subfamily-V-member 6 (TRPV6) were assessed by western blotting. 3Mast cell counts in the second part of the duodenum were significantly higher in patients with IBS than in controls. VDR protein levels were significantly elevated in the duodenum and terminal ileum of patients with IBS compared with controls, although this difference was not seen in the cecum or rectum. FXR and TGR5 protein levels did not differ in any part of the intestine. VDR-positive cryptal epithelia in IBS were distributed not only at basal crypt but also along the upper part of the basal crypt epithelial cells. In contrast, the pattern of gut somatostatin-positive cells, claudins, and TRPV6 levels did not differ. The number of mast cells in the duodenum was significantly increased, and the protein expression levels of VDR, but not those of FXR or TGR5, were elevated in the duodenal epithelial crypt in patients with IBS.

Molecular Physiology of Bile Acid Signaling in Health, Disease, and Aging.

Over the past two decades, bile acids (BAs) have become established as important signaling molecules that enable fine-tuned inter-tissue communication from the liver, their site of production, over the intestine, where they are modified by the gut microbiota, to virtually any organ, where they exert their pleiotropic physiological effects. The chemical variety of BAs, to a large extent determined by the gut microbiome, also allows for a complex fine-tuning of adaptive responses in our body. This review provides an overview of the mechanisms by which BA receptors coordinate several aspects of physiology and highlights new therapeutic strategies for diseases underlying pathological BA signaling.

Reply to “Letter to Editor: Takeda-G Protein Receptor-5 signaling mitigates parenteral nutrition-associated liver disease: public health impact”

Reply to “Letter to Editor: Takeda-G Protein Receptor-5 signaling mitigates parenteral nutrition-associated liver disease: public health impact”

Brown Adipose Tissue, Diet-Induced Thermogenesis, and Thermogenic Food Ingredients: From Mice to Men.

Since the recent rediscovery of brown adipose tissue (BAT) in adult humans, this thermogenic tissue has been attracting increasing interest. The inverse relationship between BAT activity and body fatness suggests that BAT, because of its energy dissipating activity, is protective against body fat accumulation. Cold exposure activates and recruits BAT, resulting in increased energy expenditure and decreased body fatness. The stimulatory effects of cold exposure are mediated through transient receptor potential (TRP) channels and the sympathetic nervous system (SNS). Most TRP members also function as chemesthetic receptors for various food ingredients, and indeed, agonists of TRP vanilloid 1 such as capsaicin and its analog capsinoids mimic the effects of cold exposure to decrease body fatness through the activation and recruitment of BAT. The antiobesity effect of other food ingredients including tea catechins may be attributable, at least in part, to the activation of the TRP–SNS–BAT axis. BAT is also involved in the facultative thermogenesis induced by meal intake, referred to as diet-induced thermogenesis (DIT), which is a significant component of the total energy expenditure in our daily lives. Emerging evidence suggests a crucial role for the SNS in BAT-associated DIT, particularly during the early phase, but several gut-derived humoral factors may also participate in meal-induced BAT activation. One intriguing factor is bile acids, which activate BAT directly through Takeda G-protein receptor 5 (TGR5) in brown adipocytes. Given the apparent beneficial effects of some TRP agonists and bile acids on whole-body substrate and energy metabolism, the TRP/TGR5–BAT axis represents a promising target for combating obesity and related metabolic disorders in humans.

Pharmacological activation of TGR5 promotes intestinal growth via a GLP-2-dependent pathway in mice.

Glucagon-like peptide (GLP)-1 and -2-secreting L cells have been shown to express the bile acid receptor Takeda G protein-receptor-5 (TGR5) and increase secretion upon receptor activation. Previous studies have explored GLP-1 secretion following acute TGR5 activation, but chronic activation and GLP-2 responses have not been characterized. In this study, we aimed to investigate the consequences of pharmacological TGR5 receptor activation on L cell hormone production in vivo using the specific TGR5 agonist RO5527239 and the GLP-2 receptor knockout mouse. Here, we show that 1) TGR5 receptor activation led to increased GLP-1 and GLP-2 content in the colon, which 2) was associated with an increased small intestinal weight that 3) was GLP-2 dependent. Additionally, we report that TGR5-mediated gallbladder filling occurred independently of GLP-2 signaling. In conclusion, we demonstrate that pharmacological TGR5 receptor activation stimulates L cells, triggering GLP-2-dependent intestinal adaption in mice.NEW & NOTEWORTHY Using the specific Takeda G protein-receptor-5 (TGR5) agonist RO5527239 and GLP-2 receptor knockout mice, we show that activation of TGR5 led to the increase in colonic GLP-1 and GLP-2 concomitant with a GLP-2 dependent growth response in the proximal portion of the small intestine.

Evaluation of the Antidiabetic Effect of Clerodendrum volubile P. Beauv leaves: In vivo and In Silico Approach

Clerodendrum volubile leaf is commonly used in traditional medicine for the management of various diseases including diabetes in Nigeria. This study sought to propose the possible mechanism underlying the antidiabetic effect of C. volubile leaves in streptozotocin (STZ)‐induced diabetic rats using in vivo and in silico approach. Aqueous extract of C. volubile was prepared and its effect assessed on relevant enzymes associated with diabetes. Fifty Male Wistar rats (n=5) were randomly separated into ten groups. The induction of diabetes in rats was by a single intraperitoneal injection of STZ (65 mg/kg body weight) while C. volubile extract was administered orally to diabetic and non‐diabetic animals, at the doses of 50,100 and 200 mg/kg body weight respectively for 14 days. Also the interaction of compounds identified from C. volubile (HPLC‐DAD) on Takeda‐G‐protein‐receptor‐5 (TGR5), peroxisome proliferated activated receptor gamma (PPARγ) and dipeptidyl‐peptidase 4 (DPP‐4) was also investigated through molecular docking. Administration of C. volubile extract significantly reduced the elevated plasma glucose level and body weight, improved kidney functions, attenuated oxidative stress by decreasing malondialdehyde (MDA) levels, enhancing superoxide dismutase, catalase and glutathione peroxidase activities, reinstated the lipid profiles to nearly normal level and restored pancreatic histological integrity in diabetic rats. Rutin ranked highest among all the compounds identified in C. volubile with −8.3 kcal/mol binding energy with TGR5 with Glu343 and Glu291 −7.9 kcal/mol (PPARγ) and −9.5.kcal/mol (DPP‐4). Gln77, Arg125, Tyr240, Glu291, Glu343, Tyr 662 are among residues enhancing rutin binding to these proteins. The results revealed that C. volubile possess antidiabetic effects through the modulation of TGR5, PPARγ and DPP4. Rutin might be the lead compounds responsible for the antidiabetic effect of C. volubile justifying the use of this plant in traditional medicine

TGR5 promotes cholangiocarcinoma by interacting with mortalin

Takeda-G-protein-receptor-5 (TGR5) is a G-protein-coupled receptor (GPCR) activated by bile acids, and mortalin is a multipotent chaperone of the HSP70 family. In the present study, TGR5 was detected by immunohistochemistry (IHC) in extrahepatic cholangiocarcinoma (ECC) specimens, and TGR5 expression in ECC tissues and adjacent tissues was compared. In vitro TGR5 was overexpressed and knocked down in human intrahepatic cholangiocarcinoma (ICC) cell line RBE and human extrahepatic cholangiocarcinoma (ECC) cell line QBC-939 to observe its effects on the biological behavior of cholangiocarcinoma (CC) cells, including proliferation, apoptosis and migration. In vivo xenograft model was constructed to explore the role of TGR5 in CC growth. Proteins that interacted with TGR5 were screened using an immunoprecipitation spectrometry approach, and the identified protein was down-regulated to investigate its contribution to CC growth. The present study demonstrated that TGR5 is highly expressed in CC tissues, and strong TGR5 expression may indicate high malignancy in CC. Furthermore, TGR5 promotes CC cell proliferation, migration, and apoptosis resistance. TGR5 boosts CC growth in vivo. In addition, TGR5 combines with mortalin and regulates mortalin expression in the CC cell line. Mortalin participates in the TGR5-induced increase in CC cell proliferation. In conclusion, TGR5 is of clinical significance based on its implications for the degree of malignancy in patients with CC. Mortalin may be a downstream component regulated by TGR5, and TGR5 promotes cholangiocarcinoma at least partially by interacting with mortalin and upregulating its expression. Both TGR5 and mortalin are positive regulators, and may serve as potential therapeutic targets for CC.

TGR5 signaling mitigates parenteral nutrition-associated liver disease.

Bile acid receptors regulate the metabolic and immune functions of circulating enterohepatic bile acids. This process is disrupted by administration of parenteral nutrition (PN), which may induce progressive hepatic injury for unclear reasons, especially in the newborn, leading to PN-associated liver disease. To explore the role of bile acid signaling on neonatal hepatic function, we initially observed that Takeda G protein receptor 5 (TGR5)-specific bile acids were negatively correlated with worsening clinical disease markers in the plasma of human newborns with prolonged PN exposure. To test our resulting hypothesis that TGR5 regulates critical liver functions to PN exposure, we used TGR5 receptor deficient mice (TGR5-/-). We observed PN significantly increased liver weight, cholestasis, and serum hepatic stress enzymes in TGR5-/- mice compared with controls. Mechanistically, PN reduced bile acid synthesis genes in TGR5-/-. Serum bile acid composition revealed that PN increased unconjugated primary bile acids and secondary bile acids in TGR5-/- mice, while increasing conjugated primary bile acid levels in TGR5-competent mice. Simultaneously, PN elevated hepatic IL-6 expression and infiltrating macrophages in TGR5-/- mice. However, the gut microbiota of TGR5-/- mice compared with WT mice following PN administration displayed highly elevated levels of Bacteroides and Parabacteroides, and possibly responsible for the elevated levels of secondary bile acids in TGR5-/- animals. Intestinal bile acid transporters expression was unchanged. Collectively, this suggests TGR5 signaling specifically regulates fundamental aspects of liver bile acid homeostasis during exposure to PN. Loss of TGR5 is associated with biochemical evidence of cholestasis in both humans and mice on PN.NEW & NOTEWORTHY Parenteral nutrition is associated with deleterious metabolic outcomes in patients with prolonged exposure. Here, we demonstrate that accelerated cholestasis and parental nutrition-associated liver disease (PNALD) may be associated with deficiency of Takeda G protein receptor 5 (TGR5) signaling. The microbiome is responsible for production of secondary bile acids that signal through TGR5. Therefore, collectively, these data support the hypothesis that a lack of established microbiome in early life or under prolonged parenteral nutrition may underpin disease development and PNALD.

Takeda G-protein Receptor (TGR)-5 Evolves Classical Activestate Conformational Signatures in Complex with Chromolaena Odorata-derived Flavonoid-5,7-dihydroxy-6-4-dimethoxyflavanone

Background: Takeda G-protein receptor 5 (TGR5) via glucagon-like peptide release and insulin signaling underlies antidiabetic roles of TGR5 agonists. Chromolaena Odorata- derived flavonoid-5,7-dihydroxy-6-4-dimethoxyflavanone (COF) has been identified as (TGR5) agonist. The structural basis for their interaction has not been studied. Objective: This study aimed at providing both structural and dynamic insights into COF/TGR5 interaction. Methods: Classical GPCR activation signatures (TMIII-TMVI ionic lock, toggle switches, internal water pathway) using classical MD simulation have been used. Results: Y893.29, N933.33 and E1695.43 are key residues found to be involved in ligand binding; the continuous internal water pathway connects hydrophilic groups of the ligand to the TMIII-TMVI interface in COF-bound state, TMIII-TMVI ionic locks ruptures in COF-TGR5 complex but not antagonist-bound state, and ruptured ionic lock is associated with the evolution of active-state “VPVAM” (analogous to “NPxxY”) conformation. Dihedral angles (c2) calculated along the trajectory strongly suggest W2376.48 as a ligand-dependent toggle switch. Conclusion: TGR5 evolves active state conformation from a starting intermediate state conformation when bound to COF, which further supports its underlying anti-diabetic activities.

Chenodeoxycholic Acid (CDCA) Protects against the Lipopolysaccharide-Induced Impairment of the Intestinal Epithelial Barrier Function via the FXR-MLCK Pathway.

Chenodeoxycholic acid (CDCA), a primary bile acid, has been demonstrated to play important roles as a signaling molecule in various physiology functions. However, the role of CDCA in regulating intestinal barrier function remains largely unknown. This study aimed to investigate the effects of CDCA on the lipopolysaccharide (LPS)-impaired intestinal epithelial barrier function and explore the underlying mechanisms. In IPEC-J2 cells, CDCA reversed the LPS-induced increase in transepithelial electrical resistance and decrease in tight junction protein expression. In addition, we found that farnesoid X receptor (FXR) but not Takeda G-protein receptor 5 was responsible for the CDCA-improved epithelial barrier function impaired by LPS. Furthermore, CDCA blocked LPS-induced activation of the myosin light chain kinase (MLCK) pathway in a FXR-dependent manner and elicited similar effects to MLCK inhibition. In mice, CDCA supplementation restored LPS-induced elevation of intestinal permeability and MLCK expression and reduction of tight junction protein expression, thus alleviating LPS-induced intestinal barrier impairment. In conclusion, CDCA protected against the LPS-induced impairment of the intestinal epithelial barrier function via the FXR-MLCK pathway.