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Sample records for acid receptor farnesoid

  1. Bile Acid Nuclear Receptor Farnesoid X Receptor: Therapeutic Target for Nonalcoholic Fatty Liver Disease

    PubMed Central

    Kim, Sun-Gi; Kim, Byung-Kwon; Kim, Kyumin

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is one of the causes of fatty liver, occurring when fat is accumulated in the liver without alcohol consumption. NAFLD is the most common liver disorder in advanced countries. NAFLD is a spectrum of pathology involving hepatic steatosis with/without inflammation and nonalcoholic steatohepatitis with accumulation of hepatocyte damage and hepatic fibrosis. Recent studies have revealed that NAFLD results in the progression of cryptogenic cirrhosis that leads to hepatocarcinoma and cardiovascular diseases such as heart failure. The main causes of NAFLD have not been revealed yet, metabolic syndromes including obesity and insulin resistance are widely accepted for the critical risk factors for the pathogenesis of NAFLD. Nuclear receptors (NRs) are transcriptional factors that sense environmental or hormonal signals and regulate expression of genes, involved in cellular growth, development, and metabolism. Several NRs have been reported to regulate genes involved in energy and xenobiotic metabolism and inflammation. Among various NRs, farnesoid X receptor (FXR) is abundantly expressed in the liver and a key regulator to control various metabolic processes in the liver. Recent studies have shown that NAFLD is associated with inappropriate function of FXR. The impact of FXR transcriptional activity in NAFLD is likely to be potential therapeutic strategy, but still requires to elucidate underlying potent therapeutic mechanisms of FXR for the treatment of NAFLD. This article will focus the physiological roles of FXR and establish the correlation between FXR transcriptional activity and the pathogenesis of NAFLD. PMID:28029021

  2. Lithocholic acid decreases expression of bile salt export pump through farnesoid X receptor antagonist activity.

    PubMed

    Yu, Jinghua; Lo, Jane-L; Huang, Li; Zhao, Annie; Metzger, Edward; Adams, Alan; Meinke, Peter T; Wright, Samuel D; Cui, Jisong

    2002-08-30

    Bile salt export pump (BSEP) is a major bile acid transporter in the liver. Mutations in BSEP result in progressive intrahepatic cholestasis, a severe liver disease that impairs bile flow and causes irreversible liver damage. BSEP is a target for inhibition and down-regulation by drugs and abnormal bile salt metabolites, and such inhibition and down-regulation may result in bile acid retention and intrahepatic cholestasis. In this study, we quantitatively analyzed the regulation of BSEP expression by FXR ligands in primary human hepatocytes and HepG2 cells. We demonstrate that BSEP expression is dramatically regulated by ligands of the nuclear receptor farnesoid X receptor (FXR). Both the endogenous FXR agonist chenodeoxycholate (CDCA) and synthetic FXR ligand GW4064 effectively increased BSEP mRNA in both cell types. This up-regulation was readily detectable at as early as 3 h, and the ligand potency for BSEP regulation correlates with the intrinsic activity on FXR. These results suggest BSEP as a direct target of FXR and support the recent report that the BSEP promoter is transactivated by FXR. In contrast to CDCA and GW4064, lithocholate (LCA), a hydrophobic bile acid and a potent inducer of cholestasis, strongly decreased BSEP expression. Previous studies did not identify LCA as an FXR antagonist ligand in cells, but we show here that LCA is an FXR antagonist with partial agonist activity in cells. In an in vitro co-activator association assay, LCA decreased CDCA- and GW4064-induced FXR activation with an IC(50) of 1 microm. In HepG2 cells, LCA also effectively antagonized GW4064-enhanced FXR transactivation. These data suggest that the toxic and cholestatic effect of LCA in animals may result from its down-regulation of BSEP through FXR. Taken together, these observations indicate that FXR plays an important role in BSEP gene expression and that FXR ligands may be potential therapeutic drugs for intrahepatic cholestasis.

  3. Effect of common polymorphisms of the farnesoid X receptor and bile acid transporters on the pharmacokinetics of ursodeoxycholic acid.

    PubMed

    Hu, Miao; Fok, Benny S P; Wo, Siu-Kwan; Lee, Vincent H L; Zuo, Zhong; Tomlinson, Brian

    2016-01-01

    Ursodeoxycholic acid (UDCA), a natural, dihydroxy bile acid, promotes gallstone dissolution and has been attributed with several other beneficial effects. The farnesoid X receptor (FXR) may influence the pharmacokinetics of UDCA by modulating the expression of bile acid transporters. This exploratory study examined whether common functional polymorphisms in FXR and in bile acid transporter genes affect the pharmacokinetics of exogenous UDCA. Polymorphisms in genes for transporters involved in bile acid transport, solute carrier organic anion 1B1 (SLCO1B1) 388A>G and 521T>C, solute carrier 10A1 (SLC10A1) 800 C>T and ATP-binding cassette B11 (ABCB11) 1331T>C, and the FXR -1G>T polymorphism were genotyped in 26 male Chinese subjects who ingested single oral 500-mg doses of UDCA. Plasma concentrations of UDCA and its major conjugate metabolite glycoursodeoxycholic acid (GUDCA) were determined. The mean systemic exposure of UDCA was higher in the five subjects with one copy of the FXR -1G>T variant allele than in those homozygous for the wild-type allele (n = 21) (AUC0-24 h : 38.5 ± 28.2 vs. 20.9 ± 8.0 μg h/mL, P = 0.021), but this difference appeared mainly due to one outlier with the -1GT genotype and elevated baseline and post-treatment UDCA concentrations. After excluding the outlier, body weight was the only factor associated with plasma concentrations of UDCA and there were no significant associations with the other polymorphisms examined. None of the polymorphisms affected the pharmacokinetics of GUDCA. This study showed that the common polymorphisms in bile acid transporters had no significant effect on the pharmacokinetics of exogenous UDCA but an effect of the FXR polymorphism cannot be excluded.

  4. Ursodeoxycholic Acid Suppresses Lipogenesis in Mouse Liver: Possible Role of the Decrease in β-Muricholic Acid, a Farnesoid X Receptor Antagonist.

    PubMed

    Fujita, Kyosuke; Iguchi, Yusuke; Une, Mizuho; Watanabe, Shiro

    2017-03-17

    The farnesoid X receptor (FXR) is a major nuclear receptor of bile acids; its activation suppresses sterol regulatory element-binding protein 1c (SREBP1c)-mediated lipogenesis and decreases the lipid contents in the liver. There are many reports showing that the administration of ursodeoxycholic acid (UDCA) suppresses lipogenesis and reduces the lipid contents in the liver of experimental animals. Since UDCA is not recognized as an FXR agonist, these effects of UDCA cannot be readily explained by its direct activation of FXR. We observed that the dietary administration of UDCA in mice decreased the expression levels of SREBP1c and its target lipogenic genes. Alpha- and β-muricholic acids (MCA) and cholic acid (CA) were the major bile acids in the mouse liver but their contents decreased upon UDCA administration. The hepatic contents of chenodeoxycholic acid and deoxycholic acid (DCA) were relatively low but were not changed by UDCA. UDCA did not show FXR agonistic or antagonistic potency in in vitro FXR transactivation assay. Taking these together, we deduced that the above-mentioned change in hepatic bile acid composition induced upon UDCA administration might cause the relative increase in the FXR activity in the liver, mainly by the reduction in the content of β-MCA, a farnesoid X receptor antagonist, which suggests a mechanism by which UDCA suppresses lipogenesis and decreases the lipid contents in the mouse liver.

  5. Differential regulation of bile acid and cholesterol metabolism by the farnesoid X receptor in Ldlr -/- mice versus hamsters.

    PubMed

    Gardès, Christophe; Chaput, Evelyne; Staempfli, Andreas; Blum, Denise; Richter, Hans; Benson, G Martin

    2013-05-01

    Modulating bile acid synthesis has long been considered a good strategy by which to improve cholesterol homeostasis in humans. The farnesoid X receptor (FXR), the key regulator of bile acid synthesis, was, therefore, identified as an interesting target for drug discovery. We compared the effect of four, structurally unrelated, synthetic FXR agonists in two fat-fed rodent species and observed that the three most potent and selective agonists decreased plasma cholesterol in LDL receptor-deficient (Ldlr (-/-)) mice, but none did so in hamsters. Detailed investigation revealed increases in the expression of small heterodimer partner (Shp) in their livers and of intestinal fibroblast growth factor 15 or 19 (Fgf15/19) in mice only. Cyp7a1 expression and fecal bile acid (BA) excretion were strongly reduced in mice and hamsters by all four FXR agonists, whereas bile acid pool sizes were reduced in both species by all but the X-Ceptor compound in hamsters. In Ldlr (-/-) mice, the predominant bile acid changed from cholate to the more hydrophilic β-muricholate due to a strong repression of Cyp8b1 and increase in Cyp3a11 expression. However, FXR agonists caused only minor changes in the expression of Cyp8b1 and in bile acid profiles in hamsters. In summary, FXR agonist-induced decreases in bile acid pool size and lipophilicity and in cholesterol absorption and synthesis could explain the decreased plasma cholesterol in Ldlr (-/-) mice. In hamsters, FXR agonists reduced bile acid pool size to a smaller extent with minor changes in bile acid profile and reductions in sterol absorption, and consequently, plasma cholesterol was unchanged.

  6. Farnesoid X receptor signal is involved in deoxycholic acid-induced intestinal metaplasia of normal human gastric epithelial cells.

    PubMed

    Li, Shu; Chen, Xin; Zhou, Lu; Wang, Bang-Mao

    2015-11-01

    The farnesoid X receptor (FXR) signaling pathway is known to be involved in the metabolism of bile acid, glucose and lipid. In the present study, we demonstrated that 400 µmol/l deoxycholic acid (DCA) stimulation promotes the proliferation of normal human gastric epithelial cells (GES-1). In addition, DCA activated FXR and increased the expression of intestinal metaplasia genes, including caudal-related homeobox transcription factor 2 (Cdx2) and mucin 2 (MUC2). The treatment of FXR agonist GW4064/antagonist guggulsterone (Gug.) significantly increased/decreased the expression levels of FXR, Cdx2 and MUC2 protein in DCA-induced GES-1 cells. GW4064/Gug. also enhanced/reduced the nuclear factor-κB (NF-κB) activity and binding of the Cdx2 promoter region and NF-κB, the most common subunit p50 protein. Taken together, the results indicated that DCA is capable of modulating the expression of Cdx2 and the downstream MUC2 via the nuclear receptor FXR-NF-κB activity in normal gastric epithelial cells. FXR signaling pathway may therefore be involved in the intestinal metaplasia of human gastric mucosa.

  7. Bile acids override steatosis in farnesoid X receptor deficient mice in a model of non-alcoholic steatohepatitis

    SciTech Connect

    Wu, Weibin; Liu, Xijun; Peng, Xiaomin; Xue, Ruyi; Ji, Lingling; Shen, Xizhong; Chen, She; Gu, Jianxin; Zhang, Si

    2014-05-23

    Highlights: • FXR deficiency enhanced MCD diet-induced hepatic fibrosis. • FXR deficiency attenuated MCD diet-induced hepatic steatosis. • FXR deficiency repressed genes involved in fatty acid uptake and triglyceride accumulation. - Abstract: Non-alcoholic fatty liver disease (NAFLD) is one of the most common liver diseases, and the pathogenesis is still not well known. The farnesoid X receptor (FXR) is a member of the nuclear hormone receptor superfamily and plays an essential role in maintaining bile acid and lipid homeostasis. In this study, we study the role of FXR in the pathogenesis of NFALD. We found that FXR deficient (FXR{sup −/−}) mice fed methionine- and choline-deficient (MCD) diet had higher serum ALT and AST activities and lower hepatic triglyceride levels than wild-type (WT) mice fed MCD diet. Expression of genes involved in inflammation (VCAM-1) and fibrosis (α-SMA) was increased in FXR{sup −/−} mice fed MCD diet (FXR{sup −/−}/MCD) compared to WT mice fed MCD diet (WT/MCD). Although MCD diet significantly induced hepatic fibrosis in terms of liver histology, FXR{sup −/−}/MCD mice showed less degree of hepatic steatosis than WT/MCD mice. Moreover, FXR deficiency synergistically potentiated the elevation effects of MCD diet on serum and hepatic bile acids levels. The super-physiological concentrations of hepatic bile acids in FXR{sup −/−}/MCD mice inhibited the expression of genes involved in fatty acid uptake and triglyceride accumulation, which may be an explanation for less steatosis in FXR{sup −/−}/MCD mice in contrast to WT/MCD mice. These results suggest that hepatic bile acids accumulation could override simple steatosis in hepatic injury during the progression of NAFLD and further emphasize the role of FXR in maintaining hepatic bile acid homeostasis in liver disorders and in hepatic protection.

  8. Morphologic Damage of Rat Alveolar Epithelial Type II Cells Induced by Bile Acids Could Be Ameliorated by Farnesoid X Receptor Inhibitor Z-Guggulsterone In Vitro

    PubMed Central

    Huang, Yaowei; Hou, Xusheng; Wu, Wenyu; Nie, Lei; Tian, Yinghong; Lu, Yanmeng

    2016-01-01

    Objective. To determine whether bile acids (BAs) affect respiratory functions through the farnesoid X receptor (FXR) expressed in the lungs and to explore the possible mechanisms of BAs-induced respiratory disorder. Methods. Primary cultured alveolar epithelial type II cells (AECIIs) of rat were treated with different concentrations of chenodeoxycholic acid (CDCA) in the presence or absence of FXR inhibitor Z-guggulsterone (GS). Then, expression of FXR in nuclei of AECIIs was assessed by immunofluorescence microscopy. And ultrastructural changes of the cells were observed under transmission electron microscope and analyzed by Image-Pro Plus software. Results. Morphologic damage of AECIIs was exhibited in high BAs group in vitro, with high-level expression of FXR, while FXR inhibitor GS could attenuate the cytotoxicity of BAs to AECIIs. Conclusions. FXR expression was related to the morphologic damage of AECIIs induced by BAs, thus influencing respiratory functions. PMID:27340672

  9. Farnesoid X nuclear receptor ligand obeticholic acid for non-cirrhotic, non-alcoholic steatohepatitis (FLINT): a multicentre, randomised, placebo-controlled trial

    PubMed Central

    Neuschwander-Tetri, Brent A; Loomba, Rohit; Sanyal, Arun J; Lavine, Joel E; Van Natta, Mark L; Abdelmalek, Manal F; Chalasani, Naga; Dasarathy, Srinivasan; Diehl, Anna Mae; Hameed, Bilal; Kowdley, Kris V; McCullough, Arthur; Terrault, Norah; Clark, Jeanne M; Tonascia, James; Brunt, Elizabeth M; Kleiner, David E; Doo, Edward

    2015-01-01

    Summary Background The bile acid derivative 6-ethylchenodeoxycholic acid (obeticholic acid) is a potent activator of the farnesoid X nuclear receptor that reduces liver fat and fibrosis in animal models of fatty liver disease. We assessed the efficacy of obeticholic acid in adult patients with non-alcoholic steatohepatitis. Methods We did a multicentre, double-blind, placebo-controlled, parallel group, randomised clinical trial at medical centres in the USA in patients with non-cirrhotic, non-alcoholic steatohepatitis to assess treatment with obeticholic acid given orally (25 mg daily) or placebo for 72 weeks. Patients were randomly assigned 1:1 using a computer-generated, centrally administered procedure, stratified by clinical centre and diabetes status. The primary outcome measure was improvement in centrally scored liver histology defined as a decrease in non-alcoholic fatty liver disease activity score by at least 2 points without worsening of fibrosis from baseline to the end of treatment. A planned interim analysis of change in alanine aminotransferase at 24 weeks undertaken before end-of-treatment (72 weeks) biopsies supported the decision to continue the trial (relative change in alanine aminotransferase −24%, 95% CI −45 to −3). A planned interim analysis of the primary outcome showed improved efficacy of obeticholic acid (p=0·0024) and supported a decision not to do end-of-treatment biopsies and end treatment early in 64 patients, but to continue the trial to obtain the 24-week post-treatment measures. Analyses were done by intention-to-treat. This trial was registered with ClinicalTrials.gov, number NCT01265498. Findings Between March 16, 2011, and Dec 3, 2012, 141 patients were randomly assigned to receive obeticholic acid and 142 to placebo. 50 (45%) of 110 patients in the obeticholic acid group who were meant to have biopsies at baseline and 72 weeks had improved liver histology compared with 23 (21%) of 109 such patients in the placebo group

  10. Recent advances in the development of farnesoid X receptor agonists

    PubMed Central

    Carey, Elizabeth J.; Lindor, Keith D.

    2015-01-01

    Farnesoid X receptors (FXRs) are nuclear hormone receptors expressed in high amounts in body tissues that participate in bilirubin metabolism including the liver, intestines, and kidneys. Bile acids (BAs) are the natural ligands of the FXRs. FXRs regulate the expression of the gene encoding for cholesterol 7 alpha-hydroxylase, which is the rate-limiting enzyme in BA synthesis. In addition, FXRs play a critical role in carbohydrate and lipid metabolism and regulation of insulin sensitivity. FXRs also modulate live growth and regeneration during liver injury. Preclinical studies have shown that FXR activation protects against cholestasis-induced liver injury. Moreover, FXR activation protects against fatty liver injury in animal models of nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH), and improved hyperlipidemia, glucose intolerance, and insulin sensitivity. Obeticholic acid (OCA), a 6α-ethyl derivative of the natural human BA chenodeoxycholic acid (CDCA) is the first-in-class selective FXR agonist that is ~100-fold more potent than CDCA. Preliminary human clinical trials have shown that OCA is safe and effective. In a phase II clinical trial, administration of OCA was well-tolerated, increased insulin sensitivity and reduced markers of liver inflammation and fibrosis in patients with type II diabetes mellitus and NAFLD. In two clinical trials of OCA in patients with primary biliary cirrhosis (PBC), a progressive cholestatic liver disease, OCA significantly reduced serum alkaline phosphatase (ALP) levels, an important disease marker that correlates well with clinical outcomes of patients with PBC. Together, these studies suggest that FXR agonists could potentially be used as therapeutic tools in patients suffering from nonalcoholic fatty and cholestatic liver diseases. Larger and Longer-term studies are currently ongoing. PMID:25705637

  11. Farnesoid X receptor represses hepatic human APOA gene expression

    PubMed Central

    Chennamsetty, Indumathi; Claudel, Thierry; Kostner, Karam M.; Baghdasaryan, Anna; Kratky, Dagmar; Levak-Frank, Sanja; Frank, Sasa; Gonzalez, Frank J.; Trauner, Michael; Kostner, Gert M.

    2011-01-01

    High plasma concentrations of lipoprotein(a) [Lp(a), which is encoded by the APOA gene] increase an individual’s risk of developing diseases, such as coronary artery diseases, restenosis, and stroke. Unfortunately, increased Lp(a) levels are minimally influenced by dietary changes or drug treatment. Further, the development of Lp(a)-specific medications has been hampered by limited knowledge of Lp(a) metabolism. In this study, we identified patients suffering from biliary obstructions with very low plasma Lp(a) concentrations that rise substantially after surgical intervention. Consistent with this, common bile duct ligation in mice transgenic for human APOA (tg-APOA mice) lowered plasma concentrations and hepatic expression of APOA. To test whether farnesoid X receptor (FXR), which is activated by bile acids, was responsible for the low plasma Lp(a) levels in cholestatic patients and mice, we treated tg-APOA and tg-APOA/Fxr–/– mice with cholic acid. FXR activation markedly reduced plasma concentrations and hepatic expression of human APOA in tg-APOA mice but not in tg-APOA/Fxr–/– mice. Incubation of primary hepatocytes from tg-APOA mice with bile acids dose dependently downregulated APOA expression. Further analysis determined that the direct repeat 1 element between nucleotides –826 and –814 of the APOA promoter functioned as a negative FXR response element. This motif is also bound by hepatocyte nuclear factor 4α (HNF4α), which promotes APOA transcription, and FXR was shown to compete with HNF4α for binding to this motif. These findings may have important implications in the development of Lp(a)-lowering medications. PMID:21804189

  12. 5{alpha}-Bile alcohols function as farnesoid X receptor antagonists

    SciTech Connect

    Nishimaki-Mogami, Tomoko . E-mail: mogami@nihs.go.jp; Kawahara, Yosuke; Tamehiro, Norimasa; Yoshida, Takemi; Inoue, Kazuhide; Ohno, Yasuo; Nagao, Taku; Une, Mizuho

    2006-01-06

    The farnesoid X receptor (FXR) is a bile acid/alcohol-activated nuclear receptor that regulates lipid homeostasis. Unlike other steroid receptors, FXR binds bile acids in an orientation that allows the steroid nucleus A to face helix 12 in the receptor, a crucial domain for coactivator-recruitment. Because most naturally occurring bile acids and alcohols contain a cis-oriented A, which is distinct from that of other steroids and cholesterol metabolites, we investigated the role of this 5{beta}-configuration in FXR activation. The results showed that the 5{beta}-(A/B cis) bile alcohols 5{beta}-cyprinol and bufol are potent FXR agonists, whereas their 5{alpha}-(A/B trans) counterparts antagonize FXR transactivation and target gene expression. Both isomers bound to FXR, but their ability to induce coactivator-recruitment and thereby induce transactivation differed. These findings suggest a critical role for the A orientation of bile salts in agonist/antagonist function.

  13. A tea catechin, epigallocatechin-3-gallate, is a unique modulator of the farnesoid X receptor

    SciTech Connect

    Li, Guodong; Lin, Wenwei; Araya, Juan J.; Chen, Taosheng; Timmermann, Barbara N.; Guo, Grace L.

    2012-01-15

    Farnesoid X receptor (FXR) is a ligand-activated nuclear receptor and serves as a key regulator to maintain health of the liver and intestine. Bile acids are endogenous ligands of FXR, and there are increasing efforts to identify FXR modulators to serve as biological probes and/or pharmaceutical agents. Natural FXR ligands isolated from plants may serve as models to synthesize novel FXR modulators. In this study, we demonstrated that epigallocatechin-3-gallate (EGCG), a major tea catechin, specifically and dose-dependently activates FXR. In addition, EGCG induced FXR target gene expression in vitro. Surprisingly, in a co-activator (SRC2) recruitment assay, we found that EGCG does not recruit SRC2 to FXR, but it dose-dependently inhibits recruitment of SRC2 to FXR (IC{sub 50}, 1 μM) by GW6064, which is a potent FXR synthetic ligand. In addition, EGCG suppressed FXR target gene expression induced by either GW4064 or chenodeoxycholic acid in vitro. Furthermore, wild-type and FXR knockout mice treated with an acute dose of EGCG had induced mRNA expression in a subset of FXR target genes in the intestine but not in the liver. In conclusion, EGCG is a unique modulator of FXR in the intestine and may serve as an important model for future development of FXR modulators. -- Highlights: ► Epigallocatechin-3-gallate (EGCG) is a unique farnesoid X receptor (FXR) modulator. ► EGCG activates FXR by itself, but inhibits FXR transactivation by other agonists. ► Low concentration of EGCG activates FXR in mouse intestine but not liver. ► EGCG activates FXR to induce a subset of FXR target genes in mouse intestine.

  14. Quantitative High-Throughput Profiling of Environmental Chemicals and Drugs that Modulate Farnesoid X Receptor

    PubMed Central

    Hsu, Chia-Wen; Zhao, Jinghua; Huang, Ruili; Hsieh, Jui-Hua; Hamm, Jon; Chang, Xiaoqing; Houck, Keith; Xia, Menghang

    2014-01-01

    The farnesoid X receptor (FXR) regulates the homeostasis of bile acids, lipids, and glucose. Because endogenous chemicals bind and activate FXR, it is important to examine which xenobiotic compounds would disrupt normal receptor function. We used a cell-based human FXR β-lactamase (Bla) reporter gene assay to profile the Tox21 10K compound collection of environmental chemicals and drugs. Structure-activity relationships of FXR-active compounds revealed by this screening were then compared against the androgen receptor, estrogen receptor α, peroxisome proliferator-activated receptors δ and γ, and the vitamin D receptor. We identified several FXR-active structural classes including anthracyclines, benzimidazoles, dihydropyridines, pyrethroids, retinoic acids, and vinca alkaloids. Microtubule inhibitors potently decreased FXR reporter gene activity. Pyrethroids specifically antagonized FXR transactivation. Anthracyclines affected reporter activity in all tested assays, suggesting non-specific activity. These results provide important information to prioritize chemicals for further investigation, and suggest possible modes of action of compounds in FXR signaling. PMID:25257666

  15. The farnesoid X receptor is expressed in breast cancer and regulates apoptosis and aromatase expression.

    PubMed

    Swales, Karen E; Korbonits, Márta; Carpenter, Robert; Walsh, Desmond T; Warner, Timothy D; Bishop-Bailey, David

    2006-10-15

    Bile acids are present at high concentrations in breast cysts and in the plasma of postmenopausal women with breast cancer. The farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily that regulates bile acid homeostasis. FXR was detected in normal and tumor breast tissue, with a high level of expression in ductal epithelial cells of normal breast and infiltrating ductal carcinoma cells. FXR was also present in the human breast carcinoma cells, MCF-7 and MDA-MB-468. Activation of FXR by high concentrations of ligands induced MCF-7 and MDA-MB-468 apoptosis. At lower concentrations that had no direct effect on viability, the FXR agonist GW4064 induced expression of mRNA for the FXR target genes, small heterodimer partner (SHP), intestinal bile acid binding protein, and multidrug resistance-associated protein 2 (MRP-2), and repressed the expression of the SHP target gene aromatase. In contrast to MRP-2, mRNA for the breast cancer target genes MDR-3, MRP-1, and solute carrier transporter 7A5 were decreased. Although multidrug resistance transporters were regulated and are known FXR target genes, GW4064 had no effect on the cell death induced by the anticancer drug paclitaxel. Our findings show for the first time that FXR is expressed in breast cancer tissue and has multiple properties that could be used for the treatment of breast cancer.

  16. Ku proteins function as corepressors to regulate farnesoid X receptor-mediated gene expression

    SciTech Connect

    Ohno, Masae; Kunimoto, Masaaki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2009-12-18

    The farnesoid X receptor (FXR; NR1H4) is a member of the nuclear receptor superfamily and regulates the expression of genes involved in enterohepatic circulation and the metabolism of bile acids. Based on functional analyses, nuclear receptors are divided into regions A-F. To explore the cofactors interacting with FXR, we performed a pull-down assay using GST-fused to the N-terminal A/B region and the C region, which are required for the ligand-independent transactivation and DNA-binding, respectively, of FXR, and nuclear extracts from HeLa cells. We identified DNA-dependent protein kinase catalytic subunit (DNA-PKcs), Ku80, and Ku70 as FXR associated factors. These proteins are known to have an important role in DNA repair, recombination, and transcription. DNA-PKcs mainly interacted with the A/B region of FXR, whereas the Ku proteins interacted with the C region and with the D region (hinge region). Chromatin immunoprecipitation assays revealed that the Ku proteins associated with FXR on the bile salt export pump (BSEP) promoter. Furthermore, we demonstrated that ectopic expression of the Ku proteins decreased the promoter activity and expression of BSEP gene mediated by FXR. These results suggest that the Ku proteins function as corepressors for FXR.

  17. Expression and activation of the farnesoid X receptor in the vasculature

    NASA Astrophysics Data System (ADS)

    Bishop-Bailey, David; Walsh, Desmond T.; Warner, Timothy D.

    2004-03-01

    The farnesoid X receptor/bile acid receptor (FXR) is a recently discovered member of the nuclear hormone superfamily. FXR ligands have been proposed as targets in cardiovascular disease, regulating cholesterol metabolism and bile acid transport and metabolism in the liver and gastrointestinal tract. When we used a human cardiovascular tissue array, we found that FXR is expressed in a variety of normal and pathological human tissue. Particularly high levels of FXR were found in the vasculature and in a number of different metastatic cancers, as well as the previously identified target tissues of the liver, small intestine, and kidney. In vitro, FXR is present in rat and human vascular smooth muscle cells. When treated with a range of FXR ligands, vascular smooth muscle cells undergo apoptosis in a manner that correlates with the ligands' ability to activate FXR. Furthermore, FXR activators induce mRNA for the FXR target genes, phospholipid transfer protein, and the small heterodimer partner. FXR therefore is a functional protein in the vasculature that may provide a direct target for the treatment of proliferative and dyslipidaemic diseases.

  18. Farnesoid X Receptor Inhibits the Transcriptional Activity of Carbohydrate Response Element Binding Protein in Human Hepatocytes

    PubMed Central

    Caron, Sandrine; Huaman Samanez, Carolina; Dehondt, Hélène; Ploton, Maheul; Briand, Olivier; Lien, Fleur; Dorchies, Emilie; Dumont, Julie; Postic, Catherine; Cariou, Bertrand; Lefebvre, Philippe

    2013-01-01

    The glucose-activated transcription factor carbohydrate response element binding protein (ChREBP) induces the expression of hepatic glycolytic and lipogenic genes. The farnesoid X receptor (FXR) is a nuclear bile acid receptor controlling bile acid, lipid, and glucose homeostasis. FXR negatively regulates hepatic glycolysis and lipogenesis in mouse liver. The aim of this study was to determine whether FXR regulates the transcriptional activity of ChREBP in human hepatocytes and to unravel the underlying molecular mechanisms. Agonist-activated FXR inhibits glucose-induced transcription of several glycolytic genes, including the liver-type pyruvate kinase gene (L-PK), in the immortalized human hepatocyte (IHH) and HepaRG cell lines. This inhibition requires the L4L3 region of the L-PK promoter, known to bind the transcription factors ChREBP and hepatocyte nuclear factor 4α (HNF4α). FXR interacts directly with ChREBP and HNF4α proteins. Analysis of the protein complex bound to the L4L3 region reveals the presence of ChREBP, HNF4α, FXR, and the transcriptional coactivators p300 and CBP at high glucose concentrations. FXR activation does not affect either FXR or HNF4α binding to the L4L3 region but does result in the concomitant release of ChREBP, p300, and CBP and in the recruitment of the transcriptional corepressor SMRT. Thus, FXR transrepresses the expression of genes involved in glycolysis in human hepatocytes. PMID:23530060

  19. Activation of the farnesoid X receptor induces hepatic expression and secretion of fibroblast growth factor 21.

    PubMed

    Cyphert, Holly A; Ge, Xuemei; Kohan, Alison B; Salati, Lisa M; Zhang, Yanqiao; Hillgartner, F Bradley

    2012-07-20

    Previous studies have shown that starvation or consumption of a high fat, low carbohydrate (HF-LC) ketogenic diet induces hepatic fibroblast growth factor 21 (FGF21) gene expression in part by activating the peroxisome proliferator-activated receptor-α (PPARα). Using primary hepatocyte cultures to screen for endogenous signals that mediate the nutritional regulation of FGF21 expression, we identified two sources of PPARα activators (i.e. nonesterified unsaturated fatty acids and chylomicron remnants) that induced FGF21 gene expression. In addition, we discovered that natural (i.e. bile acids) and synthetic (i.e. GW4064) activators of the farnesoid X receptor (FXR) increased FGF21 gene expression and secretion. The effects of bile acids were additive with the effects of nonesterified unsaturated fatty acids in regulating FGF21 expression. FXR activation of FGF21 gene transcription was mediated by an FXR/retinoid X receptor binding site in the 5'-flanking region of the FGF21 gene. FGF19, a gut hormone whose expression and secretion is induced by intestinal bile acids, also increased hepatic FGF21 secretion. Deletion of FXR in mice suppressed the ability of an HF-LC ketogenic diet to induce hepatic FGF21 gene expression. The results of this study identify FXR as a new signaling pathway activating FGF21 expression and provide evidence that FXR activators work in combination with PPARα activators to mediate the stimulatory effect of an HF-LC ketogenic diet on FGF21 expression. We propose that the enhanced enterohepatic flux of bile acids during HF-LC consumption leads to activation of hepatic FXR and FGF19 signaling activity and an increase in FGF21 gene expression and secretion.

  20. Farnesoid X Receptor Signaling Shapes the Gut Microbiota and Controls Hepatic Lipid Metabolism

    PubMed Central

    Zhang, Limin; Xie, Cen; Nichols, Robert G.; Chan, Siu H. J.; Jiang, Changtao; Hao, Ruixin; Smith, Philip B.; Cai, Jingwei; Simons, Margaret N.; Hatzakis, Emmanuel; Maranas, Costas D.; Gonzalez, Frank J.

    2016-01-01

    ABSTRACT The gut microbiota modulates obesity and associated metabolic phenotypes in part through intestinal farnesoid X receptor (FXR) signaling. Glycine-β-muricholic acid (Gly-MCA), an intestinal FXR antagonist, has been reported to prevent or reverse high-fat diet (HFD)-induced and genetic obesity, insulin resistance, and fatty liver; however, the mechanism by which these phenotypes are improved is not fully understood. The current study investigated the influence of FXR activity on the gut microbiota community structure and function and its impact on hepatic lipid metabolism. Predictions about the metabolic contribution of the gut microbiota to the host were made using 16S rRNA-based PICRUSt (phylogenetic investigation of communities by reconstruction of unobserved states), then validated using 1H nuclear magnetic resonance-based metabolomics, and results were summarized by using genome-scale metabolic models. Oral Gly-MCA administration altered the gut microbial community structure, notably reducing the ratio of Firmicutes to Bacteroidetes and its PICRUSt-predicted metabolic function, including reduced production of short-chain fatty acids (substrates for hepatic gluconeogenesis and de novo lipogenesis) in the ceca of HFD-fed mice. Metabolic improvement was intestinal FXR dependent, as revealed by the lack of changes in HFD-fed intestine-specific Fxr-null (FxrΔIE) mice treated with Gly-MCA. Integrative analyses based on genome-scale metabolic models demonstrated an important link between Lactobacillus and Clostridia bile salt hydrolase activity and bacterial fermentation. Hepatic metabolite levels after Gly-MCA treatment correlated with altered levels of gut bacterial species. In conclusion, modulation of the gut microbiota by inhibition of intestinal FXR signaling alters host liver lipid metabolism and improves obesity-related metabolic dysfunction. IMPORTANCE The farnesoid X receptor (FXR) plays an important role in mediating the dialog between the host

  1. Activation of farnesoid X receptor attenuates hepatic injury in a murine model of alcoholic liver disease

    SciTech Connect

    Wu, Weibin; Zhu, Bo; Peng, Xiaomin; Zhou, Meiling; Jia, Dongwei; Gu, Jianxin

    2014-01-03

    Highlights: •FXR activity was impaired by chronic ethanol ingestion in a murine model of ALD. •Activation of FXR attenuated alcohol-induced liver injury and steatosis. •Activation of FXR attenuated cholestasis and oxidative stress in mouse liver. -- Abstract: Alcoholic liver disease (ALD) is a common cause of advanced liver disease, and considered as a major risk factor of morbidity and mortality worldwide. Hepatic cholestasis is a pathophysiological feature observed in all stages of ALD. The farnesoid X receptor (FXR) is a member of the nuclear hormone receptor superfamily, and plays an essential role in the regulation of bile acid, lipid and glucose homeostasis. However, the role of FXR in the pathogenesis and progression of ALD remains largely unknown. Mice were fed Lieber-DeCarli ethanol diet or an isocaloric control diet. We used a specific agonist of FXR WAY-362450 to study the effect of pharmacological activation of FXR in alcoholic liver disease. In this study, we demonstrated that FXR activity was impaired by chronic ethanol ingestion in a murine model of ALD. Activation of FXR by specific agonist WAY-362450 protected mice from the development of ALD. We also found that WAY-362450 treatment rescued FXR activity, suppressed ethanol-induced Cyp2e1 up-regulation and attenuated oxidative stress in liver. Our results highlight a key role of FXR in the modulation of ALD development, and propose specific FXR agonists for the treatment of ALD patients.

  2. Activation of farnesoid X receptor induces RECK expression in mouse liver

    SciTech Connect

    Peng, Xiaomin; Wu, Weibin; Zhu, Bo; Sun, Zhichao; Ji, Lingling; Ruan, Yuanyuan; Zhou, Meiling; Zhou, Lei; Gu, Jianxin

    2014-01-03

    Highlights: •RECK is a novel transcriptional target gene of FXR in mouse liver. •The FXR response element is located within the intron 1 of RECK gene. •FXR agonist reverses the down-regulation of RECK in the liver in mouse NASH model. -- Abstract: Farnesoid X receptor (FXR) belongs to the ligand-activated nuclear receptor superfamily, and functions as a transcription factor regulating the transcription of numerous genes involved in bile acid homeostasis, lipoprotein and glucose metabolism. In the present study, we identified RECK, a membrane-anchored inhibitor of matrix metalloproteinases, as a novel target gene of FXR in mouse liver. We found that FXR agonist substantially augmented hepatic RECK mRNA and protein expression in vivo and in vitro. FXR regulated the transcription of RECK through directly binding to FXR response element located within intron 1 of the mouse RECK gene. Moreover, FXR agonist reversed the down-regulation of RECK in the livers from mice fed a methionine and choline deficient diet. In summary, our data suggest that RECK is a novel transcriptional target of FXR in mouse liver, and provide clues to better understanding the function of FXR in liver.

  3. Farnesoid X Receptor Deficiency Improves Glucose Homeostasis in Mouse Models of Obesity

    PubMed Central

    Prawitt, Janne; Abdelkarim, Mouaadh; Stroeve, Johanna H.M.; Popescu, Iuliana; Duez, Helene; Velagapudi, Vidya R.; Dumont, Julie; Bouchaert, Emmanuel; van Dijk, Theo H.; Lucas, Anthony; Dorchies, Emilie; Daoudi, Mehdi; Lestavel, Sophie; Gonzalez, Frank J.; Oresic, Matej; Cariou, Bertrand; Kuipers, Folkert; Caron, Sandrine; Staels, Bart

    2011-01-01

    OBJECTIVE Bile acids (BA) participate in the maintenance of metabolic homeostasis acting through different signaling pathways. The nuclear BA receptor farnesoid X receptor (FXR) regulates pathways in BA, lipid, glucose, and energy metabolism, which become dysregulated in obesity. However, the role of FXR in obesity and associated complications, such as dyslipidemia and insulin resistance, has not been directly assessed. RESEARCH DESIGN AND METHODS Here, we evaluate the consequences of FXR deficiency on body weight development, lipid metabolism, and insulin resistance in murine models of genetic and diet-induced obesity. RESULTS FXR deficiency attenuated body weight gain and reduced adipose tissue mass in both models. Surprisingly, glucose homeostasis improved as a result of an enhanced glucose clearance and adipose tissue insulin sensitivity. In contrast, hepatic insulin sensitivity did not change, and liver steatosis aggravated as a result of the repression of β-oxidation genes. In agreement, liver-specific FXR deficiency did not protect from diet-induced obesity and insulin resistance, indicating a role for nonhepatic FXR in the control of glucose homeostasis in obesity. Decreasing elevated plasma BA concentrations in obese FXR-deficient mice by administration of the BA sequestrant colesevelam improved glucose homeostasis in a FXR-dependent manner, indicating that the observed improvements by FXR deficiency are not a result of indirect effects of altered BA metabolism. CONCLUSIONS Overall, FXR deficiency in obesity beneficially affects body weight development and glucose homeostasis. PMID:21593203

  4. Induction of farnesoid X receptor signaling in germ-free mice colonized with a human microbiota.

    PubMed

    Wahlström, Annika; Kovatcheva-Datchary, Petia; Ståhlman, Marcus; Khan, Muhammad-Tanweer; Bäckhed, Fredrik; Marschall, Hanns-Ulrich

    2017-02-01

    The gut microbiota influences the development and progression of metabolic diseases partly by metabolism of bile acids (BAs) and modified signaling through the farnesoid X receptor (FXR). In this study, we aimed to determine how the human gut microbiota metabolizes murine BAs and affects FXR signaling in colonized mice. We colonized germ-free mice with cecal content from a mouse donor or feces from a human donor and euthanized the mice after short-term (2 weeks) or long-term (15 weeks) colonization. We analyzed the gut microbiota and BA composition and expression of FXR target genes in ileum and liver. We found that cecal microbiota composition differed between mice colonized with mouse and human microbiota and was stable over time. Human and mouse microbiota reduced total BA levels similarly, but the humanized mice produced less secondary BAs. The human microbiota was able to reduce the levels of tauro-β-muricholic acid and induce expression of FXR target genes Fgf15 and Shp in ileum after long-term colonization. We show that a human microbiota can change BA composition and induce FXR signaling in colonized mice, but the levels of secondary BAs produced are lower than in mice colonized with a mouse microbiota.

  5. Intestine-selective farnesoid X receptor inhibition improves obesity-related metabolic dysfunction

    PubMed Central

    Jiang, Changtao; Xie, Cen; Lv, Ying; Li, Jing; Krausz, Kristopher W.; Shi, Jingmin; Brocker, Chad N.; Desai, Dhimant; Amin, Shantu G.; Bisson, William H.; Liu, Yulan; Gavrilova, Oksana; Patterson, Andrew D.; Gonzalez, Frank J.

    2015-01-01

    The farnesoid X receptor (FXR) regulates bile acid, lipid and glucose metabolism. Here we show that treatment of mice with glycine-β-muricholic acid (Gly-MCA) inhibits FXR signalling exclusively in intestine, and improves metabolic parameters in mouse models of obesity. Gly-MCA is a selective high-affinity FXR inhibitor that can be administered orally and prevents, or reverses, high-fat diet-induced and genetic obesity, insulin resistance and hepatic steatosis in mice. The high-affinity FXR agonist GW4064 blocks Gly-MCA action in the gut, and intestine-specific Fxr-null mice are unresponsive to the beneficial effects of Gly-MCA. Mechanistically, the metabolic improvements with Gly-MCA depend on reduced biosynthesis of intestinal-derived ceramides, which directly compromise beige fat thermogenic function. Consequently, ceramide treatment reverses the action of Gly-MCA in high-fat diet-induced obese mice. We further show that FXR signalling in ileum biopsies of humans positively correlates with body mass index. These data suggest that Gly-MCA may be a candidate for the treatment of metabolic disorders. PMID:26670557

  6. Farnesoid X Receptor Protects against Kidney Injury in Uninephrectomized Obese Mice.

    PubMed

    Gai, Zhibo; Gui, Ting; Hiller, Christian; Kullak-Ublick, Gerd A

    2016-01-29

    Activation of the farnesoid X receptor (FXR) has indicated a therapeutic potential for this nuclear bile acid receptor in the prevention of diabetic nephropathy and obesity-induced renal damage. Here, we investigated the protective role of FXR against kidney damage induced by obesity in mice that had undergone uninephrectomy, a model resembling the clinical situation of kidney donation by obese individuals. Mice fed a high-fat diet developed the core features of metabolic syndrome, with subsequent renal lipid accumulation and renal injury, including glomerulosclerosis, interstitial fibrosis, and albuminuria. The effects were accentuated by uninephrectomy. In human renal biopsies, staining of 4-hydroxynonenal (4-HNE), glucose-regulated protein 78 (Grp78), and C/EBP-homologous protein, markers of endoplasmic reticulum stress, was more prominent in the proximal tubules of 15 obese patients compared with 16 non-obese patients. In mice treated with the FXR agonist obeticholic acid, renal injury, renal lipid accumulation, apoptosis, and changes in lipid peroxidation were attenuated. Moreover, disturbed mitochondrial function was ameliorated and the mitochondrial respiratory chain recovered following obeticholic acid treatment. Culturing renal proximal tubular cells with free fatty acid and FXR agonists showed that FXR activation protected cells from free fatty acid-induced oxidative stress and endoplasmic reticulum stress, as denoted by a reduction in the level of reactive oxygen species staining and Grp78 immunostaining, respectively. Several genes involved in glutathione metabolism were induced by FXR activation in the remnant kidney, which was consistent with a decreased glutathione disulfide/glutathione ratio. In summary, FXR activation maintains endogenous glutathione homeostasis and protects the kidney in uninephrectomized mice from obesity-induced injury.

  7. Small heterodimer partner overexpression partially protects against liver tumor development in farnesoid X receptor knockout mice

    SciTech Connect

    Li, Guodong; Kong, Bo; Zhu, Yan; Zhan, Le; Williams, Jessica A.; Tawfik, Ossama; Kassel, Karen M.; Luyendyk, James P.; Wang, Li; Guo, Grace L.

    2013-10-15

    Farnesoid X receptor (FXR, Nr1h4) and small heterodimer partner (SHP, Nr0b2) are nuclear receptors that are critical to liver homeostasis. Induction of SHP serves as a major mechanism of FXR in suppressing gene expression. Both FXR{sup −/−} and SHP{sup −/−} mice develop spontaneous hepatocellular carcinoma (HCC). SHP is one of the most strongly induced genes by FXR in the liver and is a tumor suppressor, therefore, we hypothesized that deficiency of SHP contributes to HCC development in the livers of FXR{sup −/−} mice and therefore, increased SHP expression in FXR{sup −/−} mice reduces liver tumorigenesis. To test this hypothesis, we generated FXR{sup −/−} mice with overexpression of SHP in hepatocytes (FXR{sup −/−}/SHP{sup Tg}) and determined the contribution of SHP in HCC development in FXR{sup −/−} mice. Hepatocyte-specific SHP overexpression did not affect liver tumor incidence or size in FXR{sup −/−} mice. However, SHP overexpression led to a lower grade of dysplasia, reduced indicator cell proliferation and increased apoptosis. All tumor-bearing mice had increased serum bile acid levels and IL-6 levels, which was associated with activation of hepatic STAT3. In conclusion, SHP partially protects FXR{sup −/−} mice from HCC formation by reducing tumor malignancy. However, disrupted bile acid homeostasis by FXR deficiency leads to inflammation and injury, which ultimately results in uncontrolled cell proliferation and tumorigenesis in the liver. - Highlights: • SHP does not prevent HCC incidence nor size in FXR KO mice but reduces malignancy. • Increased SHP promotes apoptosis. • Bile acids and inflammation maybe critical for HCC formation with FXR deficiency.

  8. Effects of coumestrol on lipid and glucose metabolism as a farnesoid X receptor ligand

    SciTech Connect

    Takahashi, Miki; Kanayama, Tomohiko; Yashiro, Takuya; Kondo, Hidehiko; Murase, Takatoshi; Hase, Tadashi; Tokimitsu, Ichiro; Nishikawa, Jun-ichi; Sato, Ryuichiro

    2008-08-01

    In the course of an effort to identify novel agonists of the farnesoid X receptor (FXR), coumestrol was determined to be one such ligand. Reporter and in vitro coactivator interaction assays revealed that coumestrol bound and activated FXR. Treatment of Hep G2 cells with coumestrol stimulated the expression of FXR target genes, thereby regulating the expression of target genes of the liver X receptor and hepatocyte nuclear factor-4{alpha}. Through these actions, coumestrol is expected to exert beneficial effects on lipid and glucose metabolism.

  9. Synthesis and biological evaluations of chalcones, flavones and chromenes as farnesoid x receptor (FXR) antagonists.

    PubMed

    Zhang, Guoning; Liu, Shuainan; Tan, Wenjuan; Verma, Ruchi; Chen, Yuan; Sun, Deyang; Huan, Yi; Jiang, Qian; Wang, Xing; Wang, Na; Xu, Yang; Wong, Chiwai; Shen, Zhufang; Deng, Ruitang; Liu, Jinsong; Zhang, Yanqiao; Fang, Weishuo

    2017-03-31

    Farnesoid X receptor (FXR), a nuclear receptor mainly distributed in liver and intestine, has been regarded as a potential target for the treatment of various metabolic diseases, cancer and infectious diseases related to liver. Starting from two previously identified chalcone-based FXR antagonists, we tried to increase the activity through the design and synthesis of a library containing chalcones, flavones and chromenes, based on substitution manipulation and conformation (ring closure) restriction strategy. Many chalcones and four chromenes were identified as microM potent FXR antagonists, among which chromene 11c significantly decreased the plasma and hepatic triglyceride level in KKay mice.

  10. SUMOylation of the Farnesoid X Receptor (FXR) Regulates the Expression of FXR Target Genes*

    PubMed Central

    Balasubramaniyan, Natarajan; Luo, Yuhuan; Sun, An-Qiang; Suchy, Frederick J.

    2013-01-01

    The farnesoid X receptor (FXR) belongs to a family of ligand-activated transcription factors that regulate many aspects of metabolism including bile acid homeostasis. Here we show that FXR is covalently modified by the small ubiquitin-like modifier (Sumo1), an important regulator of cell signaling and transcription. Well conserved consensus sites at lysine 122 and 275 in the AF-1 and ligand binding domains, respectively, of FXR were subject to SUMOylation in vitro and in vivo. Chromatin immunoprecipitation (ChIP) analysis showed that Sumo1 was recruited to the bile salt export pump (BSEP), the small heterodimer partner (SHP), and the OSTα-OSTβ organic solute transporter loci in a ligand-dependent fashion. Sequential chromatin immunoprecipitation (ChIP-ReChIP) verified the concurrent binding of FXR and Sumo1 to the BSEP and SHP promoters. Overexpression of Sumo1 markedly decreased binding and/or recruitment of FXR to the BSEP and SHP promoters on ChIP-ReChIP. SUMOylation did not have an apparent effect on nuclear localization of FXR. Expression of Sumo1 markedly inhibited the ligand-dependent, transactivation of BSEP and SHP promoters by FXR/retinoid X receptor α (RXRα) in HepG2 cells. In contrast, mutations that abolished SUMOylation of FXR or siRNA knockdown of Sumo1 expression augmented the transactivation of BSEP and SHP promoters by FXR. Pathways for SUMOylation were significantly altered during obstructive cholestasis with differential Sumo1 recruitment to the promoters of FXR target genes. In conclusion, FXR is subject to SUMOylation that regulates its capacity to transactivate its target genes in normal liver and during obstructive cholestasis. PMID:23546875

  11. Farnesoid X Receptor Activation Attenuates Intestinal Ischemia Reperfusion Injury in Rats

    PubMed Central

    Decuypere, Jean-Paul; Farré, Ricard; De Hertogh, Gert; Lenaerts, Kaatje; Jochmans, Ina; Monbaliu, Diethard; Nevens, Frederik; Tack, Jan; Laleman, Wim; Pirenne, Jacques

    2017-01-01

    Introduction The farnesoid X receptor (FXR) is abundantly expressed in the ileum, where it exerts an enteroprotective role as a key regulator of intestinal innate immunity and homeostasis, as shown in pre-clinical models of inflammatory bowel disease. Since intestinal ischemia reperfusion injury (IRI) is characterized by hyperpermeability, bacterial translocation and inflammation, we aimed to investigate, for the first time, if the FXR-agonist obeticholic acid (OCA) could attenuate intestinal ischemia reperfusion injury. Material and Methods In a validated rat model of intestinal IRI (laparotomy + temporary mesenteric artery clamping), 3 conditions were tested (n = 16/group): laparotomy only (sham group); ischemia 60min+ reperfusion 60min + vehicle pretreatment (IR group); ischemia 60min + reperfusion 60min + OCA pretreatment (IR+OCA group). Vehicle or OCA (INT-747, 2*30mg/kg) was administered by gavage 24h and 4h prior to IRI. The following end-points were analyzed: 7-day survival; biomarkers of enterocyte viability (L-lactate, I-FABP); histology (morphologic injury to villi/crypts and villus length); intestinal permeability (Ussing chamber); endotoxin translocation (Lipopolysaccharide assay); cytokines (IL-6, IL-1-β, TNFα, IFN-γ IL-10, IL-13); apoptosis (cleaved caspase-3); and autophagy (LC3, p62). Results It was found that intestinal IRI was associated with high mortality (90%); loss of intestinal integrity (structurally and functionally); increased endotoxin translocation and pro-inflammatory cytokine production; and inhibition of autophagy. Conversely, OCA-pretreatment improved 7-day survival up to 50% which was associated with prevention of epithelial injury, preserved intestinal architecture and permeability. Additionally, FXR-agonism led to decreased pro-inflammatory cytokine release and alleviated autophagy inhibition. Conclusion Pretreatment with OCA, an FXR-agonist, improves survival in a rodent model of intestinal IRI, preserves the gut barrier

  12. Identification of Trisubstituted-pyrazol Carboxamide Analogs as Novel and Potent Antagonists of Farnesoid X Receptor

    PubMed Central

    Yu, Donna D.; Lin, Wenwei; Forman, Barry M.; Chen, Taosheng

    2014-01-01

    Farnesoid X receptor (FXR, NRIH4) plays a major role in the control of cholesterol metabolism. This suggests that antagonizing the transcriptional activity of FXR is a potential means to treat cholestasis and related metabolic disorders. Here we describe the synthesis, biological evaluation, and structure-activity relationship (SAR) studies of trisubstituted-pyrazol carboxamides as novel and potent FXR antagonists. One of these novel FXR antagonists, 4j has an IC50 of 7.5 nM in an FXR binding assay and 468.5 nM in a cell-based FXR antagonistic assay. Compound 4j has no detectable FXR agonistic activity or cytotoxicity. Notably, 4j is the most potent FXR antagonist identified to date; it has a promising in vitro profile and could serve as an excellent chemical tool to elucidate the biological function of FXR. PMID:24775917

  13. Farnesoid X receptor immunolocalization in reproductive tissues of adult female rabbits.

    PubMed

    Anaya-Hernández, Arely; Méndez-Tepepa, Maribel; Hernández-Aragón, Laura G; Pacheco, Pablo; Martínez-Gómez, Margarita; Castelán, Francisco; Cuevas, Estela

    2014-07-01

    Farnesoid X receptor (FXR) has been involved in lipid metabolism, cell proliferation, apoptosis, and aromatase expression, as well as in the steroid synthesis and signaling. Considering that these events occur in reproductive tissues in females, the aim of the present study was to determine the immunolocalization of FXR in the ovary, oviduct, uterus, and vagina of rabbits. Rabbits were sacrificed and their reproductive tissues were excised and histologically processed. Immunohistochemistry for FXR was done and reproductive tissues were photographed. FXR immunoreactivity was found in all types of ovarian follicles, ovarian stroma, and corpus luteum of virgin and pregnant rabbits. Also, oviductal and vaginal epithelium of virgins, as well as the oviductal smooth muscle, showed anti-FXR immunoreactivity. The uterine epithelium and musculature of virgins had scarce anti-FXR immunoreactivity. Although the role of FXR in female reproductive tissues is still not known, it is possible to consider various functions related to the reproductive tissue.

  14. A formulation-enabled preclinical efficacy assessment of a farnesoid X receptor agonist, GW4064, in hamsters and cynomolgus monkeys.

    PubMed

    Chiang, Po-Chang; Thompson, David C; Ghosh, Sarbani; Heitmeier, Monique R

    2011-11-01

    The farnesoid X receptor (FXR) belongs to one of the human nuclear receptor superfamilies that regulate gene transcription. FXR is widely expressed in liver, gall bladder, intestine, kidney, and adrenal glands. It serves as a key controller of bile acid homeostasis through its regulation of bile acid synthesis, conjugation, secretion, and absorption. FXR is also known to play a role in lipid regulation, triglyceride synthesis, and lipoprotein metabolism and clearance. We used a commercially available FXR agonist, GW4064, as a model compound to assess preclinical efficacy in two species (hamster and cynomolgus monkey). The crystalline GW4064, however, was found to have limited solubility, which resulted in poor oral bioavailability. This made it difficult to assess in vivo efficacy at the exposure levels desired. The physiochemical properties of GW4064 were assessed and both salt and self-emulsifying drug delivery system (SEDDS) formulation were developed and tested. The SEDDS formulation was found to greatly improve the oral bioavailability of GW4064, and permitted the evaluation of FXR agonist target efficacy.

  15. The farnesoid-X-receptor in myeloid cells controls CNS autoimmunity in an IL-10-dependent fashion.

    PubMed

    Hucke, Stephanie; Herold, Martin; Liebmann, Marie; Freise, Nicole; Lindner, Maren; Fleck, Ann-Katrin; Zenker, Stefanie; Thiebes, Stephanie; Fernandez-Orth, Juncal; Buck, Dorothea; Luessi, Felix; Meuth, Sven G; Zipp, Frauke; Hemmer, Bernhard; Engel, Daniel Robert; Roth, Johannes; Kuhlmann, Tanja; Wiendl, Heinz; Klotz, Luisa

    2016-09-01

    Innate immune responses by myeloid cells decisively contribute to perpetuation of central nervous system (CNS) autoimmunity and their pharmacologic modulation represents a promising strategy to prevent disease progression in Multiple Sclerosis (MS). Based on our observation that peripheral immune cells from relapsing-remitting and primary progressive MS patients exhibited strongly decreased levels of the bile acid receptor FXR (farnesoid-X-receptor, NR1H4), we evaluated its potential relevance as therapeutic target for control of established CNS autoimmunity. Pharmacological FXR activation promoted generation of anti-inflammatory macrophages characterized by arginase-1, increased IL-10 production, and suppression of T cell responses. In mice, FXR activation ameliorated CNS autoimmunity in an IL-10-dependent fashion and even suppressed advanced clinical disease upon therapeutic administration. In analogy to rodents, pharmacological FXR activation in human monocytes from healthy controls and MS patients induced an anti-inflammatory phenotype with suppressive properties including control of effector T cell proliferation. We therefore, propose an important role of FXR in control of T cell-mediated autoimmunity by promoting anti-inflammatory macrophage responses.

  16. Farnesoid X receptor activation improves erectile dysfunction in models of metabolic syndrome and diabetes.

    PubMed

    Morelli, Annamaria; Vignozzi, Linda; Maggi, Mario; Adorini, Luciano

    2011-08-01

    The metabolic syndrome (MetS) is an insulin-resistant state characterized by a cluster of cardiovascular risk factors, including abdominal obesity, hyperglycemia, elevated blood pressure and combined dyslipidemia. In this review, we discuss the potential of farnesoid X receptor (FXR) agonists in the treatment of erectile dysfunction (ED), a multifactorial disorder often comorbid with MetS. FXR not only regulates lipid and glucose homeostasis but also influences endothelial function and atherosclerosis, suggesting a regulatory role for this hormone nuclear receptor in the cardiovascular complications associated with the MetS, including ED. MetS induces ED via several mechanisms, and in particular through endothelial dysfunction in penile vessels. In a high-fat diet rabbit model of MetS, a 3-month treatment with the potent and selective FXR agonist INT-747 restores endothelium-dependent relaxation in isolated cavernous tissue, normalizing responsiveness to acetylcholine and to electrical field stimulation. Accordingly, eNOS expression in the penis is greatly up-regulated by INT-747 treatment. Experiments in a rat model of chemically-induced type 1 diabetes further demonstrate that INT-747 treatment preserves erectile function induced by electrical stimulation of the cavernous nerve. These results add a new facet to the pleiotropic activities mediated by FXR, and reveal novel beneficial effects of FXR activation with potential clinical relevance. This article is part of a Special Issue entitled: Translating nuclear receptors from health to disease.

  17. Farnesoid X receptor directly regulates xenobiotic detoxification genes in the long-lived Little mice.

    PubMed

    Jiang, Yanjun; Jin, Jingling; Iakova, Polina; Hernandez, Julio Cesar; Jawanmardi, Nicole; Sullivan, Emily; Guo, Grace L; Timchenko, Nikolai A; Darlington, Gretchen J

    2013-09-01

    Activation of xenobiotic metabolism pathways has been linked to lifespan extension in different models of aging. However, the mechanisms underlying activation of xenobiotic genes remain largely unknown. Here we showed that although farnesoid X receptor (FXR, Nr1h4) mRNA levels do not change significantly, FXR protein levels are elevated in the livers of the long-lived Little mice, leading to increased DNA binding activity of FXR. Hepatic FXR expression is sex-dependent in wild-type mice but not in Little mice, implying that up-regulation of FXR might be dependent on the reduction of growth hormone in Little mice. Growth hormone treatment decreased hepatic expression of FXR and xenobiotic genes Abcb1a, Fmo3 and Gsta2 in both wild-type and Little mice, suggesting an association between FXR and xenobiotic gene expression. We found that Abcb1a is transactivated by FXR via direct binding of FXR/retinoid X receptor α (RXRα) heterodimer to a response element at the proximal promoter. FXR also positively controls Fmo3 and Gsta2 expression through direct interaction with the response elements in these genes. Our study demonstrates that xenobiotic genes are direct transcriptional targets of FXR and suggests that FXR signaling may play a critical role in the lifespan extension observed in Little mice.

  18. Glutamine attenuates obstructive cholestasis in rats via farnesoid X receptor-mediated regulation of Bsep and Mrp2.

    PubMed

    Liu, Bingli; Li, Yiming; Ji, Hong; Lu, Hongwei; Li, Hua; Shi, Yakun

    2017-02-01

    To investigate the protective effect of glutamine (Gln) against obstructive cholestasis in association with farnesoid X receptor (FXR) activation, an obstructive cholestasis model was established in male Sprague-Dawley rats by bile duct ligation (BDL). Serum biomarkers and hematoxylin plus eosin staining were used to identify the degree of hepatic injury in the rats with obstructive cholestasis after Gln treatment. Immunohistochemistry, real-time PCR, Western blot, cultured primary rat hepatocytes with FXR knockdown, and dual-luciferase reporter assay were performed to elucidate the mechanisms underlying Gln hepatoprotection. We found that Gln treatment protected against obstructive cholestasis induced by BDL through reducing hepatocyte injury. Upregulation of the hepatic efflux transporters small heterodimer partner (Shp), bile salt export pump (Bsep), and multidrug resistance-associated protein 2 (Mrp2), and inhibition of the hepatic uptake transporter Na(+)/taurocholate cotransporting polypeptide (Ntcp) and the bile acid synthesis enzyme cholesterol 7α-hydroxylase (Cyp7a1) expression were observed in rats with BDL treated with Gln in vivo. Furthermore, the regulatory effect of Gln on Bsep and Mrp2 expression was abrogated after FXR knockdown in rat primary cultured hepatocytes. Luciferase assay HepG2 cells also illustrated FXR was a direct target for Gln treatment. In conclusion, the regulation of Bsep and Mrp2 expression mediated by FXR might be an important mechanism for Gln against obstructive cholestasis.

  19. Role of farnesoid X receptor in establishment of ontogeny of phase-I drug metabolizing enzyme genes in mouse liver.

    PubMed

    Peng, Lai; Piekos, Stephanie; Guo, Grace L; Zhong, Xiao-Bo

    2016-09-01

    The expression of phase-I drug metabolizing enzymes in liver changes dramatically during postnatal liver maturation. Farnesoid X receptor (FXR) is critical for bile acid and lipid homeostasis in liver. However, the role of FXR in regulating ontogeny of phase-I drug metabolizing genes is not clear. Hence, we applied RNA-sequencing to quantify the developmental expression of phase-I genes in both Fxr-null and control (C57BL/6) mouse livers during development. Liver samples of male C57BL/6 and Fxr-null mice at 6 different ages from prenatal to adult were used. The Fxr-null showed an overall effect to diminish the "day-1 surge" of phase-I gene expression, including cytochrome P450s at neonatal ages. Among the 185 phase-I genes from 12 different families, 136 were expressed, and differential expression during development occurred in genes from all 12 phase-I families, including hydrolysis: carboxylesterase (Ces), paraoxonase (Pon), and epoxide hydrolase (Ephx); reduction: aldoketo reductase (Akr), quinone oxidoreductase (Nqo), and dihydropyrimidine dehydrogenase (Dpyd); and oxidation: alcohol dehydrogenase (Adh), aldehyde dehydrogenase (Aldh), flavin monooxygenases (Fmo), molybdenum hydroxylase (Aox and Xdh), cytochrome P450 (P450), and cytochrome P450 oxidoreductase (Por). The data also suggested new phase-I genes potentially targeted by FXR. These results revealed an important role of FXR in regulation of ontogeny of phase-I genes.

  20. Bile Salt Export Pump is Dysregulated with Altered Farnesoid X Receptor Isoform Expression in Patients with Hepatocellular Carcinoma

    PubMed Central

    Chen, Yuan; Song, Xiulong; Valanejad, Leila; Vasilenko, Alexander; More, Vijay; Qiu, Xi; Chen, Weikang; Lai, Yurong; Slitt, Angela; Stoner, Matthew; Yan, Bingfang; Deng, Ruitang

    2012-01-01

    As a canalicular bile acid effluxer, bile salt export pump (BSEP) plays a vital role in maintaining bile acid homeostasis. BSEP deficiency leads to severe cholestasis and hepatocellular carcinoma (HCC) in young children. Regardless of the etiology, chronic inflammation is the common pathological process for HCC development. Clinical studies showed that bile acid homeostasis is disrupted in HCC patients with elevated serum bile acid level as a proposed marker for HCC. However, the underlying mechanisms remain largely unknown. In this study, we found that BSEP expression was severely diminished in HCC tissues and markedly reduced in adjacent non-tumor tissues. In contrast to mouse, human BSEP was regulated by farnesoid x receptor (FXR) in an isoform-dependent manner. FXRα2 exhibited a much more potent activity than FXRα1 in transactivating human BSEP in vitro and in vivo. The decreased BSEP expression in HCC was associated with altered relative expression of FXRα1 and FXRα2. The FXRα1/FXRα2 ratios were significantly increased with undetectable FXRα2 expression in one third of the HCC tumor samples. Similar correlation between BSEP and FXR isoform expression was confirmed in hepatoma Huh 7 and HepG2 cells. Further studies showed that intrahepatic proinflammatory cytokines interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were significantly elevated in HCC tissues. Treatment of Huh 7 cells with IL-6 and TNF-α resulted in a marked increase in the FXRα1/FXRα2 ratio concurrent with a significant decrease in BSEP expression. In conclusion, BSEP expression was severely diminished in HCC patients associated with alteration of FXR isoform expression induced by inflammation, and the restoration of BSEP expression through suppressing inflammation in the liver may re-establish the bile acid homeostasis. PMID:23213087

  1. The Halicylindramides, Farnesoid X Receptor Antagonizing Depsipeptides from a Petrosia sp. Marine Sponge Collected in Korea.

    PubMed

    Hahn, Dongyup; Kim, Hiyoung; Yang, Inho; Chin, Jungwook; Hwang, Hoosang; Won, Dong Hwan; Lee, Byoungchan; Nam, Sang-Jip; Ekins, Merrick; Choi, Hyukjae; Kang, Heonjoong

    2016-03-25

    Three new structurally related depsipeptides, halicylindramides F-H (1-3), and two known halicylindramides were isolated from a Petrosia sp. marine sponge collected off the shore of Youngdeok-Gun, East Sea, Republic of Korea. Their planar structures were elucidated by extensive spectroscopic data analyses including 1D and 2D NMR data as well as MS data. The absolute configurations of halicylindramides F-H (1-3) were determined by Marfey's method in combination with Edman degradation. The absolute configurations at C-4 of the dioxyindolyl alanine (Dioia) residues of halicylindramides G (2) and H (3) were determined as 4S and 4R, respectively, based on ECD spectroscopy. The C-2 configurations of Dioia in 2 and 3 were speculated to both be 2R based on the shared biogenesis of the halicylindramides. Halicylindramides F (1), A (4), and C (5) showed human farnesoid X receptor (hFXR) antagonistic activities, but did not bind directly to hFXR.

  2. Reciprocal regulation of farnesoid X receptor α activity and hepatitis B virus replication in differentiated HepaRG cells and primary human hepatocytes.

    PubMed

    Radreau, Pauline; Porcherot, Marine; Ramière, Christophe; Mouzannar, Karim; Lotteau, Vincent; André, Patrice

    2016-09-01

    Hepatitis B virus (HBV) and bile salt metabolism seem tightly connected. HBV enters hepatocytes by binding to sodium taurocholate cotransporting polypeptide (NTCP), the genome of which contains 2 active farnesoid X receptor (FXR) α response elements that participate in HBV transcriptional activity. We investigated in differentiated HepaRG cells and in primary human hepatocytes (PHHs) effects of FXR activation on HBV replication and of infection on the FXR pathway. In HepaRG cells, FXR agonists (6-ethyl chenodeoxycholic acid and GW4064), but no antagonist, and an FXR-unrelated bile salt inhibited viral mRNA, DNA, and protein production (IC50, 0.1-0.5 μM) and reduced covalently closed circular DNA pool size. These effects were independent of the NTCP inhibitor cyclosporine-A, which suggests inhibition occurred at a postentry step. Similar results were obtained in PHHs with GW4064. Infection of these cells increased expression of FXR and modified expression of FXR-regulated genes SHP, APOA1, NTCP, CYP7A1, and CYP8B1 with a more pronounced effect in PHHs than in HepaRG cells. FXR agonists reversed all but one of the HBV-induced FXR gene profile modifications. HBV replication and FXR regulation seem to be interdependent, and altered bile salt metabolism homeostasis might contribute to the persistence of HBV infection.-Radreau, P., Porcherot, M., Ramière, C., Mouzannar, K., Lotteau, V., André, P. Reciprocal regulation of farnesoid X receptor α activity and hepatitis B virus replication in differentiated HepaRG cells and primary human hepatocytes.

  3. Farnesoid X receptor-dependent and -independent pathways mediate the transcriptional control of human fibroblast growth factor 19 by vitamin A.

    PubMed

    Jahn, Daniel; Sutor, Dominic; Dorbath, Donata; Weiß, Johannes; Götze, Oliver; Schmitt, Johannes; Hermanns, Heike M; Geier, Andreas

    2016-02-01

    Fibroblast growth factor 19 (FGF19) is a gut-derived hormone that controls bile acid (BA), carbohydrate and lipid metabolism. Whereas strong evidence supports a key role of BAs and farnesoid X receptor (FXR) for the control of FGF19 expression, information on other regulators is limited. In mice, FGF15 expression (ortholog of human FGF19) is induced by vitamin A (VitA) in an FXR-dependent manner. However, the significance of this finding for human FGF19 is currently unclear. Here, we demonstrate that VitA derivatives induce FGF19 in human intestinal cell lines by a direct transcriptional mechanism. In contrast to mouse FGF15, however, this direct regulation is not dependent on FXR but mediated by retinoic acid receptors (RARs) and their interaction with a novel DR-5 element in the human FGF19 gene. In addition to this direct effect, VitA derivatives impacted on the BA-mediated control of FGF19 by regulation of FXR protein levels. In conclusion, VitA regulates human FGF19 expression through FXR-dependent and -independent pathways. Moreover, we suggest that considerable mechanistic differences exist between humans and mice with regard to the nuclear receptors controlling the VitA-FGF15/19 axis. These findings may implicate a clinical relevance of RAR-activating VitA derivatives for the regulation of FGF19 levels in humans.

  4. Xanthohumol, the chalcone from beer hops (Humulus lupulus L.), is the ligand for farnesoid X receptor and ameliorates lipid and glucose metabolism in KK-A(y) mice.

    PubMed

    Nozawa, Hajime

    2005-10-28

    We have examined the modulating action of xanthohumol (XN) on the farnesoid X receptor (FXR) in vitro and in vivo. In the transient transfection assay, XN dose-dependently increased the BSEP promoter-driven luciferase activity. XN-fed KK-A(y) mice exhibited lowered levels of plasma glucose, plasma, and hepatic triglyceride. They also showed decreased amounts of water intake, lowered weights of white adipose tissue, and exhibited increased levels of plasma adiponectin, indicating that XN attenuated diabetes in KK-A(y) mice. The hepatic gene expression of XN-fed mice showed lowered levels of SREBP-1c including its targets involved in fatty acid synthesis and lowered levels of gluconeogenetic genes. However, the expression of cholesterol 7-hydroxylase (CYP7A1) was significantly induced in the liver of XN-fed mice. From the present results, it is suggested that XN acts on FXR through a selective bile acid receptor modulator (SBARM) like guggulsterone or polyunsaturated fatty acids, which have previously been reported as SBARMs.

  5. The evolution of farnesoid X, vitamin D, and pregnane X receptors: insights from the green-spotted pufferfish (Tetraodon nigriviridis) and other non-mammalian species

    PubMed Central

    2011-01-01

    Background The farnesoid X receptor (FXR), pregnane X receptor (PXR), and vitamin D receptor (VDR) are three closely related nuclear hormone receptors in the NR1H and 1I subfamilies that share the property of being activated by bile salts. Bile salts vary significantly in structure across vertebrate species, suggesting that receptors binding these molecules may show adaptive evolutionary changes in response. We have previously shown that FXRs from the sea lamprey (Petromyzon marinus) and zebrafish (Danio rerio) are activated by planar bile alcohols found in these two species. In this report, we characterize FXR, PXR, and VDR from the green-spotted pufferfish (Tetraodon nigriviridis), an actinopterygian fish that unlike the zebrafish has a bile salt profile similar to humans. We utilize homology modelling, docking, and pharmacophore studies to understand the structural features of the Tetraodon receptors. Results Tetraodon FXR has a ligand selectivity profile very similar to human FXR, with strong activation by the synthetic ligand GW4064 and by the primary bile acid chenodeoxycholic acid. Homology modelling and docking studies suggest a ligand-binding pocket architecture more similar to human and rat FXRs than to lamprey or zebrafish FXRs. Tetraodon PXR was activated by a variety of bile acids and steroids, although not by the larger synthetic ligands that activate human PXR such as rifampicin. Homology modelling predicts a larger ligand-binding cavity than zebrafish PXR. We also demonstrate that VDRs from the pufferfish and Japanese medaka were activated by small secondary bile acids such as lithocholic acid, whereas the African clawed frog VDR was not. Conclusions Our studies provide further evidence of the relationship between both FXR, PXR, and VDR ligand selectivity and cross-species variation in bile salt profiles. Zebrafish and green-spotted pufferfish provide a clear contrast in having markedly different primary bile salt profiles (planar bile alcohols for

  6. Overexpression of farnesoid X receptor in small airways contributes to epithelial to mesenchymal transition and COX-2 expression in chronic obstructive pulmonary disease

    PubMed Central

    Chen, Bi; You, Wen-Jie; Xue, Shan; Qin, Hui; Zhao, Xu-Ji; Zhang, Miao; Liu, Xue-Qing; Zhu, Shu-Yang

    2016-01-01

    Background Epithelial-mesenchymal transition (EMT) and cyclooxygenase-2 (COX-2) contribute to airway remodelling and inflammation in chronic obstructive pulmonary disease (COPD). Recent data suggest that the farnesoid X receptor (FXR), a nuclear receptor traditionally considered as bile acid-activated receptor, is also expressed in non-classical bile acids target tissues with novel functions beyond regulating bile acid homeostasis. This study aimed to investigate the potential role of FXR in the development of COPD, as well as factors that affect FXR expression. Methods Expression of FXR, EMT biomarkers and COX-2 was examined by immunohistochemistry in lung tissues from non-smokers, smokers, and smokers with COPD. The role of FXR in TGF-β1-induced EMT and COX-2 expression in human bronchial epithelial (HBE) cells was evaluated in vitro. Factors regulating FXR expression were assessed in cultured HBE cells and a cigarette smoke-induced rat model of COPD. Results Expression of FXR, EMT markers and COX-2 was significantly elevated in small airway epithelium of COPD patients compared with controls. The staining scores of FXR in small airway epithelium were negatively related with FEV1% of predicted of smokers without and with COPD. FXR agonist GW4064 remarkably enhanced and FXR antagonist Z-Guggulsterone significantly inhibited EMT changes in TGF-β1-treated HBE cells. Both chenodeoxycholic acid (CDCA) and GW4064 increased COX-2 expression in HBE cells, whereas Z-Guggulsterone dramatically restrained CDCA-induced COX-2 expression. Finally, FXR expression is induced by IL-4 and IL-13 in HBE cells, as well as by cigarette smoke exposure in a rat model of COPD. Conclusions Overexpression of FXR in small airway may contribute to airway remodelling and inflammation in COPD by regulating EMT and COX-2 expression. PMID:28066584

  7. Dietary cholesterol stimulates CYP7A1 in rats because farnesoid X receptor is not activated.

    PubMed

    Xu, Guorong; Pan, Lu-Xing; Li, Hai; Shang, Quan; Honda, Akira; Shefer, Sarah; Bollineni, Jaya; Matsuzaki, Yasushi; Tint, G Stephen; Salen, Gerald

    2004-05-01

    Cholesterol feeding upregulates CYP7A1 in rats but downregulates CYP7A1 in rabbits. To clarify the mechanism responsible for the upregulation of CYP7A1 in cholesterol-fed rats, the effects of dietary cholesterol (Ch) and cholic acid (CA) on the activation of the nuclear receptors, liver X-receptor (LXR-alpha) and farsenoid X-receptor (FXR), which positively and negatively regulate CYP7A1, were investigated in rats. Studies were carried out in four groups (n = 12/group) of male Sprague-Dawley rats fed regular chow (control), 2% Ch, 2% Ch + 1% CA, and 1% CA alone for 1 wk. Changes in mRNA expression of short heterodimer partner (SHP) and bile salt export pump (BSEP), target genes for FXR, were determined to indicate FXR activation, whereas the expression of ABCA1 and lipoprotein lipase (LPL), target genes for LXR-alpha, reflected activation. CYP7A1 mRNA and activity increased twofold and 70%, respectively, in rats fed Ch alone when the bile acid pool size was stable but decreased 43 and 49%, respectively, after CA was added to the Ch diet, which expanded the bile acid pool 3.4-fold. SHP and BSEP mRNA levels did not change after feeding Ch but increased 88 and 37% in rats fed Ch + CA. This indicated that FXR was activated by the expanded bile acid pool. When Ch or Ch + CA were fed, hepatic concentrations of oxysterols, ligands for LXR-alpha increased to activate LXR-alpha, as evidenced by increased mRNA levels of ABCA1 and LPL. Feeding CA alone enlarged the bile acid pool threefold and increased the expression of both SHP and BSEP. These results suggest that LXR-alpha was activated in rats fed both Ch or Ch + CA, whereas CYP7A1 mRNA and activity were induced only in Ch-fed rats where the bile acid pool was not enlarged such that FXR was not activated. In rats fed Ch + CA, the bile acid pool expanded, which activated FXR to offset the stimulatory effects of LXR-alpha on CYP7A1.

  8. GW4064, an Agonist of Farnesoid X Receptor, Represses CYP3A4 Expression in Human Hepatocytes by Inducing Small Heterodimer Partner Expression

    PubMed Central

    Zhang, Shu; Pan, Xian

    2015-01-01

    Farnesoid X receptor (FXR) functions as a regulator of bile acid and lipid homeostasis and is recognized as a promising therapeutic target for metabolic diseases. The biologic function of FXR is mediated in part by a small heterodimer partner (SHP); ligand-activated FXR enhances SHP expression, and SHP in turn represses the activity of multiple transcription factors. This study aimed to investigate the effect of FXR activation on expression of the major drug-metabolizing enzyme CYP3A4. The effects of 3-(2,6-dichlorophenyl)-4-(3′-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylisoxazole (GW4064), a synthetic agonist of FXR, on the expression and activity of CYP3A4 were examined in primary human hepatocytes by using quantitative real-time polymerase chain reaction and S9 phenotyping. In human hepatocytes, treatment of GW4064 (1 μM) for 48 hours resulted in a 75% decrease in CYP3A4 mRNA expression and a 25% decrease in CYP3A4 activity, accompanied by ∼3-fold increase in SHP mRNA expression. In HepG2 cells, SHP repressed transactivation of CYP3A4 promoter by pregnane X receptor (PXR), constitutive androstane receptor (CAR), and glucocorticoid receptor. Interestingly, GW4064 did not repress expression of CYP2B6, another target gene of PXR and CAR; GW4064 enhanced CYP2B6 promoter activity. In conclusion, GW4064 represses CYP3A4 expression in human hepatocytes, potentially through upregulation of SHP expression and subsequent repression of CYP3A4 promoter activity. Clinically significant drug-drug interaction involving FXR agonists and CYP3A4 substrates may occur. PMID:25725071

  9. Boldine enhances bile production in rats via osmotic and Farnesoid X receptor dependent mechanisms

    SciTech Connect

    Cermanova, Jolana; Kadova, Zuzana; Zagorova, Marie; Hroch, Milos; Tomsik, Pavel; Nachtigal, Petr; Kudlackova, Zdenka; Pavek, Petr; Dubecka, Michaela; Ceckova, Martina; Staud, Frantisek; Laho, Tomas; Micuda, Stanislav

    2015-05-15

    Boldine, the major alkaloid from the Chilean Boldo tree, is used in traditional medicine to support bile production, but evidence to support this function is controversial. We analyzed the choleretic potential of boldine, including its molecular background. The acute- and long-term effects of boldine were evaluated in rats either during intravenous infusion or after 28-day oral treatment. Infusion of boldine instantly increased the bile flow 1.4-fold in healthy rats as well as in animals with Mrp2 deficiency or ethinylestradiol induced cholestasis. This effect was not associated with a corresponding increase in bile acid or glutathione biliary excretion, indicating that the effect is not related to stimulation of either bile acid dependent or independent mechanisms of bile formation and points to the osmotic activity of boldine itself. We subsequently analyzed bile production under conditions of changing biliary excretion of boldine after bolus intravenous administration and found strong correlations between both parameters. HPLC analysis showed that bile concentrations of boldine above 10 μM were required for induction of choleresis. Importantly, long-term pretreatment, when the bile collection study was performed 24-h after the last administration of boldine, also accelerated bile formation despite undetectable levels of the compound in bile. The effect paralleled upregulation of the Bsep transporter and increased biliary clearance of its substrates, bile acids. We consequently confirmed the ability of boldine to stimulate the Bsep transcriptional regulator, FXR receptor. In conclusion, our study clarified the mechanisms and circumstances surrounding the choleretic activity of boldine. - Highlights: • Boldine may increase bile production by direct as well as indirect mechanisms. • Biliary concentrations of boldine above 10 μM directly stimulate bile production. • Long-term oral boldine administration increases bile acid (BA) biliary secretion. • Boldine

  10. Hypothyroidism Induces a Moderate Steatohepatitis Accompanied by Liver Regeneration, Mast Cells Infiltration, and Changes in the Expression of the Farnesoid X Receptor.

    PubMed

    Rodríguez-Castelán, J; Corona-Pérez, A; Nicolás-Toledo, L; Martínez-Gómez, M; Castelán, F; Cuevas-Romero, E

    2017-03-01

    Hypothyroidism is associated with the development of non-alcoholic steatohepatitis, but cellular mechanisms have been scarcely analyzed. Thyroid hormones regulate the synthesis and secretion of bile acids that are endogenous ligands of the farnesoid receptor (FXRα), which have been involved in the development of non-alcoholic steatohepatitis. However, the relationship between thyroid hormones and FXRα expression in the liver is yet unknown. Control (n=6) and methimazole-induced hypothyroid (n=6) female rabbits were used to evaluate the amount of lipids and glycogen, vascularization, hepatocytes proliferation, immune cells infiltration, and expression of FXRα. Student-t or Mann-Whitney U tests were carried out to determine significant differences. Hypothyroidism induced steatosis, glycogen loss, fibrosis, and a minor vascularization in the liver. In contrast, hypothyroidism increased the proliferation of hepatocytes and the infiltration of mast cells, but did not modify the number of immune cells into sinusoids. These changes were associated with a minor anti-FXRα immunoreactivity of periportal hepatocytes and pericentral immune cells. Our results suggest that hypothyroidism induces a moderate non-alcoholic steatohepatitis, alllowing the hepatic regeneration. The FXRα may be involved in the development of non-alcoholic steatohepatitis in hypothyroid subjects.

  11. Farnesoid X receptor ligand CDCA suppresses human prostate cancer cells growth by inhibiting lipid metabolism via targeting sterol response element binding protein 1

    PubMed Central

    Liu, Nian; Zhao, Jun; Wang, Jinguo; Teng, Haolin; Fu, Yaowen; Yuan, Hang

    2016-01-01

    Aim: A wealth of studies have demonstrated that abnormal cellular lipid metabolism plays an important role in prostate cancer (PCa) development. Therefore, manipulating lipid metabolism is a potential PCa therapy strategy. In this study, our goal is to investigate the role of farnesoid X receptor (FXR) in regulating the proliferation and lipid metabolism of human PCa cells following its ligand chenodexycholic acid (CDCA) treatment. Methods: Oil Red O was used to stain lipid contents in PCa cells, and siRNA knockdown was performed to deplete FXR expression. To study the cell proliferation when treated by CDCA or FXR knockdown, cell counting kit 8 (CCK8) was adopted to evaluate tumor cell growth. Western blot was used for protein analysis. Results: Our data suggest that activation of FXR by CDCA reduces lipid accumulation and significantly inhibits cells proliferation in prostate tumor cells. Instead, CDCA treatment doesn’t affect normal prostate epithelial RWPE-1 cells growth in vitro. FXR activation decreases mRNA and protein levels of sterol regulatory element binding protein 1 (SREBP1) and some other key regulators involved in lipid metabolism. Depletion of FXR by siRNA attenuates the inhibitory effects. Conclusion: Our study indicates that activation of FXR inhibits lipid metabolism via SREBP1 pathway and further suppresses prostate tumor growth in vitro. PMID:27904713

  12. Farnesoid X receptor inhibits glucagon-like peptide-1 production by enteroendocrine L cells.

    PubMed

    Trabelsi, Mohamed-Sami; Daoudi, Mehdi; Prawitt, Janne; Ducastel, Sarah; Touche, Véronique; Sayin, Sama I; Perino, Alessia; Brighton, Cheryl A; Sebti, Yasmine; Kluza, Jérôme; Briand, Olivier; Dehondt, Hélène; Vallez, Emmanuelle; Dorchies, Emilie; Baud, Grégory; Spinelli, Valeria; Hennuyer, Nathalie; Caron, Sandrine; Bantubungi, Kadiombo; Caiazzo, Robert; Reimann, Frank; Marchetti, Philippe; Lefebvre, Philippe; Bäckhed, Fredrik; Gribble, Fiona M; Schoonjans, Kristina; Pattou, François; Tailleux, Anne; Staels, Bart; Lestavel, Sophie

    2015-07-02

    Bile acids are signalling molecules, which activate the transmembrane receptor TGR5 and the nuclear receptor FXR. BA sequestrants (BAS) complex bile acids in the intestinal lumen and decrease intestinal FXR activity. The BAS-BA complex also induces glucagon-like peptide-1 (GLP-1) production by L cells which potentiates β-cell glucose-induced insulin secretion. Whether FXR is expressed in L cells and controls GLP-1 production is unknown. Here, we show that FXR activation in L cells decreases proglucagon expression by interfering with the glucose-responsive factor Carbohydrate-Responsive Element Binding Protein (ChREBP) and GLP-1 secretion by inhibiting glycolysis. In vivo, FXR deficiency increases GLP-1 gene expression and secretion in response to glucose hence improving glucose metabolism. Moreover, treatment of ob/ob mice with the BAS colesevelam increases intestinal proglucagon gene expression and improves glycaemia in a FXR-dependent manner. These findings identify the FXR/GLP-1 pathway as a new mechanism of BA control of glucose metabolism and a pharmacological target for type 2 diabetes.

  13. Boldine enhances bile production in rats via osmotic and farnesoid X receptor dependent mechanisms.

    PubMed

    Cermanova, Jolana; Kadova, Zuzana; Zagorova, Marie; Hroch, Milos; Tomsik, Pavel; Nachtigal, Petr; Kudlackova, Zdenka; Pavek, Petr; Dubecka, Michaela; Ceckova, Martina; Staud, Frantisek; Laho, Tomas; Micuda, Stanislav

    2015-05-15

    Boldine, the major alkaloid from the Chilean Boldo tree, is used in traditional medicine to support bile production, but evidence to support this function is controversial. We analyzed the choleretic potential of boldine, including its molecular background. The acute- and long-term effects of boldine were evaluated in rats either during intravenous infusion or after 28-day oral treatment. Infusion of boldine instantly increased the bile flow 1.4-fold in healthy rats as well as in animals with Mrp2 deficiency or ethinylestradiol induced cholestasis. This effect was not associated with a corresponding increase in bile acid or glutathione biliary excretion, indicating that the effect is not related to stimulation of either bile acid dependent or independent mechanisms of bile formation and points to the osmotic activity of boldine itself. We subsequently analyzed bile production under conditions of changing biliary excretion of boldine after bolus intravenous administration and found strong correlations between both parameters. HPLC analysis showed that bile concentrations of boldine above 10 μM were required for induction of choleresis. Importantly, long-term pretreatment, when the bile collection study was performed 24-h after the last administration of boldine, also accelerated bile formation despite undetectable levels of the compound in bile. The effect paralleled upregulation of the Bsep transporter and increased biliary clearance of its substrates, bile acids. We consequently confirmed the ability of boldine to stimulate the Bsep transcriptional regulator, FXR receptor. In conclusion, our study clarified the mechanisms and circumstances surrounding the choleretic activity of boldine.

  14. Farnesoid X Receptor Inhibits Glucagon-Like Peptide-1 Production by Enteroendocrine L-cells

    PubMed Central

    TRABELSI, Mohamed-Sami; DAOUDI, Mehdi; PRAWITT, Janne; DUCASTEL, Sarah; TOUCHE, Véronique; SAYIN, Sama I.; PERINO, Alessia; BRIGHTON, Cheryl A.; SEBTI, Yasmine; KLUZA, Jérôme; BRIAND, Olivier; DEHONDT, Hélène; VALLEZ, Emmanuelle; DORCHIES, Emilie; BAUD, Grégory; SPINELLI, Valeria; HENNUYER, Nathalie; CARON, Sandrine; BANTUBUNGI, Kadiombo; CAIAZZO, Robert; REIMANN, Frank; MARCHETTI, Philippe; LEFEBVRE, Philippe; BÄCKHED, Fredrik; GRIBBLE, Fiona M.; SCHOONJANS, Kristina; PATTOU, François; TAILLEUX, Anne; STAELS, Bart; LESTAVEL, Sophie

    2015-01-01

    Bile acids (BA) are signalling molecules which activate the transmembrane receptor TGR5 and the nuclear receptor FXR. BA sequestrants (BAS) complex BA in the intestinal lumen and decrease intestinal FXR activity. The BAS-BA complex also induces Glucagon-Like Peptide-1 (GLP-1) production by L-cells which potentiates β-cell glucose-induced insulin secretion. Whether FXR is expressed in L-cells and controls GLP-1 production is unknown. Here we show that FXR activation in L-cells decreases proglucagon expression by interfering with the glucose-responsive factor Carbohydrate-Responsive Element Binding Protein (ChREBP) and GLP-1 secretion by inhibiting glycolysis. In vivo, FXR-deficiency increases GLP-1 gene expression and secretion in response to glucose hence improving glucose metabolism. Moreover, treatment of ob/ob mice with the BAS colesevelam increases intestinal proglucagon gene expression and improves glycemia in a FXR-dependent manner. These findings identify the FXR/GLP-1 pathway as a new mechanism of BA control of glucose metabolism and a pharmacological target for type 2 diabetes. PMID:26134028

  15. Dihydroartemisinin counteracts fibrotic portal hypertension via farnesoid X receptor-dependent inhibition of hepatic stellate cell contraction.

    PubMed

    Xu, Wenxuan; Lu, Chunfeng; Zhang, Feng; Shao, Jiangjuan; Yao, Shunyu; Zheng, Shizhong

    2017-01-01

    Portal hypertension is a frequent pathological symptom occurring especially in hepatic fibrosis and cirrhosis. Current paradigms indicate that inhibition of hepatic stellate cell (HSC) activation and contraction is anticipated to be an attractive therapeutic strategy, because activated HSC dominantly facilitates an increase in intrahepatic vein pressure through secreting extracellular matrix and contracting. Our previous in vitro study indicated that dihydroartemisinin (DHA) inhibited contractility of cultured HSC by activating intracellular farnesoid X receptor (FXR). However, the effect of DHA on fibrosis-related portal hypertension still requires clarification. In this study, gain- and loss-of-function models of FXR in HSC were established to investigate the mechanisms underlying DHA protection against chronic CCl4 -caused hepatic fibrosis and portal hypertension. Immunofluorescence staining visually showed a decrease in FXR expression in CCl4 -administrated rat HSC but an increase in that in DHA-treated rat HSC. Serum diagnostics and morphological analyses consistently indicated that DHA exhibited hepatoprotective effects on CCl4 -induced liver injury. DHA also reduced CCl4 -caused inflammatory mediator expression and inflammatory cell infiltration. These improvements were further enhanced by INT-747 but weakened by Z-guggulsterone. Noteworthily, DHA, analogous to INT-747, significantly lowered portal vein pressure and suppressed fibrogenesis. Experiments on mice using FXR shRNA lentivirus consolidated the results above. Mechanistically, inhibition of HSC activation and contraction was found as a cellular basis for DHA to relieve portal hypertension. These findings demonstrated that DHA attenuated portal hypertension in fibrotic rodents possibly by targeting HSC contraction via a FXR activation-dependent mechanism. FXR could be a target molecule for reducing portal hypertension during hepatic fibrosis.

  16. Farnesoid X Receptor Regulation of the NLRP3 Inflammasome Underlies Cholestasis-Associated Sepsis.

    PubMed

    Hao, Haiping; Cao, Lijuan; Jiang, Changtao; Che, Yuan; Zhang, Songyang; Takahashi, Shogo; Wang, Guangji; Gonzalez, Frank J

    2017-04-04

    Cholestasis is a common complication of sepsis, and the increased plasma levels of bile acids are predictive of sepsis-associated mortality. However, the exact mechanism by which cholestasis aggravates sepsis development remains elusive. Here, we show that bile acids are danger-associated molecular patterns (DAMPs) that can activate both signal 1 and 2 of the NLRP3 inflammasome in inflammatory macrophages. Mechanistically, bile acids induce a prolonged calcium influx and activate the NLRP3 inflammasome synergistically with ATP. Experimental cholestasis sensitizes, while cholestyramine, a bile acid sequestrant, protects mice from LPS-induced sepsis. FXR negatively regulates the NLRP3 inflammasome via physical interaction with NLRP3 and caspase 1. Fxr-null mice are more sensitive, while FXR-overexpressing mice are more resistant, to endoxemia shock. These findings suggest that bile acids and FXR play pivotal roles in sepsis via controlling the NLRP3 inflammasome, and that targeting FXR may represent a therapeutic strategy for cholestasis-associated sepsis.

  17. Structural Basis for Small Molecule NDB (N-Benzyl-N-(3-(tert-butyl)-4-hydroxyphenyl)-2,6-dichloro-4-(dimethylamino) Benzamide) as a Selective Antagonist of Farnesoid X Receptor α (FXRα) in Stabilizing the Homodimerization of the Receptor*

    PubMed Central

    Xu, Xing; Xu, Xin; Liu, Peng; Zhu, Zhi-yuan; Chen, Jing; Fu, Hai-an; Chen, Li-li; Hu, Li-hong; Shen, Xu

    2015-01-01

    Farnesoid X receptor α (FXRα) as a bile acid sensor plays potent roles in multiple metabolic processes, and its antagonist has recently revealed special interests in the treatment of metabolic disorders, although the underlying mechanisms still remain unclear. Here, we identified that the small molecule N-benzyl-N-(3-(tert-butyl)-4-hydroxyphenyl)-2,6-dichloro-4-(dimethylamino) benzamide (NDB) functioned as a selective antagonist of human FXRα (hFXRα), and the crystal structure of hFXRα ligand binding domain (hFXRα-LBD) in complex with NDB was analyzed. It was unexpectedly discovered that NDB induced rearrangements of helix 11 (H11) and helix 12 (H12, AF-2) by forming a homodimer of hFXRα-LBD, totally different from the active conformation in monomer state, and the binding details were further supported by the mutation analysis. Moreover, functional studies demonstrated that NDB effectively antagonized the GW4064-stimulated FXR/RXR interaction and FXRα target gene expression in primary mouse hepatocytes, including the small heterodimer partner (SHP) and bile-salt export pump (BSEP); meanwhile, administration of NDB to db/db mice efficiently decreased the gene expressions of phosphoenolpyruvate carboxykinase (PEPCK), glucose 6-phosphatase (G6-pase), small heterodimer partner, and BSEP. It is expected that our first analyzed crystal structure of hFXRα-LBD·NDB will help expound the antagonistic mechanism of the receptor, and NDB may find its potential as a lead compound in anti-diabetes research. PMID:26100621

  18. Modulation of farnesoid X receptor results in post-translational modification of poly (ADP-ribose) polymerase 1 in the liver

    SciTech Connect

    Zhu, Yan; Li, Guodong; Dong, Yafeng; Zhou, Helen H.; Kong, Bo; Aleksunes, Lauren M.; Richardson, Jason R.; Li, Fei; Guo, Grace L.

    2013-01-15

    The farnesoid X receptor (FXR) is a bile acid-activated transcription factor belonging to the nuclear receptor superfamily. FXR deficiency in mice results in cholestasis, metabolic disorders, and tumorigenesis in liver and intestine. FXR is known to contribute to pathogenesis by regulating gene transcription; however, changes in the post-transcriptional modification of proteins associated with FXR modulation have not been determined. In the current study, proteomic analysis of the livers of wild-type (WT) and FXR knockout (FXR-KO) mice treated with a FXR synthetic ligand or vehicle was performed. The results identified five proteins as novel FXR targets. Since FXR deficiency in mice leads to liver tumorigenesis, poly (ADP-ribose) polymerase family, member 1 (Parp1) that is important for DNA repair, was validated in the current study by quantitative real-time PCR, and 1- and 2-dimensional gel electrophoresis/western blot. The results showed that Parp1 mRNA levels were not altered by FXR genetic status or by agonist treatment. However, total Parp1 protein levels were increased in FXR-KO mice as early as 3 month old. Interestingly, total Parp1 protein levels were increased in WT mice in an age-dependent manner (from 3 to 18 months), but not in FXR-KO mice. Finally, activation of FXR in WT mice resulted in reduction of phosporylated Parp1 protein in the liver without affecting total Parp1 protein levels. In conclusion, this study reveals that FXR genetic status and agonist treatment affects basal levels and phosphorylation state of Parp1, respectively. These alterations, in turn, may be associated with the hepatobiliary alterations observed in FXR-KO mice and participate in FXR agonist-induced protection in the liver. -- Highlights: ► Proteomic analysis identified novel FXR targets. ► FXR modification altered post-translational modification of the Parp1 protein. ► Altered Parp1 function may contribute to mechanisms of FXR regulation of liver functions.

  19. Optimization of a novel class of benzimidazole-based farnesoid X receptor (FXR) agonists to improve physicochemical and ADME properties.

    PubMed

    Richter, Hans G F; Benson, G M; Bleicher, K H; Blum, D; Chaput, E; Clemann, N; Feng, S; Gardes, C; Grether, U; Hartman, P; Kuhn, B; Martin, R E; Plancher, J-M; Rudolph, M G; Schuler, F; Taylor, S

    2011-02-15

    Structure-guided lead optimization of recently described benzimidazolyl acetamides addressed the key liabilities of the previous lead compound 1. These efforts culminated in the discovery of 4-{(S)-2-[2-(4-chloro-phenyl)-5,6-difluoro-benzoimidazol-1-yl]-2-cyclohexyl-acetylamino}-3-fluoro-benzoic acid 7g, a highly potent and selective FXR agonist with excellent physicochemical and ADME properties and potent lipid lowering activity after oral administration to LDL receptor deficient mice.

  20. Polyphenols rich Passiflora leschenaultii leaves modulating Farnesoid X Receptor and Pregnane X Receptor against paracetamol-induced hepatotoxicity in rats.

    PubMed

    Shanmugam, Saravanan; Sivaraj, Dhivya; Dos Santos Lima, Bruno; Dos Passos Menezes, Paula; de Carvalho, Yasmim Maria Barbosa Gomes; de Souza Araújo, Adriano Antunes; Narain, Narendra; Serafini, Mairim Russo; Quintans Júnior, Lucindo José; Scotti, Luciana; Scotti, Marcus Tulius; Parimelazhagan, Thangaraj

    2017-04-01

    The hepatoprotective effect of P. leschenaultii. (DC) leaves was investigated in rats under paracetamol induced oxidative stress. Leaf acetone extract (200 and 400mg/kg) were administered daily via gavage for 14days before paracetamol (2000mg/kg, p.o.) treatment. After the experiment, the levels of serum biochemical parameters and enzymatic antioxidant levels were determined. Furthermore, liver tissues were analyzed histopathologically. Additionally, the molecular docking studies of the identified compounds against PXR and FXR proteins were also performed. The assessment revealed that the acetone extract significantly reduced the elevated levels of SGPT, SGOT and ALP in serum. Moreover, the enzymatic antioxidants such as SOD, CAT and LPO were also retained normally by the plant extract. From histopathological analysis, it was clearly evident that the cellular architecture of plant extract treated rat liver tissues were not affected by the paracetamol induction at the higher dose. The results of docking studies also revealed that the identified compounds showed steric interactions (between nonpolar atoms) with amino acid groups. Collectively, the present study suggests that P. leschenaultii leaves extract protects the liver from paracetamol induced hepatic damage.

  1. Farnesoid X receptor up-regulates expression of Lipid transfer inhibitor protein in liver cells and mice

    SciTech Connect

    Li, Liangpeng; Liu, Hong; Peng, Jiahe; Wang, Yongchao; Zhang, Yan; Dong, Jinyu; Liu, Xiaohua; Guo, Dongmei; Jiang, Yu

    2013-11-29

    Highlights: •FXR up-regulates apoF. •It binds to ER1 element. •It activates apoF gene promoter. -- Abstract: Apolipoprotein F is a component protein mainly secreted by liver and resides on several lipoprotein classes. It can inhibit lipids transfer between different lipoproteins. FXR is a member of the nuclear receptor superfamily which is also highly expressed in the liver. It modulates bile acids synthesis and lipids metabolism by transcriptional regulation. We aimed to determine whether apoF can be regulated by FXR. The FXR agonist Chenodeoxycholic acid (CDCA) and GW4064 both can activate the expression of apoF in liver cell lines and in C57/BL6 mouse liver. This is dependent on the binding of FXR to the FXR element ER1 (−2904 to −2892 bp) in the apoF gene promoter. Taken together, we have identified apoF as likely another target gene of FXR.

  2. Interleukin-18 Down-Regulates Multidrug Resistance-Associated Protein 2 Expression through Farnesoid X Receptor Associated with Nuclear Factor Kappa B and Yin Yang 1 in Human Hepatoma HepG2 Cells.

    PubMed

    Liu, Xiao-cong; Lian, Wei; Zhang, Liang-jun; Feng, Xin-chan; Gao, Yu; Li, Shao-xue; Liu, Chang; Cheng, Ying; Yang, Long; Wang, Xiao-Juan; Chen, Lei; Wang, Rong-quan; Chai, Jin; Chen, Wen-sheng

    2015-01-01

    Multidrug resistance-associated protein 2 (MRP2) plays an important role in bile acid metabolism by transporting toxic organic anion conjugates, including conjugated bilirubin, glutathione, sulfate, and multifarious drugs. MRP2 expression is reduced in cholestatic patients and rodents. However, the molecular mechanism of MRP2 down-regulation remains elusive. In this report, we treated human hepatoma HepG2 cells with interleukin-18 (IL-18) and measured the expression of MRP2, nuclear factor kappa B (NF-κB), farnesoid X receptor (FXR), and the transcription factor Yin Yang 1 (YY1) by quantitative real-time quantitative polymerase chain reaction (PCR) and western blotting. We found that expression of MRP2 was repressed by IL-18 at both the mRNA and protein levels in a dose- and time-dependent manner. Furthermore, the activated NF-κB pathway increased YY1 and reduced FXR. These changes were all attenuated in HepG2 cells with knockdown of the NF-κB subunit, p65. The reduced expression of FXR and MRP2 in HepG2 cells that had been caused by IL-18 treatment was also attenuated by YY1 knockdown. We further observed significantly elevated IL-18, NF-κB, and YY1 expression and decreased FXR and MRP2 expression in bile duct-ligated Sprague Dawley rat livers. Chromatin immunoprecipitation assays also showed that FXR bound to the promoter region in MRP2 was less abundant in liver extracts from bile duct-ligated rats than sham-operated rats. Our findings indicate that IL-18 down-regulates MRP2 expression through the nuclear receptor FXR in HepG2 cells, and may be mediated by NF-κB and YY1.

  3. Activation of the Farnesoid X-receptor in breast cancer cell lines results in cytotoxicity but not increased migration potential.

    PubMed

    Alasmael, Noura; Mohan, Rati; Meira, Lisiane B; Swales, Karen E; Plant, Nick J

    2016-01-28

    Breast cancer is the commonest form of cancer in women, but successful treatment is confounded by the heterogeneous nature of breast tumours: Effective treatments exist for hormone-sensitive tumours, but triple-negative breast cancer results in poor survival. An area of increasing interest is metabolic reprogramming, whereby drug-induced alterations in the metabolic landscape of a tumour slow tumour growth and/or increase sensitivity to existing therapeutics. Nuclear receptors are transcription factors central to the expression of metabolic and transport proteins, and thus represent potential targets for metabolic reprogramming. We show that activation of the nuclear receptor FXR, either by its endogenous ligand CDCA or the synthetic GW4064, leads to cell death in four breast cancer cell lines with distinct phenotypes: MCF-10A (normal), MCF-7 (receptor positive), MDA-MB-231 and MDA-MB-468 (triple negative). Furthermore, we show that the mechanism of cell death is predominantly through the intrinsic apoptotic pathway. Finally, we demonstrate that FXR agonists do not stimulate migration in breast cancer cell lines, an important potential adverse effect. Together, our data support the continued examination of FXR agonists as a novel class of therapeutics for the treatment of breast cancer.

  4. Hepatic Farnesoid X-Receptor Isoforms α2 and α4 Differentially Modulate Bile Salt and Lipoprotein Metabolism in Mice

    PubMed Central

    Boesjes, Marije; Bloks, Vincent W.; Hageman, Jurre; Bos, Trijnie; van Dijk, Theo H.; Havinga, Rick; Wolters, Henk; Jonker, Johan W.; Kuipers, Folkert; Groen, Albert K.

    2014-01-01

    The nuclear receptor FXR acts as an intracellular bile salt sensor that regulates synthesis and transport of bile salts within their enterohepatic circulation. In addition, FXR is involved in control of a variety of crucial metabolic pathways. Four FXR splice variants are known, i.e. FXRα1-4. Although these isoforms show differences in spatial and temporal expression patterns as well as in transcriptional activity, the physiological relevance hereof has remained elusive. We have evaluated specific roles of hepatic FXRα2 and FXRα4 by stably expressing these isoforms using liver-specific self-complementary adeno-associated viral vectors in total body FXR knock-out mice. The hepatic gene expression profile of the FXR knock-out mice was largely normalized by both isoforms. Yet, differential effects were also apparent; FXRα2 was more effective in reducing elevated HDL levels and transrepressed hepatic expression of Cyp8b1, the regulator of cholate synthesis. The latter coincided with a switch in hydrophobicity of the bile salt pool. Furthermore, FXRα2-transduction caused an increased neutral sterol excretion compared to FXRα4 without affecting intestinal cholesterol absorption. Our data show, for the first time, that hepatic FXRα2 and FXRα4 differentially modulate bile salt and lipoprotein metabolism in mice. PMID:25506828

  5. Bile acid receptors and nonalcoholic fatty liver disease

    PubMed Central

    Yuan, Liyun; Bambha, Kiran

    2015-01-01

    With the high prevalence of obesity, diabetes, and other features of the metabolic syndrome in United States, nonalcoholic fatty liver disease (NAFLD) has inevitably become a very prevalent chronic liver disease and is now emerging as one of the leading indications for liver transplantation. Insulin resistance and derangement of lipid metabolism, accompanied by activation of the pro-inflammatory response and fibrogenesis, are essential pathways in the development of the more clinically significant form of NAFLD, known as nonalcoholic steatohepatitis (NASH). Recent advances in the functional characterization of bile acid receptors, such as farnesoid X receptor (FXR) and transmembrane G protein-coupled receptor (TGR) 5, have provided further insight in the pathophysiology of NASH and have led to the development of potential therapeutic targets for NAFLD and NASH. Beyond maintaining bile acid metabolism, FXR and TGR5 also regulate lipid metabolism, maintain glucose homeostasis, increase energy expenditure, and ameliorate hepatic inflammation. These intriguing features have been exploited to develop bile acid analogues to target pathways in NAFLD and NASH pathogenesis. This review provides a brief overview of the pathogenesis of NAFLD and NASH, and then delves into the biological functions of bile acid receptors, particularly with respect to NASH pathogenesis, with a description of the associated experimental data, and, finally, we discuss the prospects of bile acid analogues in the treatment of NAFLD and NASH. PMID:26668692

  6. Retinoic Acid-mediated Nuclear Receptor Activation and Hepatocyte Proliferation

    PubMed Central

    Bushue, Nathan; Wan, Yu-Jui Yvonne

    2016-01-01

    Due to their well-known differentiation and apoptosis-inducing abilities, retinoic acid (RA) and its analogs have strong anti-cancer efficacy in human cancers. However, in vivo RA is a liver mitogen. While speculation has persisted that RA-mediated signaling is likely involved in hepatocyte proliferation during liver regeneration, direct evidence is still required. Findings in support of this proposition include observations that a release of retinyl palmitate (the precursor of RA) occurs in liver stellate cells following liver injury. Nevertheless, the biological action of this released vitamin A is virtually unknown. More likely is that the released vitamin A is converted to RA, the biological form, and then bound to a specific receptor (retinoid x receptor; RXRα), which is most abundantly expressed in the liver. Considering the mitogenic effects of RA, the RA-activated RXRα would likely then influence hepatocyte proliferation and liver tissue repair. At present, the mechanism by which RA stimulates hepatocyte proliferation is largely unknown. This review summarizes the activation of nuclear receptors (peroxisome proliferator activated receptor-α, pregnane x receptor, constitutive androstane receptor, and farnesoid x receptor) in an RXRα dependent manner to induce hepatocyte proliferation, providing a link between RA and its proliferative role. PMID:27635169

  7. Mutations in the nuclear bile acid receptor FXR cause progressive familial intrahepatic cholestasis

    PubMed Central

    Gomez-Ospina, Natalia; Potter, Carol J.; Xiao, Rui; Manickam, Kandamurugu; Kim, Mi-Sun; Kim, Kang Ho; Shneider, Benjamin L.; Picarsic, Jennifer L.; Jacobson, Theodora A.; Zhang, Jing; He, Weimin; Liu, Pengfei; Knisely, A. S.; Finegold, Milton J.; Muzny, Donna M.; Boerwinkle, Eric; Lupski, James R.; Plon, Sharon E.; Gibbs, Richard A.; Eng, Christine M.; Yang, Yaping; Washington, Gabriel C.; Porteus, Matthew H.; Berquist, William E.; Kambham, Neeraja; Singh, Ravinder J.; Xia, Fan; Enns, Gregory M.; Moore, David D.

    2016-01-01

    Neonatal cholestasis is a potentially life-threatening condition requiring prompt diagnosis. Mutations in several different genes can cause progressive familial intrahepatic cholestasis, but known genes cannot account for all familial cases. Here we report four individuals from two unrelated families with neonatal cholestasis and mutations in NR1H4, which encodes the farnesoid X receptor (FXR), a bile acid-activated nuclear hormone receptor that regulates bile acid metabolism. Clinical features of severe, persistent NR1H4-related cholestasis include neonatal onset with rapid progression to end-stage liver disease, vitamin K-independent coagulopathy, low-to-normal serum gamma-glutamyl transferase activity, elevated serum alpha-fetoprotein and undetectable liver bile salt export pump (ABCB11) expression. Our findings demonstrate a pivotal function for FXR in bile acid homeostasis and liver protection. PMID:26888176

  8. Regulation of antibacterial defense in the small intestine by the nuclear bile acid receptor

    PubMed Central

    Inagaki, Takeshi; Moschetta, Antonio; Lee, Youn-Kyoung; Peng, Li; Zhao, Guixiang; Downes, Michael; Yu, Ruth T.; Shelton, John M.; Richardson, James A.; Repa, Joyce J.; Mangelsdorf, David J.; Kliewer, Steven A.

    2006-01-01

    Obstruction of bile flow results in bacterial proliferation and mucosal injury in the small intestine that can lead to the translocation of bacteria across the epithelial barrier and systemic infection. These adverse effects of biliary obstruction can be inhibited by administration of bile acids. Here we show that the farnesoid X receptor (FXR), a nuclear receptor for bile acids, induces genes involved in enteroprotection and inhibits bacterial overgrowth and mucosal injury in ileum caused by bile duct ligation. Mice lacking FXR have increased ileal levels of bacteria and a compromised epithelial barrier. These findings reveal a central role for FXR in protecting the distal small intestine from bacterial invasion and suggest that FXR agonists may prevent epithelial deterioration and bacterial translocation in patients with impaired bile flow. PMID:16473946

  9. Acetylated deoxycholic (DCA) and cholic (CA) acids are potent ligands of pregnane X (PXR) receptor.

    PubMed

    Carazo, Alejandro; Hyrsova, Lucie; Dusek, Jan; Chodounska, Hana; Horvatova, Alzbeta; Berka, Karel; Bazgier, Vaclav; Gan-Schreier, Hongying; Chamulitrat, Waleé; Kudova, Eva; Pavek, Petr

    2017-01-04

    The Pregnane X (PXR), Vitamin D (VDR) and Farnesoid X (FXR) nuclear receptors have been shown to be receptors of bile acids controlling their detoxification or synthesis. Chenodeoxycholic (CDCA) and lithocholic (LCA) acids are ligands of FXR and VDR, respectively, whereas 3-keto and acetylated derivates of LCA have been described as ligands for all three receptors. In this study, we hypothesized that oxidation or acetylation at position 3, 7 and 12 of bile acids DCA (deoxycholic acid), LCA, CA (cholic acid), and CDCA by detoxification enzymes or microbiome may have an effect on the interactions with bile acid nuclear receptors. We employed reporter gene assays in HepG2 cells, the TR-FRET assay with recombinant PXR and RT-PCR to study the effects of acetylated and keto bile acids on the nuclear receptors activation and their target gene expression in differentiated hepatic HepaRG cells. We demonstrate that the DCA 3,12-diacetate and CA 3,7,12-triacetate derivatives are ligands of PXR and DCA 3,12-diacetate induces PXR target genes such as CYP3A4, CYP2B6 and ABCB1/MDR1. In conclusion, we found that acetylated DCA and CA are potent ligands of PXR. Whether the acetylated bile acid derivatives are novel endogenous ligands of PXR with detoxification or physiological functions should be further studied in ongoing experiments.

  10. Bile Acids Trigger GLP-1 Release Predominantly by Accessing Basolaterally Located G Protein-Coupled Bile Acid Receptors.

    PubMed

    Brighton, Cheryl A; Rievaj, Juraj; Kuhre, Rune E; Glass, Leslie L; Schoonjans, Kristina; Holst, Jens J; Gribble, Fiona M; Reimann, Frank

    2015-11-01

    Bile acids are well-recognized stimuli of glucagon-like peptide-1 (GLP-1) secretion. This action has been attributed to activation of the G protein-coupled bile acid receptor GPBAR1 (TGR5), although other potential bile acid sensors include the nuclear farnesoid receptor and the apical sodium-coupled bile acid transporter ASBT. The aim of this study was to identify pathways important for GLP-1 release and to determine whether bile acids target their receptors on GLP-1-secreting L-cells from the apical or basolateral compartment. Using transgenic mice expressing fluorescent sensors specifically in L-cells, we observed that taurodeoxycholate (TDCA) and taurolithocholate (TLCA) increased intracellular cAMP and Ca(2+). In primary intestinal cultures, TDCA was a more potent GLP-1 secretagogue than taurocholate (TCA) and TLCA, correlating with a stronger Ca(2+) response to TDCA. Using small-volume Ussing chambers optimized for measuring GLP-1 secretion, we found that both a GPBAR1 agonist and TDCA stimulated GLP-1 release better when applied from the basolateral than from the luminal direction and that luminal TDCA was ineffective when intestinal tissue was pretreated with an ASBT inhibitor. ASBT inhibition had no significant effect in nonpolarized primary cultures. Studies in the perfused rat gut confirmed that vascularly administered TDCA was more effective than luminal TDCA. Intestinal primary cultures and Ussing chamber-mounted tissues from GPBAR1-knockout mice did not secrete GLP-1 in response to either TLCA or TDCA. We conclude that the action of bile acids on GLP-1 secretion is predominantly mediated by GPBAR1 located on the basolateral L-cell membrane, suggesting that stimulation of gut hormone secretion may include postabsorptive mechanisms.

  11. G-protein-coupled bile acid receptor plays a key role in bile acid metabolism and fasting-induced hepatic steatosis in mice.

    PubMed

    Donepudi, Ajay C; Boehme, Shannon; Li, Feng; Chiang, John Y L

    2017-03-01

    Bile acids are signaling molecules that play a critical role in regulation of hepatic metabolic homeostasis by activating nuclear farnesoid X receptor (Fxr) and membrane G-protein-coupled receptor (Takeda G-protein-coupled receptor 5; Tgr5). The role of FXR in regulation of bile acid synthesis and hepatic metabolism has been studied extensively. However, the role of TGR5 in hepatic metabolism has not been explored. The liver plays a central role in lipid metabolism, and impaired response to fasting and feeding contributes to steatosis and nonalcoholic fatty liver and obesity. We have performed a detailed analysis of gallbladder bile acid and lipid metabolism in Tgr5(-/-) mice in both free-fed and fasted conditions. Lipid profiles of serum, liver and adipose tissues, bile acid composition, energy metabolism, and messenger RNA and protein expression of the genes involved in lipid metabolism were analyzed. Results showed that deficiency of the Tgr5 gene in mice alleviated fasting-induced hepatic lipid accumulation. Expression of liver oxysterol 7α-hydroxylase in the alternative bile acid synthesis pathway was reduced. Analysis of gallbladder bile acid composition showed marked increase of taurocholic acid and decrease of tauro-α and β-muricholic acid in Tgr5(-/-) mice. Tgr5(-/-) mice had increased hepatic fatty acid oxidation rate and decreased hepatic fatty acid uptake. Interestingly, fasting induction of fibroblast growth factor 21 in liver was attenuated. In addition, fasted Tgr5(-/-) mice had increased activation of hepatic growth hormone-signal transducer and activator of transcription 5 (GH-Stat5) signaling compared to wild-type mice.

  12. Transcriptional integration of metabolism by the nuclear sterol-activated receptors LXR and FXR.

    PubMed

    Calkin, Anna C; Tontonoz, Peter

    2012-03-14

    Nuclear receptors are integrators of hormonal and nutritional signals, mediating changes to metabolic pathways within the body. Given that modulation of lipid and glucose metabolism has been linked to diseases including type 2 diabetes, obesity and atherosclerosis, a greater understanding of pathways that regulate metabolism in physiology and disease is crucial. The liver X receptors (LXRs) and the farnesoid X receptors (FXRs) are activated by oxysterols and bile acids, respectively. Mounting evidence indicates that these nuclear receptors have essential roles, not only in the regulation of cholesterol and bile acid metabolism but also in the integration of sterol, fatty acid and glucose metabolism.

  13. Transcriptional integration of metabolism by the nuclear sterol-activated receptors LXR and FXR

    PubMed Central

    2013-01-01

    Nuclear receptors are integrators of hormonal and nutritional signals, mediating changes to metabolic pathways within the body. Given that modulation of lipid and glucose metabolism has been linked to diseases including type 2 diabetes, obesity and atherosclerosis, a greater understanding of pathways that regulate metabolism in physiology and disease is crucial. The liver X receptors (LXRs) and the farnesoid X receptors (FXRs) are activated by oxysterols and bile acids, respectively. Mounting evidence indicates that these nuclear receptors have essential roles, not only in the regulation of cholesterol and bile acid metabolism but also in the integration of sterol, fatty acid and glucose metabolism. PMID:22414897

  14. Obeticholic Acid

    MedlinePlus

    Obeticholic acid is used alone or in combination with ursodiol (Actigall, Urso) to treat primary biliary cholangitis (PBC; a ... were not treated successfully with ursodiol alone. Obeticholic acid is in a class of medications called farnesoid ...

  15. Activation of transmembrane bile acid receptor TGR5 stimulates insulin secretion in pancreatic β cells

    PubMed Central

    Kumar, Divya P.; Rajagopal, Senthilkumar; Mahavadi, Sunila; Mirshahi, Faridoddin; Grider, John R.; Murthy, Karnam S.; Sanyal, Arun J.

    2012-01-01

    Bile acids act as signaling molecules and stimulate the G protein coupled receptor, TGR5, in addition to nuclear farnesoid X receptor to regulate lipid, glucose and energy metabolism. Bile acid induced activation of TGR5 in the enteroendocrine cells promotes glucagon like peptide-1 (GLP-1) release, which has insulinotropic effect in the pancreatic β cells. In the present study, we have identified the expression of TGR5 in pancreatic β cell line MIN6 and also in mouse and human pancreatic islets. TGR5 selective ligands, oleanolic acid (OA) and INT-777 selectively activated Gαs and caused an increase in intracellular cAMP and Ca2+. OA and INT-777 also increased phosphoinositide (PI) hydrolysis and the increase was blocked by NF449 (a selective Gαs inhibitor) or U73122 (PI hydrolysis inhibitor). OA, INT-777 and lithocholic acid increased insulin release in MIN6 and human islets and the increase was inhibited by treatment with NF449, U73122 or BAPTA-AM (chelator of calcium), but not with myristoylated PKI (PKA inhibitor), suggesting that the release is dependent on Gs/cAMP/Ca2+ pathway. 8-pCPT-2′-O-Me-cAMP, a cAMP analogue, which activates Epac, but not PKA also stimulated PI hydrolysis. In conclusion, our study demonstrates that the TGR5 expressed in the pancreatic β cells regulates insulin secretion and highlights the importance of ongoing therapeutic strategies targeting TGR5 in the control of glucose homeostasis. PMID:23022524

  16. Altered activity profile of a tertiary silanol analog of multi-targeting nuclear receptor modulator T0901317.

    PubMed

    Toyama, Hirozumi; Sato, Shoko; Shirakawa, Hitoshi; Komai, Michio; Hashimoto, Yuichi; Fujii, Shinya

    2016-04-01

    We report the design, synthesis, and physicochemical/biological evaluation of novel silanol derivative 6 (sila-T) as a silanol analog of multi-target nuclear receptor modulator T0901317 (5). Compound 6 showed intermediate hydrophobicity between the corresponding alcohol 13 and perfluoroalcohol 5. While 5 exhibited potent activities toward liver X receptor α and β, farnesoid X receptor, pregnane X receptor (PXR) and retinoic acid receptor-related orphan receptor (ROR)γ, silanol 6 exhibited activity only toward PXR and RORs. Incorporation of silanol instead of perfluoroalcohol is a promising option for developing novel target-selective, biologically active compounds.

  17. Bile Acids, Obesity, and the Metabolic Syndrome

    PubMed Central

    Ma, Huijuan; Patti, Mary Elizabeth

    2014-01-01

    Bile acids are increasingly recognized as key regulators of systemic metabolism. While bile acids have long been known to play important and direct roles in nutrient absorption, bile acids also serve as signaling molecules. Bile acid interactions with the nuclear hormone receptor farnesoid X receptor (FXR) and the membrane receptor G-protein-coupled bile acid receptor 5 (TGR5) can regulate incretin hormone and fibroblast growth factor 19 (FGF19) secretion, cholesterol metabolism, and systemic energy expenditure. Bile acid levels and distribution are altered in type 2 diabetes and increased following bariatric procedures, in parallel with reduced body weight and improved insulin sensitivity and glycemic control. Thus, modulation of bile acid levels and signaling, using bile acid binding resins, TGR5 agonists, and FXR agonists, may serve as a potent therapeutic approach for the treatment of obesity, type 2 diabetes, and other components of the metabolic syndrome in humans. PMID:25194176

  18. Activation of the nuclear receptor FXR improves hyperglycemia and hyperlipidemia in diabetic mice

    NASA Astrophysics Data System (ADS)

    Zhang, Yanqiao; Lee, Florence Ying; Barrera, Gabriel; Lee, Hans; Vales, Charisse; Gonzalez, Frank J.; Willson, Timothy M.; Edwards, Peter A.

    2006-01-01

    Farnesoid X receptor (FXR) plays an important role in maintaining bile acid and cholesterol homeostasis. Here we demonstrate that FXR also regulates glucose metabolism. Activation of FXR by the synthetic agonist GW4064 or hepatic overexpression of constitutively active FXR by adenovirus-mediated gene transfer significantly lowered blood glucose levels in both diabetic db/db and wild-type mice. Consistent with these data, FXR null mice exhibited glucose intolerance and insulin insensitivity. We further demonstrate that activation of FXR in db/db mice repressed hepatic gluconeogenic genes and increased hepatic glycogen synthesis and glycogen content by a mechanism that involves enhanced insulin sensitivity. In view of its central roles in coordinating regulation of both glucose and lipid metabolism, we propose that FXR agonists are promising therapeutic agents for treatment of diabetes mellitus. glucose | GW4064 | farnesoid X receptor-VP16 | triglyceride | cholesterol

  19. Activation of transmembrane bile acid receptor TGR5 stimulates insulin secretion in pancreatic {beta} cells

    SciTech Connect

    Kumar, Divya P.; Rajagopal, Senthilkumar; Mahavadi, Sunila; Mirshahi, Faridoddin; Grider, John R.; Murthy, Karnam S.; Sanyal, Arun J.

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer G protein coupled receptor TGR5 is expressed in mouse and human islets. Black-Right-Pointing-Pointer TGR5 is coupled to activation of Gs and Ca{sup 2+} release via cAMP/Epac/PLC-{epsilon} pathway. Black-Right-Pointing-Pointer Activation of TGR5 by bile salts and selective ligands causes insulin secretion. Black-Right-Pointing-Pointer TGR5 could be a potential therapeutic target to treat diabetes. -- Abstract: Bile acids act as signaling molecules and stimulate the G protein coupled receptor, TGR5, in addition to nuclear farnesoid X receptor to regulate lipid, glucose and energy metabolism. Bile acid induced activation of TGR5 in the enteroendocrine cells promotes glucagon like peptide-1 (GLP-1) release, which has insulinotropic effect in the pancreatic {beta} cells. In the present study, we have identified the expression of TGR5 in pancreatic {beta} cell line MIN6 and also in mouse and human pancreatic islets. TGR5 selective ligands, oleanolic acid (OA) and INT-777 selectively activated G{alpha}{sub s} and caused an increase in intracellular cAMP and Ca{sup 2+}. OA and INT-777 also increased phosphoinositide (PI) hydrolysis and the increase was blocked by NF449 (a selective G{alpha}{sub s} inhibitor) or (U73122) (PI hydrolysis inhibitor). OA, INT-777 and lithocholic acid increased insulin release in MIN6 and human islets and the increase was inhibited by treatment with NF449, (U73122) or BAPTA-AM (chelator of calcium), but not with myristoylated PKI (PKA inhibitor), suggesting that the release is dependent on G{sub s}/cAMP/Ca{sup 2+} pathway. 8-pCPT-2 Prime -O-Me-cAMP, a cAMP analog, which activates Epac, but not PKA also stimulated PI hydrolysis. In conclusion, our study demonstrates that the TGR5 expressed in the pancreatic {beta} cells regulates insulin secretion and highlights the importance of ongoing therapeutic strategies targeting TGR5 in the control of glucose homeostasis.

  20. Acid detection by taste receptor cells.

    PubMed

    DeSimone, J A; Lyall, V; Heck, G L; Feldman, G M

    2001-12-01

    Sourness is a primary taste quality that evokes an innate rejection response in humans and many other animals. Acidic stimuli are the unique sources of sour taste so a rejection response may serve to discourage ingestion of foods spoiled by acid producing microorganisms. The investigation of mechanisms by which acids excite taste receptor cells (TRCs) is complicated by wide species variability and within a species, apparently different mechanisms for strong and weak acids. The problem is further complicated by the fact that the receptor cells are polarized epithelial cells with different apical and basolateral membrane properties. The cellular mechanisms proposed for acid sensing in taste cells include, the direct blockage of apical K(+) channels by protons, an H(+)-gated Ca(2+) channel, proton conduction through apical amiloride-blockable Na(+) channels, a Cl(-) conductance blocked by NPPB, the activation of the proton-gated channel, BNC-1, a member of the Na(+) channel/degenerin super family, and by stimulus-evoked changes in intracellular pH. Acid-induced intracellular pH changes appear to be similar to those reported in other mammalian acid-sensing cells, such as type-I cells of the carotid body, and neurons found in the ventrolateral medulla, nucleus of the solitary tract, the medullary raphe, and the locus coceuleus. Like type-I carotid body cells and brainstem neurons, isolated TRCs demonstrate a linear relationship between intracellular pH (pH(i)) and extracellular pH (pH(o)) with slope, DeltapH(i)/DeltapH(o) near unity. Acid-sensing cells also appear to regulate pH(i) when intracellular pH changes occur under iso-extracellular pH conditions, but fail to regulate their pH when pH(i) changes are induced by decreasing extracellular pH. We shall discuss the current status of proposed acid-sensing taste mechanisms, emphasizing pH-tracking in receptor cells.

  1. Evolution of retinoic acid receptors and retinoic acid signaling.

    PubMed

    Gutierrez-Mazariegos, Juliana; Schubert, Michael; Laudet, Vincent

    2014-01-01

    Retinoic acid (RA) is a vitamin A-derived morphogen controlling important developmental processes in vertebrates, and more generally in chordates, including axial patterning and tissue formation and differentiation. In the embryo, endogenous RA levels are controlled by RA synthesizing and degrading enzymes and the RA signal is transduced by two retinoid receptors: the retinoic acid receptor (RAR) and the retinoid X receptor (RXR). Both RAR and RXR are members of the nuclear receptor superfamily of ligand-activated transcription factors and mainly act as heterodimers to activate the transcription of target genes in the presence of their ligand, all-trans RA. This signaling pathway was long thought to be a chordate innovation, however, recent findings of gene homologs involved in RA signaling in the genomes of a wide variety of non-chordate animals, including ambulacrarians (sea urchins and acorn worms) and lophotrochozoans (annelids and mollusks), challenged this traditional view and suggested that the RA signaling pathway might have a more ancient evolutionary origin than previously thought. In this chapter, we discuss the evolutionary history of the RA signaling pathway, and more particularly of the RARs, which might have experienced independent gene losses and duplications in different animal lineages. In sum, the available data reveal novel insights into the origin of the RA signaling pathway as well as into the evolutionary history of the RARs.

  2. Role of bile acids in the regulation of the metabolic pathways

    PubMed Central

    Taoka, Hiroki; Yokoyama, Yoko; Morimoto, Kohkichi; Kitamura, Naho; Tanigaki, Tatsuya; Takashina, Yoko; Tsubota, Kazuo; Watanabe, Mitsuhiro

    2016-01-01

    Recent studies have revealed that bile acids (BAs) are not only facilitators of dietary lipid absorption but also important signaling molecules exerting multiple physiological functions. Some major signaling pathways involving the nuclear BAs receptor farnesoid X receptor and the G protein-coupled BAs receptor TGR5/M-BAR have been identified to be the targets of BAs. BAs regulate their own homeostasis via signaling pathways. BAs also affect diverse metabolic pathways including glucose metabolism, lipid metabolism and energy expenditure. This paper suggests the mechanism of controlling metabolism via BA signaling and demonstrates that BA signaling is an attractive therapeutic target of the metabolic syndrome. PMID:27433295

  3. Bile acids: regulation of synthesis.

    PubMed

    Chiang, John Y L

    2009-10-01

    Bile acids are physiological detergents that generate bile flow and facilitate intestinal absorption and transport of lipids, nutrients, and vitamins. Bile acids also are signaling molecules and inflammatory agents that rapidly activate nuclear receptors and cell signaling pathways that regulate lipid, glucose, and energy metabolism. The enterohepatic circulation of bile acids exerts important physiological functions not only in feedback inhibition of bile acid synthesis but also in control of whole-body lipid homeostasis. In the liver, bile acids activate a nuclear receptor, farnesoid X receptor (FXR), that induces an atypical nuclear receptor small heterodimer partner, which subsequently inhibits nuclear receptors, liver-related homolog-1, and hepatocyte nuclear factor 4alpha and results in inhibiting transcription of the critical regulatory gene in bile acid synthesis, cholesterol 7alpha-hydroxylase (CYP7A1). In the intestine, FXR induces an intestinal hormone, fibroblast growth factor 15 (FGF15; or FGF19 in human), which activates hepatic FGF receptor 4 (FGFR4) signaling to inhibit bile acid synthesis. However, the mechanism by which FXR/FGF19/FGFR4 signaling inhibits CYP7A1 remains unknown. Bile acids are able to induce FGF19 in human hepatocytes, and the FGF19 autocrine pathway may exist in the human livers. Bile acids and bile acid receptors are therapeutic targets for development of drugs for treatment of cholestatic liver diseases, fatty liver diseases, diabetes, obesity, and metabolic syndrome.

  4. Nutritional Signaling via Free Fatty Acid Receptors

    PubMed Central

    Miyamoto, Junki; Hasegawa, Sae; Kasubuchi, Mayu; Ichimura, Atsuhiko; Nakajima, Akira; Kimura, Ikuo

    2016-01-01

    Excess energy is stored primarily as triglycerides, which are mobilized when demand for energy arises. Dysfunction of energy balance by excess food intake leads to metabolic diseases, such as obesity and diabetes. Free fatty acids (FFAs) provided by dietary fat are not only important nutrients, but also contribute key physiological functions via FFA receptor (FFAR)-mediated signaling molecules, which depend on FFAs’ carbon chain length and the ligand specificity of the receptors. Functional analyses have revealed that FFARs are critical for metabolic functions, such as peptide hormone secretion and inflammation, and contribute to energy homeostasis. In particular, recent studies have shown that the administration of selective agonists of G protein-coupled receptor (GPR) 40 and GPR120 improved glucose metabolism and systemic metabolic disorders. Furthermore, the anti-inflammation and energy metabolism effects of short chain FAs have been linked to the activation of GPR41 and GPR43. In this review, we summarize recent progress in research on FFAs and their physiological roles in the regulation of energy metabolism. PMID:27023530

  5. Differential activation of pregnane X receptor by carnosic acid, carnosol, ursolic acid, and rosmarinic acid.

    PubMed

    Seow, Chun Ling; Lau, Aik Jiang

    2017-03-10

    Pregnane X receptor (PXR) regulates the expression of many genes, including those involved in drug metabolism and transport, and has been linked to various diseases, including inflammatory bowel disease. In the present study, we determined whether carnosic acid and other chemicals in rosemary extract (carnosol, ursolic acid, and rosmarinic acid) are PXR activators. As assessed in dual-luciferase reporter gene assays, carnosic acid, carnosol, and ursolic acid, but not rosmarinic acid, activated human PXR (hPXR) and mouse PXR (mPXR), whereas carnosol and ursolic acid, but not carnosic acid or rosmarinic acid, activated rat PXR (rPXR). Dose-response experiments indicated that carnosic acid, carnosol, and ursolic acid activated hPXR with EC50 values of 0.79, 2.22, and 10.77μM, respectively. Carnosic acid, carnosol, and ursolic acid, but not rosmarinic acid, transactivated the ligand-binding domain of hPXR and recruited steroid receptor coactivator-1 (SRC-1), SRC-2, and SRC-3 to the ligand-binding domain of hPXR. Carnosic acid, carnosol, and ursolic acid, but not rosmarinic acid, increased hPXR target gene expression, as shown by an increase in CYP3A4, UGT1A3, and ABCB1 mRNA expression in LS180 human colon adenocarcinoma cells. Rosmarinic acid did not attenuate the extent of hPXR activation by rifampicin, suggesting it is not an antagonist of hPXR. Overall, carnosic acid, carnosol, and ursolic acid, but not rosmarinic acid, are hPXR agonists, and carnosic acid shows species-dependent activation of hPXR and mPXR, but not rPXR. The findings provide new mechanistic insight on the effects of carnosic acid, carnosol, and ursolic acid on PXR-mediated biological effects.

  6. Transcriptional coordination of hepatic autophagy by nutrient-sensing nuclear receptor PPARα and FXR

    PubMed Central

    2016-01-01

    Nuclear receptors are in general ligand-dependent transcription factors that control a variety of mammalian physiologies including development, differentiation, proliferation, and homeostasis. Recent studies have found that two nutrient-sensing nuclear receptors, peroxisome proliferator-activated receptor α and farnesoid x receptor, responding to fasting or feeding state, respectively are able to regulate autophagy, an evolutionarily conserved catabolic process involved in lysosomal degradation. In this review, we discuss the role of these nutrient-sensing nuclear receptors in an aspect of transcriptional regulation of autophagy, and how these nuclear receptor-driven transcriptional programs integrate lipophagy, a lipid autophagy with fatty acid oxidation to coordinate hepatic lipid metabolism in the fasted state of the liver. PMID:28164071

  7. Transcriptional coordination of hepatic autophagy by nutrient-sensing nuclear receptor PPARα and FXR.

    PubMed

    Lee, Jae Man

    2016-12-01

    Nuclear receptors are in general ligand-dependent transcription factors that control a variety of mammalian physiologies including development, differentiation, proliferation, and homeostasis. Recent studies have found that two nutrient-sensing nuclear receptors, peroxisome proliferator-activated receptor α and farnesoid x receptor, responding to fasting or feeding state, respectively are able to regulate autophagy, an evolutionarily conserved catabolic process involved in lysosomal degradation. In this review, we discuss the role of these nutrient-sensing nuclear receptors in an aspect of transcriptional regulation of autophagy, and how these nuclear receptor-driven transcriptional programs integrate lipophagy, a lipid autophagy with fatty acid oxidation to coordinate hepatic lipid metabolism in the fasted state of the liver.

  8. Identifying the receptor subtype selectivity of retinoid X and retinoic acid receptors via quantum mechanics.

    PubMed

    Tsuji, Motonori; Shudo, Koichi; Kagechika, Hiroyuki

    2017-03-01

    Understanding and identifying the receptor subtype selectivity of a ligand is an important issue in the field of drug discovery. Using a combination of classical molecular mechanics and quantum mechanical calculations, this report assesses the receptor subtype selectivity for the human retinoid X receptor (hRXR) and retinoic acid receptor (hRAR) ligand-binding domains (LBDs) complexed with retinoid ligands. The calculated energies show good correlation with the experimentally reported binding affinities. The technique proposed here is a promising method as it reveals the origin of the receptor subtype selectivity of selective ligands.

  9. Pharmacophore model for bile acids recognition by the FPR receptor

    NASA Astrophysics Data System (ADS)

    Ferrari, Cristina; Macchiarulo, Antonio; Costantino, Gabriele; Pellicciari, Roberto

    2006-05-01

    Formyl-peptide receptors (FPRs) belong to the family A of the G-protein coupled receptor superfamily and include three subtypes: FPR, FPR-like-1 and FPR-like-2. They have been involved in the control of␣many inflammatory processes promoting the recruitment and infiltration of leukocytes in regions of inflammation through the molecular recognition of chemotactic factors. A large number of structurally diverse chemotypes modulate the activity of FPRs. Newly identified antagonists include bile acids deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA). The molecular recognition of these compounds at FPR receptor was computationally investigated using both ligand- and structure-based approaches. Our findings suggest that all antagonists bind at the first third of the seven helical bundles. A closer inspection of bile acid interaction reveals a number of unexploited anchor points in the binding site that may be used to aid the design of new potent and selective bile acids derivatives at FPR.

  10. Bile acids: emerging role in management of liver diseases

    PubMed Central

    Asgharpour, Amon; Kumar, Divya

    2016-01-01

    Bile acids are well known for their effects on cholesterol homeostasis and lipid digestion. Since the discovery of bile acid receptors, of which there are farnesoid X receptor (FXR), a nuclear receptor, and the plasma membrane G-protein receptor, as well as Takeda G-protein coupled receptor clone 5, further roles have been elucidated for bile acids including glucose and lipid metabolism as well as inflammation. Additionally, treatment with bile acid receptor agonists has shown a decrease in the amount of atherosclerosis plaque formation and decreased portal vascular resistance and portal hypotension in animal models. Furthermore, rodent models have demonstrated antifibrotic activity using bile acid receptor agonists. Early human data using a FXR agonist, obeticholic acid, have shown promising results with improvement of histological activity and even a reduction of fibrosis. Human studies are ongoing and will provide further information on bile acid receptor agonist therapies. Thus, bile acids and their derivatives have the potential for management of liver diseases and potentially other disease states including diabetes and the metabolic syndrome. PMID:26320013

  11. Receptor-level interrelationships of amino acids and the adequate amino acid type hormones in Tetrahymena: a receptor evolution model.

    PubMed

    Csaba, G; Darvas, Z

    1986-01-01

    Histidine stimulates the phagocytosis of Tetrahymena to the same extent as histamine, and also stimulates its division, which histamine does not. Tyrosine and diiodotyrosine equally stimulate the growth of the Tetrahymena. Both amino acids inhibit the characteristic influence of the adequate amino acid hormone when added to Tetrahymena culture 72 h in advance of it. Primary interaction with diiodotyrosine and tyrosine notably increases the cellular growth rate. Histamine has a similar, although less notable effect than histidine. In the light of these experimental observations there is reason to postulate that the receptors of the amino acid hormones have developed from amino acid receptors.

  12. Bile acid signaling in metabolic disease and drug therapy.

    PubMed

    Li, Tiangang; Chiang, John Y L

    2014-10-01

    Bile acids are the end products of cholesterol catabolism. Hepatic bile acid synthesis accounts for a major fraction of daily cholesterol turnover in humans. Biliary secretion of bile acids generates bile flow and facilitates hepatobiliary secretion of lipids, lipophilic metabolites, and xenobiotics. In the intestine, bile acids are essential for the absorption, transport, and metabolism of dietary fats and lipid-soluble vitamins. Extensive research in the last 2 decades has unveiled new functions of bile acids as signaling molecules and metabolic integrators. The bile acid-activated nuclear receptors farnesoid X receptor, pregnane X receptor, constitutive androstane receptor, vitamin D receptor, and G protein-coupled bile acid receptor play critical roles in the regulation of lipid, glucose, and energy metabolism, inflammation, and drug metabolism and detoxification. Bile acid synthesis exhibits a strong diurnal rhythm, which is entrained by fasting and refeeding as well as nutrient status and plays an important role for maintaining metabolic homeostasis. Recent research revealed an interaction of liver bile acids and gut microbiota in the regulation of liver metabolism. Circadian disturbance and altered gut microbiota contribute to the pathogenesis of liver diseases, inflammatory bowel diseases, nonalcoholic fatty liver disease, diabetes, and obesity. Bile acids and their derivatives are potential therapeutic agents for treating metabolic diseases of the liver.

  13. Obeticholic acid for the treatment of primary biliary cirrhosis.

    PubMed

    Trivedi, Palak J; Hirschfield, Gideon M; Gershwin, M Eric

    2016-01-01

    Primary biliary cirrhosis (PBC) is characterized by progressive nonsuppurative destruction of small bile ducts, resulting in intrahepatic cholestasis, fibrosis and ultimately end-stage liver disease. Timely intervention with ursodeoxycholic acid is associated with excellent survival, although approximately one-third of all patients fail to achieve biochemical response, signifying a critical need for additional therapeutic strategies. Obeticholic acid (OCA) is a potent ligand of the nuclear hormone receptor farnesoid X receptor (FXR). Activation of FXR inhibits bile acid synthesis and protects against toxic accumulation in models of cholestasis and facilitates hepatic regeneration in preclinical studies. Data from recent Phase II and III controlled trials suggest a therapeutic impact of OCA in PBC biochemical nonresponders, as evidenced by change in proven laboratory surrogates of long-term outcome. Dose-dependent pruritus is a common adverse effect, but may be overcome through dose-titration. Longer term studies are needed with focus on safety and long-term clinical efficacy.

  14. Bile Acid Signaling in Metabolic Disease and Drug Therapy

    PubMed Central

    Li, Tiangang

    2014-01-01

    Bile acids are the end products of cholesterol catabolism. Hepatic bile acid synthesis accounts for a major fraction of daily cholesterol turnover in humans. Biliary secretion of bile acids generates bile flow and facilitates hepatobiliary secretion of lipids, lipophilic metabolites, and xenobiotics. In the intestine, bile acids are essential for the absorption, transport, and metabolism of dietary fats and lipid-soluble vitamins. Extensive research in the last 2 decades has unveiled new functions of bile acids as signaling molecules and metabolic integrators. The bile acid–activated nuclear receptors farnesoid X receptor, pregnane X receptor, constitutive androstane receptor, vitamin D receptor, and G protein–coupled bile acid receptor play critical roles in the regulation of lipid, glucose, and energy metabolism, inflammation, and drug metabolism and detoxification. Bile acid synthesis exhibits a strong diurnal rhythm, which is entrained by fasting and refeeding as well as nutrient status and plays an important role for maintaining metabolic homeostasis. Recent research revealed an interaction of liver bile acids and gut microbiota in the regulation of liver metabolism. Circadian disturbance and altered gut microbiota contribute to the pathogenesis of liver diseases, inflammatory bowel diseases, nonalcoholic fatty liver disease, diabetes, and obesity. Bile acids and their derivatives are potential therapeutic agents for treating metabolic diseases of the liver. PMID:25073467

  15. Suppressed production of methyl farnesoid hormones yields developmental defects and lethality in Drosophila larvae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A long-unresolved question in the developmental biology of Drosophila melanogaster has been whether methyl farnesoid hormones secreted by the ring gland are necessary for larval maturation and metamorphosis. In this study, we have used RNAi techniques to inhibit 3-Hydroxy-3-Methylglutaryl CoA Reduct...

  16. Therapeutic targeting of bile acids

    PubMed Central

    Gores, Gregory J.

    2015-01-01

    The first objectives of this article are to review the structure, chemistry, and physiology of bile acids and the types of bile acid malabsorption observed in clinical practice. The second major theme addresses the classical or known properties of bile acids, such as the role of bile acid sequestration in the treatment of hyperlipidemia; the use of ursodeoxycholic acid in therapeutics, from traditional oriental medicine to being, until recently, the drug of choice in cholestatic liver diseases; and the potential for normalizing diverse bowel dysfunctions in irritable bowel syndrome, either by sequestering intraluminal bile acids for diarrhea or by delivering more bile acids to the colon to relieve constipation. The final objective addresses novel concepts and therapeutic opportunities such as the interaction of bile acids and the microbiome to control colonic infections, as in Clostridium difficile-associated colitis, and bile acid targeting of the farnesoid X receptor and G protein-coupled bile acid receptor 1 with consequent effects on energy expenditure, fat metabolism, and glycemic control. PMID:26138466

  17. Regulation of the cytosolic sulfotransferases by nuclear receptors

    PubMed Central

    Runge-Morris, Melissa; Kocarek, Thomas A.; Falany, Charles N.

    2013-01-01

    The cytosolic sulfotransferases (SULTs) are a multigene family of enzymes that catalyze the transfer of a sulfonate group from the physiologic sulfate donor, 3′-phosphoadenosine-5′-phosphosulfate, to a nucleophilic substrate to generate a polar product that is more amenable to elimination from the body. As catalysts of both xenobiotic and endogenous metabolism, the SULTs are major points of contact between the external and physiological environments, and modulation of SULT-catalyzed metabolism can not only affect xenobiotic disposition, but it can also alter endogenous metabolic processes. Therefore, it is not surprising that SULT expression is regulated by numerous members of the nuclear receptor (NR) superfamily that function as sensors of xenobiotics as well as endogenous molecules, such as fatty acids, bile acids, and oxysterols. These NRs include the peroxisome proliferator-activated receptors, pregnane X receptor, constitutive androstane receptor, vitamin D receptor, liver X receptors, farnesoid X receptor, retinoid-related orphan receptors, and estrogen-related receptors. This review summarizes current information about NR regulation of SULT expression. Because species differences in SULT subfamily composition and tissue-, sex-, development-, and inducer-dependent regulation are prominent, these differences will be emphasized throughout the review. In addition, because of the central role of the SULTs in cellular physiology, the effect of NR-mediated SULT regulation on physiological and pathophysiological processes will be discussed. Gaps in current knowledge that require further investigation are also highlighted. PMID:23330539

  18. Carbobenzoxy amino acids: Structural requirements for cholecystokinin receptor antagonist activity

    SciTech Connect

    Maton, P.N.; Sutliff, V.E.; Jensen, R.T.; Gardner, J.D.

    1985-04-01

    The authors used dispersed acini prepared from guinea pig pancreas to examine 28 carbobenzoxy (CBZ) amino acids for their abilities to function as cholecystokinin receptor antagonists. All amino acid derivatives tested, except for CBZ-alanine, CBZ-glycine, and N alpha-CBZ- lysine, were able to inhibit the stimulation of amylase secretion caused by the C-terminal octapeptide of cholecystokinin. In general, there was a good correlation between the ability of a carbobenzoxy amino acid to inhibit stimulated amylase secretion and the ability of the amino acid derivative to inhibit binding of /sup 125/I-cholecystokinin. The inhibition of cholecystokinin-stimulated amylase secretion was competitive, fully reversible, and specific for those secretagogues that interact with the cholecystokinin receptor. The potencies with which the various carbobenzoxy amino acids inhibited the action of cholecystokinin varied 100-fold and CBZ-cystine was the most potent cholecystokinin receptor antagonist. This variation in potency was primarily but not exclusively a function of the hydrophobicity of the amino acid side chain.

  19. Tauroursodeoxycholic acid protects bile acid homeostasis under inflammatory conditions and dampens Crohn's disease-like ileitis.

    PubMed

    Van den Bossche, Lien; Borsboom, Daniel; Devriese, Sarah; Van Welden, Sophie; Holvoet, Tom; Devisscher, Lindsey; Hindryckx, Pieter; De Vos, Martine; Laukens, Debby

    2017-02-06

    Bile acids regulate the expression of intestinal bile acid transporters and are natural ligands for nuclear receptors controlling inflammation. Accumulating evidence suggests that signaling through these receptors is impaired in inflammatory bowel disease. We investigated whether tauroursodeoxycholic acid (TUDCA), a secondary bile acid with cytoprotective properties, regulates ileal nuclear receptor and bile acid transporter expression and assessed its therapeutic potential in an experimental model of Crohn's disease (CD). Gene expression of the nuclear receptors farnesoid X receptor, pregnane X receptor and vitamin D receptor and the bile acid transporters apical sodium-dependent bile acid transporter and organic solute transporter α and β was analyzed in Caco-2 cell monolayers exposed to tumor necrosis factor (TNF)α, in ileal tissue of TNF(ΔARE/WT) mice and in inflamed ileal biopsies from CD patients by quantitative real-time polymerase chain reaction. TNF(ΔARE/WT) mice and wild-type littermates were treated with TUDCA or placebo for 11 weeks and ileal histopathology and expression of the aforementioned genes were determined. Exposing Caco-2 cell monolayers to TNFα impaired the mRNA expression of nuclear receptors and bile acid transporters, whereas co-incubation with TUDCA antagonized their downregulation. TNF(ΔARE/WT) mice displayed altered ileal bile acid homeostasis that mimicked the situation in human CD ileitis. Administration of TUDCA attenuated ileitis and alleviated the downregulation of nuclear receptors and bile acid transporters in these mice. These results show that TUDCA protects bile acid homeostasis under inflammatory conditions and suppresses CD-like ileitis. Together with previous observations showing similar efficacy in experimental colitis, we conclude that TUDCA could be a promising therapeutic agent for inflammatory bowel disease, warranting a clinical trial.Laboratory Investigation advance online publication, 6 February 2017; doi:10

  20. The Contributing Role of Bile Acids to Metabolic Improvements After Obesity and Metabolic Surgery.

    PubMed

    Fouladi, Farnaz; Mitchell, James E; Wonderlich, Joseph A; Steffen, Kristine J

    2016-10-01

    Obesity and metabolic surgery (OMS) leads to several metabolic improvements, which often occur prior to substantial weight loss. Therefore, other factors in addition to weight loss contribute to the metabolic benefits. This literature review offers an overview of studies investigating bile acids (BAs) and their metabolic effects after OMS. Rearrangement of enterohepatic circulation, changes in BA synthesis, BA conjugation, intestinal reabsorption, and alterations in the gut microbiota are potential mechanisms for altered BA profiles after surgery. Increased BA levels are associated with improved glucose homeostasis and lipid profiles, which are mediated by two major receptors: the Transmembrane G-protein Coupled Receptor and the Farnesoid X Receptor. Therefore, pharmacological manipulation of BAs and their receptors may be viable targets for less invasive obesity treatment.

  1. Selective Orthosteric Free Fatty Acid Receptor 2 (FFA2) Agonists

    PubMed Central

    Schmidt, Johannes; Smith, Nicola J.; Christiansen, Elisabeth; Tikhonova, Irina G.; Grundmann, Manuel; Hudson, Brian D.; Ward, Richard J.; Drewke, Christel; Milligan, Graeme; Kostenis, Evi; Ulven, Trond

    2011-01-01

    Free fatty acid receptor 2 (FFA2; GPR43) is a G protein-coupled seven-transmembrane receptor for short-chain fatty acids (SCFAs) that is implicated in inflammatory and metabolic disorders. The SCFA propionate has close to optimal ligand efficiency for FFA2 and can hence be considered as highly potent given its size. Propionate, however, does not discriminate between FFA2 and the closely related receptor FFA3 (GPR41). To identify FFA2-selective ligands and understand the molecular basis for FFA2 selectivity, a targeted library of small carboxylic acids was examined using holistic, label-free dynamic mass redistribution technology for primary screening and the receptor-proximal G protein [35S]guanosine 5′-(3-O-thio)triphosphate activation, inositol phosphate, and cAMP accumulation assays for hit confirmation. Structure-activity relationship analysis allowed formulation of a general rule to predict selectivity for small carboxylic acids at the orthosteric binding site where ligands with substituted sp3-hybridized α-carbons preferentially activate FFA3, whereas ligands with sp2- or sp-hybridized α-carbons prefer FFA2. The orthosteric binding mode was verified by site-directed mutagenesis: replacement of orthosteric site arginine residues by alanine in FFA2 prevented ligand binding, and molecular modeling predicted the detailed mode of binding. Based on this, selective mutation of three residues to their non-conserved counterparts in FFA3 was sufficient to transfer FFA3 selectivity to FFA2. Thus, selective activation of FFA2 via the orthosteric site is achievable with rather small ligands, a finding with significant implications for the rational design of therapeutic compounds selectively targeting the SCFA receptors. PMID:21220428

  2. Seizure control by decanoic acid through direct AMPA receptor inhibition

    PubMed Central

    Chang, Pishan; Augustin, Katrin; Boddum, Kim; Williams, Sophie; Sun, Min; Terschak, John A.; Hardege, Jörg D.; Chen, Philip E.

    2016-01-01

    See Rogawski (doi:10.1093/awv369) for a scientific commentary on this article.  The medium chain triglyceride ketogenic diet is an established treatment for drug-resistant epilepsy that increases plasma levels of decanoic acid and ketones. Recently, decanoic acid has been shown to provide seizure control in vivo, yet its mechanism of action remains unclear. Here we show that decanoic acid, but not the ketones β-hydroxybutryate or acetone, shows antiseizure activity in two acute ex vivo rat hippocampal slice models of epileptiform activity. To search for a mechanism of decanoic acid, we show it has a strong inhibitory effect on excitatory, but not inhibitory, neurotransmission in hippocampal slices. Using heterologous expression of excitatory ionotropic glutamate receptor AMPA subunits in Xenopus oocytes, we show that this effect is through direct AMPA receptor inhibition, a target shared by a recently introduced epilepsy treatment perampanel. Decanoic acid acts as a non-competitive antagonist at therapeutically relevant concentrations, in a voltage- and subunit-dependent manner, and this is sufficient to explain its antiseizure effects. This inhibitory effect is likely to be caused by binding to sites on the M3 helix of the AMPA-GluA2 transmembrane domain; independent from the binding site of perampanel. Together our results indicate that the direct inhibition of excitatory neurotransmission by decanoic acid in the brain contributes to the anti-convulsant effect of the medium chain triglyceride ketogenic diet. PMID:26608744

  3. Complement receptor 1 is a sialic acid-independent erythrocyte receptor of Plasmodium falciparum.

    PubMed

    Spadafora, Carmenza; Awandare, Gordon A; Kopydlowski, Karen M; Czege, Jozsef; Moch, J Kathleen; Finberg, Robert W; Tsokos, George C; Stoute, José A

    2010-06-17

    Plasmodium falciparum is a highly lethal malaria parasite of humans. A major portion of its life cycle is dedicated to invading and multiplying inside erythrocytes. The molecular mechanisms of erythrocyte invasion are incompletely understood. P. falciparum depends heavily on sialic acid present on glycophorins to invade erythrocytes. However, a significant proportion of laboratory and field isolates are also able to invade erythrocytes in a sialic acid-independent manner. The identity of the erythrocyte sialic acid-independent receptor has been a mystery for decades. We report here that the complement receptor 1 (CR1) is a sialic acid-independent receptor for the invasion of erythrocytes by P. falciparum. We show that soluble CR1 (sCR1) as well as polyclonal and monoclonal antibodies against CR1 inhibit sialic acid-independent invasion in a variety of laboratory strains and wild isolates, and that merozoites interact directly with CR1 on the erythrocyte surface and with sCR1-coated microspheres. Also, the invasion of neuraminidase-treated erythrocytes correlates with the level of CR1 expression. Finally, both sialic acid-independent and dependent strains invade CR1 transgenic mouse erythrocytes preferentially over wild-type erythrocytes but invasion by the latter is more sensitive to neuraminidase. These results suggest that both sialic acid-dependent and independent strains interact with CR1 in the normal red cell during the invasion process. However, only sialic acid-independent strains can do so without the presence of glycophorin sialic acid. Our results close a longstanding and important gap in the understanding of the mechanism of erythrocyte invasion by P. falciparum that will eventually make possible the development of an effective blood stage vaccine.

  4. Emerging role of obeticholic acid in the management of nonalcoholic fatty liver disease

    PubMed Central

    Makri, Evangelia; Cholongitas, Evangelos; Tziomalos, Konstantinos

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is the commonest chronic liver disease and its prevalence is increasing driven by the pandemic of obesity and type 2 diabetes mellitus. NAFLD can progress to cirrhosis and is associated with increased risk for cardiovascular disease and hepatocellular cancer. Diet and exercise are limited by suboptimal long-term adherence in patients with NAFLD. On the other hand, current pharmacological treatment of NAFLD has limited efficacy and unfavorable safety profile. In this context, obeticholic acid (OCA), a selective agonist of the farnesoid X receptors, might represent a useful option in these patients. Preclinical studies suggest that OCA improves hepatic steatosis, inflammation and fibrosis. A proof-of-concept study and the randomized, placebo-controlled Farnesoid X Receptor Ligand Obeticholic Acid in non-alcoholic steatohepatitis Treatment (FLINT) trial also showed improvements in liver histology in patients with NAFLD who received OCA. Weight loss and reduction in blood pressure were also observed. However, the effects of OCA on insulin resistance are conflicting and the lipid profile is adversely affected by this agent. In addition, pruritus is frequently observed during treatment with OCA and might lead to treatment discontinuation. However, given the limitations of existing treatments for NAFLD, OCA might represent a useful therapeutic option in selected patients with NAFLD. PMID:27895393

  5. Hyodeoxycholic acid derivatives as liver X receptor α and G-protein-coupled bile acid receptor agonists

    PubMed Central

    De Marino, Simona; Carino, Adriana; Masullo, Dario; Finamore, Claudia; Marchianò, Silvia; Cipriani, Sabrina; Di Leva, Francesco Saverio; Catalanotti, Bruno; Novellino, Ettore; Limongelli, Vittorio; Fiorucci, Stefano; Zampella, Angela

    2017-01-01

    Bile acids are extensively investigated for their potential in the treatment of human disorders. The liver X receptors (LXRs), activated by oxysterols and by a secondary bile acid named hyodeoxycholic acid (HDCA), have been found essential in the regulation of lipid homeostasis in mammals. Unfortunately, LXRα activates lipogenic enzymes causing accumulation of lipid in the liver. In addition to LXRs, HDCA has been also shown to function as ligand for GPBAR1, a G protein coupled receptor for secondary bile acids whose activation represents a promising approach to liver steatosis. In the present study, we report a library of HDCA derivatives endowed with modulatory activity on the two receptors. The lead optimization of HDCA moiety was rationally driven by the structural information on the binding site of the two targets and results from pharmacological characterization allowed the identification of hyodeoxycholane derivatives with selective agonistic activity toward LXRα and GPBAR1 and notably to the identification of the first example of potent dual LXRα/GPBAR1 agonists. The new chemical entities might hold utility in the treatment of dyslipidemic disorders. PMID:28233865

  6. Hyodeoxycholic acid derivatives as liver X receptor α and G-protein-coupled bile acid receptor agonists

    NASA Astrophysics Data System (ADS)

    de Marino, Simona; Carino, Adriana; Masullo, Dario; Finamore, Claudia; Marchianò, Silvia; Cipriani, Sabrina; di Leva, Francesco Saverio; Catalanotti, Bruno; Novellino, Ettore; Limongelli, Vittorio; Fiorucci, Stefano; Zampella, Angela

    2017-02-01

    Bile acids are extensively investigated for their potential in the treatment of human disorders. The liver X receptors (LXRs), activated by oxysterols and by a secondary bile acid named hyodeoxycholic acid (HDCA), have been found essential in the regulation of lipid homeostasis in mammals. Unfortunately, LXRα activates lipogenic enzymes causing accumulation of lipid in the liver. In addition to LXRs, HDCA has been also shown to function as ligand for GPBAR1, a G protein coupled receptor for secondary bile acids whose activation represents a promising approach to liver steatosis. In the present study, we report a library of HDCA derivatives endowed with modulatory activity on the two receptors. The lead optimization of HDCA moiety was rationally driven by the structural information on the binding site of the two targets and results from pharmacological characterization allowed the identification of hyodeoxycholane derivatives with selective agonistic activity toward LXRα and GPBAR1 and notably to the identification of the first example of potent dual LXRα/GPBAR1 agonists. The new chemical entities might hold utility in the treatment of dyslipidemic disorders.

  7. Identification of COUP-TFII Orphan Nuclear Receptor as a Retinoic Acid-Activated Receptor

    SciTech Connect

    Kruse, Schoen W; Suino-Powell, Kelly; Zhou, X Edward; Kretschman, Jennifer E; Reynolds, Ross; Vonrhein, Clemens; Xu, Yong; Wang, Liliang; Tsai, Sophia Y; Tsai, Ming-Jer; Xu, H Eric

    2010-01-12

    The chicken ovalbumin upstream promoter-transcription factors (COUP-TFI and II) make up the most conserved subfamily of nuclear receptors that play key roles in angiogenesis, neuronal development, organogenesis, cell fate determination, and metabolic homeostasis. Although the biological functions of COUP-TFs have been studied extensively, little is known of their structural features or aspects of ligand regulation. Here we report the ligand-free 1.48 {angstrom} crystal structure of the human COUP-TFII ligand-binding domain. The structure reveals an autorepressed conformation of the receptor, where helix {alpha}10 is bent into the ligand-binding pocket and the activation function-2 helix is folded into the cofactor binding site, thus preventing the recruitment of coactivators. In contrast, in multiple cell lines, COUP-TFII exhibits constitutive transcriptional activity, which can be further potentiated by nuclear receptor coactivators. Mutations designed to disrupt cofactor binding, dimerization, and ligand binding, substantially reduce the COUP-TFII transcriptional activity. Importantly, retinoid acids are able to promote COUP-TFII to recruit coactivators and activate a COUP-TF reporter construct. Although the concentration needed is higher than the physiological levels of retinoic acids, these findings demonstrate that COUP-TFII is a ligand-regulated nuclear receptor, in which ligands activate the receptor by releasing it from the autorepressed conformation.

  8. Cholestenoic acids regulate motor neuron survival via liver X receptors

    PubMed Central

    Theofilopoulos, Spyridon; Griffiths, William J.; Crick, Peter J.; Yang, Shanzheng; Meljon, Anna; Ogundare, Michael; Kitambi, Satish Srinivas; Lockhart, Andrew; Tuschl, Karin; Clayton, Peter T.; Morris, Andrew A.; Martinez, Adelaida; Reddy, M. Ashwin; Martinuzzi, Andrea; Bassi, Maria T.; Honda, Akira; Mizuochi, Tatsuki; Kimura, Akihiko; Nittono, Hiroshi; De Michele, Giuseppe; Carbone, Rosa; Criscuolo, Chiara; Yau, Joyce L.; Seckl, Jonathan R.; Schüle, Rebecca; Schöls, Ludger; Sailer, Andreas W.; Kuhle, Jens; Fraidakis, Matthew J.; Gustafsson, Jan-Åke; Steffensen, Knut R.; Björkhem, Ingemar; Ernfors, Patrik; Sjövall, Jan; Arenas, Ernest; Wang, Yuqin

    2014-01-01

    Cholestenoic acids are formed as intermediates in metabolism of cholesterol to bile acids, and the biosynthetic enzymes that generate cholestenoic acids are expressed in the mammalian CNS. Here, we evaluated the cholestenoic acid profile of mammalian cerebrospinal fluid (CSF) and determined that specific cholestenoic acids activate the liver X receptors (LXRs), enhance islet-1 expression in zebrafish, and increase the number of oculomotor neurons in the developing mouse in vitro and in vivo. While 3β,7α-dihydroxycholest-5-en-26-oic acid (3β,7α-diHCA) promoted motor neuron survival in an LXR-dependent manner, 3β-hydroxy-7-oxocholest-5-en-26-oic acid (3βH,7O-CA) promoted maturation of precursors into islet-1+ cells. Unlike 3β,7α-diHCA and 3βH,7O-CA, 3β-hydroxycholest-5-en-26-oic acid (3β-HCA) caused motor neuron cell loss in mice. Mutations in CYP7B1 or CYP27A1, which encode enzymes involved in cholestenoic acid metabolism, result in different neurological diseases, hereditary spastic paresis type 5 (SPG5) and cerebrotendinous xanthomatosis (CTX), respectively. SPG5 is characterized by spastic paresis, and similar symptoms may occur in CTX. Analysis of CSF and plasma from patients with SPG5 revealed an excess of the toxic LXR ligand, 3β-HCA, while patients with CTX and SPG5 exhibited low levels of the survival-promoting LXR ligand 3β,7α-diHCA. Moreover, 3β,7α-diHCA prevented the loss of motor neurons induced by 3β-HCA in the developing mouse midbrain in vivo.Our results indicate that specific cholestenoic acids selectively work on motor neurons, via LXR, to regulate the balance between survival and death. PMID:25271621

  9. Beyond intestinal soap--bile acids in metabolic control.

    PubMed

    Kuipers, Folkert; Bloks, Vincent W; Groen, Albert K

    2014-08-01

    Over the past decade, it has become apparent that bile acids are involved in a host of activities beyond their classic functions in bile formation and fat absorption. The identification of the farnesoid X receptor (FXR) as a nuclear receptor directly activated by bile acids and the discovery that bile acids are also ligands for the membrane-bound, G-protein coupled bile acid receptor 1 (also known as TGR5) have opened new avenues of research. Both FXR and TGR5 regulate various elements of glucose, lipid and energy metabolism. Consequently, a picture has emerged of bile acids acting as modulators of (postprandial) metabolism. Therefore, strategies that interfere with either bile acid metabolism or signalling cascades mediated by bile acids may represent novel therapeutic approaches for metabolic diseases. Synthetic modulators of FXR have been designed and tested, primarily in animal models. Furthermore, the use of bile acid sequestrants to reduce plasma cholesterol levels has unexpected benefits. For example, treatment of patients with type 2 diabetes mellitus (T2DM) with sequestrants causes substantial reductions in plasma levels of glucose and HbA1c. This Review aims to provide an overview of the molecular mechanisms by which bile acids modulate glucose and energy metabolism, particularly focusing on the glucose-lowering actions of bile acid sequestrants in insulin resistant states and T2DM.

  10. Electrophysiological evidence for acidic, basic, and neutral amino acid olfactory receptor sites in the catfish

    PubMed Central

    1984-01-01

    Electrophysiological experiments indicate that olfactory receptors of the channel catfish, Ictalurus punctatus, contain different receptor sites for the acidic (A), basic (B), and neutral amino acids; further, at least two partially interacting neutral sites exist, one for the hydrophilic neutral amino acids containing short side chains (SCN), and the second for the hydrophobic amino acids containing long side chains (LCN). The extent of cross-adaptation was determined by comparing the electro-olfactogram (EOG) responses to 20 "test" amino acids during continuous bathing of the olfactory mucosa with water only (control) to those during each of the eight "adapting" amino acid regimes. Both the adapting and test amino acids were adjusted in concentrations to provide approximately equal response magnitudes in the unadapted state. Under all eight adapting regimes, the test EOG responses were reduced from those obtained in the unadapted state, but substantial quantitative differences resulted, depending upon the molecular structure of the adapting stimulus. Analyses of the patterns of EOG responses to the test stimuli identified and characterized the respective "transduction processes," a term used to describe membrane events initiated by a particular subset of amino acid stimuli that are intricately linked to the origin of the olfactory receptor potential. Only when the stimulus compounds interact with different transduction processes are the stimuli assumed to bind to different membrane "sites." Four relatively independent L-alpha-amino acid transduction processes (and thus at least four binding sites) identified in this report include: (a) the A process for aspartic and glutamic acids; (b) the B process for arginine and lysine; (c) the SCN process for glycine, alanine, serine, glutamine, and possibly cysteine; (d) the LCN process for methionine, ethionine, valine, norvaline, leucine, norleucine, glutamic acid-gamma-methyl ester, histidine, phenylalanine, and also

  11. Receptor for protons: First observations on Acid Sensing Ion Channels.

    PubMed

    Krishtal, Oleg

    2015-07-01

    The history of ASICs began in 1980 with unexpected observation. The concept of highly selective Na(+) current gated by specific receptors for protons was not easily accepted. It took 16 years to get these receptor/channels cloned and start a new stage in their investigation. "The receptor for protons" became ASIC comprising under this name a family of receptor/channels ubiquitous for mammalian nervous system, both peripheral and central. The role of ASICs as putative nociceptors was suggested almost immediately after their discovery. This role subsequently was proven in many forms of pain-related phenomena. Many other functions of ASICs have been also found or primed for speculations both in physiology and in disease. Despite the width of field and strength of efforts, numerous basic questions are to be answered before we understand how the local changes in pH in the nervous tissue transform into electric and messenger signaling via ASICs as transducers. This article is part of the Special Issue entitled 'Acid-Sensing Ion Channels in the Nervous System'.

  12. Pentagastrin gastroprotection against acid is related to H2 receptor activation but not acid secretion

    PubMed Central

    Tanaka, S; Akiba, Y; Kaunitz, J

    1998-01-01

    Background—Pentagastrin enhances gastric mucosal defence mechanisms against acid and protects the gastric mucosa from experimental injury. 
Aims—To investigate whether this gastroprotection is mediated by histamine receptors or occurs as a secondary effect of acid secretion stimulation. 
Methods—The effects of omeprazole (100 µmol/kg), ranitidine (20 mg/kg), and pyrilamine (10 mg/kg) on pentagastrin (80 µg/kg/h) induced gastroprotection against acidified aspirin injury were examined in a luminal pH controlled model. The effects of these compounds on pentagastrin enhanced gastroprotective mechanisms were investigated using intravital microscopy, in which intracellular pH of gastric surface cells (pHi), mucus gel thickness, gastric mucosal blood flow, and acid output were measured simultaneously. 
Results—Pentagastrin protected rat gastric mucosa from acidified aspirin injury. This gastroprotection was abolished by ranitidine, but not omeprazole or pyrilamine. Pentagastrin induced a hyperaemic response to luminal acid challenge, increased mucus gel thickness, and elevated pHi during acid challenge. Ranitidine reversed these enhanced defence mechanisms, whereas omeprazole and pyrilamine preserved these effects. 
Conclusions—These data indicate that pentagastrin associated gastroprotection and enhanced defence mechanisms against acid result mainly from activation of histamine H2 receptors, and not as an effect of the stimulation of acid secretion. 

 Keywords: gastric injury; gastric defence mechanisms; omeprazole; pyrilamine; ranitidine; intracellular pH PMID:9863477

  13. Azadirachtin interacts with retinoic acid receptors and inhibits retinoic acid-mediated biological responses.

    PubMed

    Thoh, Maikho; Babajan, Banaganapalli; Raghavendra, Pongali B; Sureshkumar, Chitta; Manna, Sunil K

    2011-02-11

    Considering the role of retinoids in regulation of more than 500 genes involved in cell cycle and growth arrest, a detailed understanding of the mechanism and its regulation is useful for therapy. The extract of the medicinal plant Neem (Azadirachta indica) is used against several ailments especially for anti-inflammatory, anti-itching, spermicidal, anticancer, and insecticidal activities. In this report we prove the detailed mechanism on the regulation of retinoic acid-mediated cell signaling by azadirachtin, active components of neem extract. Azadirachtin repressed all trans-retinoic acid (ATRA)-mediated nuclear transcription factor κB (NF-κB) activation, not the DNA binding but the NF-κB-dependent gene expression. It did not inhibit IκBα degradation, IκBα kinase activity, or p65 phosphorylation and its nuclear translocation but inhibited NF-κB-dependent reporter gene expression. Azadirachtin inhibited TRAF6-mediated, but not TRAF2-mediated NF-κB activation. It inhibited ATRA-induced Sp1 and CREB (cAMP-response element-binding protein) DNA binding. Azadirachtin inhibited ATRA binding with retinoid receptors, which is supported by biochemical and in silico evidences. Azadirachtin showed strong interaction with retinoid receptors. It suppressed ATRA-mediated removal of retinoid receptors, bound with DNA by inhibiting ATRA binding to its receptors. Overall, our data suggest that azadirachtin interacts with retinoic acid receptors and suppresses ATRA binding, inhibits falling off the receptors, and activates transcription factors like CREB, Sp1, NF-κB, etc. Thus, azadirachtin exerts anti-inflammatory and anti-metastatic responses by a novel pathway that would be beneficial for further anti-inflammatory and anti-cancer therapies.

  14. Fatty acids and retinoids control lipid metabolism through activation of peroxisome proliferator-activated receptor-retinoid X receptor heterodimers.

    PubMed Central

    Keller, H; Dreyer, C; Medin, J; Mahfoudi, A; Ozato, K; Wahli, W

    1993-01-01

    The nuclear hormone receptors called PPARs (peroxisome proliferator-activated receptors alpha, beta, and gamma) regulate the peroxisomal beta-oxidation of fatty acids by induction of the acyl-CoA oxidase gene that encodes the rate-limiting enzyme of the pathway. Gel retardation and cotransfection assays revealed that PPAR alpha heterodimerizes with retinoid X receptor beta (RXR beta; RXR is the receptor for 9-cis-retinoic acid) and that the two receptors cooperate for the activation of the acyl-CoA oxidase gene promoter. The strongest stimulation of this promoter was obtained when both receptors were exposed simultaneously to their cognate activators. Furthermore, we show that natural fatty acids, and especially polyunsaturated fatty acids, activate PPARs as potently as does the hypolipidemic drug Wy 14,643, the most effective activator known so far. Moreover, we discovered that the synthetic arachidonic acid analogue 5,8,11,14-eicosatetraynoic acid is 100 times more effective than Wy 14,643 in the activation of PPAR alpha. In conclusion, our data demonstrate a convergence of the PPAR and RXR signaling pathways in the regulation of the peroxisomal beta-oxidation of fatty acids by fatty acids and retinoids. Images Fig. 1 Fig. 2 PMID:8384714

  15. Pharmacological activation of lysophosphatidic acid receptors regulates erythropoiesis

    PubMed Central

    Lin, Kuan-Hung; Ho, Ya-Hsuan; Chiang, Jui-Chung; Li, Meng-Wei; Lin, Shi-Hung; Chen, Wei-Min; Chiang, Chi-Ling; Lin, Yu-Nung; Yang, Ya-Jan; Chen, Chiung-Nien; Lu, Jenher; Huang, Chang-Jen; Tigyi, Gabor; Yao, Chao-Ling; Lee, Hsinyu

    2016-01-01

    Lysophosphatidic acid (LPA), a growth factor-like phospholipid, regulates numerous physiological functions, including cell proliferation and differentiation. In a previous study, we have demonstrated that LPA activates erythropoiesis by activating the LPA 3 receptor subtype (LPA3) under erythropoietin (EPO) induction. In the present study, we applied a pharmacological approach to further elucidate the functions of LPA receptors during red blood cell (RBC) differentiation. In K562 human erythroleukemia cells, knockdown of LPA2 enhanced erythropoiesis, whereas knockdown of LPA3 inhibited RBC differentiation. In CD34+ human hematopoietic stem cells (hHSC) and K526 cells, the LPA3 agonist 1-oleoyl-2-methyl-sn-glycero-3-phosphothionate (2S-OMPT) promoted erythropoiesis, whereas the LPA2 agonist dodecyl monophosphate (DMP) and the nonlipid specific agonist GRI977143 (GRI) suppressed this process. In zebrafish embryos, hemoglobin expression was significantly increased by 2S-OMPT treatment but was inhibited by GRI. Furthermore, GRI treatment decreased, whereas 2S-OMPT treatment increased RBC counts and amount of hemoglobin level in adult BALB/c mice. These results indicate that LPA2 and LPA3 play opposing roles during RBC differentiation. The pharmacological activation of LPA receptor subtypes represent a novel strategies for augmenting or inhibiting erythropoiesis. PMID:27244685

  16. Leveraging abscisic acid receptors for efficient water use in Arabidopsis

    PubMed Central

    Yang, Zhenyu; Liu, Jinghui; Tischer, Stefanie V.; Christmann, Alexander; Windisch, Wilhelm; Schnyder, Hans; Grill, Erwin

    2016-01-01

    Plant growth requires the influx of atmospheric CO2 through stomatal pores, and this carbon uptake for photosynthesis is inherently associated with a large efflux of water vapor. Under water deficit, plants reduce transpiration and are able to improve carbon for water exchange leading to higher water use efficiency (WUE). Whether increased WUE can be achieved without trade-offs in plant growth is debated. The signals mediating the WUE response under water deficit are not fully elucidated but involve the phytohormone abscisic acid (ABA). ABA is perceived by a family of related receptors known to mediate acclimation responses and to reduce transpiration. We now show that enhanced stimulation of ABA signaling via distinct ABA receptors can result in plants constitutively growing at high WUE in the model species Arabidopsis. WUE was assessed by three independent approaches involving gravimetric analyses, 13C discrimination studies of shoots and derived cellulose fractions, and by gas exchange measurements of whole plants and individual leaves. Plants expressing the ABA receptors RCAR6/PYL12 combined up to 40% increased WUE with high growth rates, i.e., are water productive. Water productivity was associated with maintenance of net carbon assimilation by compensatory increases of leaf CO2 gradients, thereby sustaining biomass acquisition. Leaf surface temperatures and growth potentials of plants growing under well-watered conditions were found to be reliable indicators for water productivity. The study shows that ABA receptors can be explored to generate more plant biomass per water transpired, which is a prime goal for a more sustainable water use in agriculture. PMID:27247417

  17. Positive and Negative Cross-Talk between Lysophosphatidic Acid Receptor 1, Free Fatty Acid Receptor 4, and Epidermal Growth Factor Receptor in Human Prostate Cancer Cells.

    PubMed

    Hopkins, Mandi M; Liu, Ze; Meier, Kathryn E

    2016-10-01

    Lysophosphatidic acid (LPA) is a lipid mediator that mediates cellular effects via G protein-coupled receptors (GPCRs). Epidermal growth factor (EGF) is a peptide that acts via a receptor tyrosine kinase. LPA and EGF both induce proliferation of prostate cancer cells and can transactivate each other's receptors. The LPA receptor LPA1 is particularly important for LPA response in human prostate cancer cells. Previous work in our laboratory has demonstrated that free fatty acid 4 (FFA4), a GPCR activated by ω-3 fatty acids, inhibits responses to both LPA and EGF in these cells. One potential mechanism for the inhibition involves negative interactions between FFA4 and LPA1, thereby suppressing responses to EGF that require LPA1 In the current study, we examined the role of LPA1 in mediating EGF and FFA4 agonist responses in two human prostate cancer cell lines, DU145 and PC-3. The results show that an LPA1-selective antagonist inhibits proliferation and migration to both LPA and EGF. Knockdown of LPA1 expression, using silencing RNA, blocks responses to LPA and significantly inhibits responses to EGF. The partial response to EGF that is observed after LPA1 knockdown is not inhibited by FFA4 agonists. Finally, the role of arrestin-3, a GPCR-binding protein that mediates many actions of activated GPCRs, was tested. Knockdown of arrestin-3 completely inhibits responses to both LPA and EGF in prostate cancer cells. Taken together, these results suggest that LPA1 plays a critical role in EGF responses and that FFA4 agonists inhibit proliferation by suppressing positive cross-talk between LPA1 and the EGF receptor.

  18. Retinal pigment epithelial acid lipase activity and lipoprotein receptors: effects of dietary omega-3 fatty acids.

    PubMed Central

    Elner, Victor M

    2002-01-01

    PURPOSE: To show that fish oil-derived omega-3 polyunsaturated fatty acids, delivered to the retinal pigment epithelium (RPE) by circulating low-density lipoproteins (LDL), enhance already considerable RPE lysosomal acid lipase activity, providing for more efficient hydrolysis of intralysosomal RPE lipids, an effect that may help prevent development of age-related macular degeneration (ARMD). METHODS: Colorimetric biochemical and histochemical techniques were used to demonstrate RPE acid lipase in situ, in vitro, and after challenge with phagocytic stimuli. Receptor-mediated RPE uptake of fluorescently labeled native, aceto-acetylated, and oxidized LDL was studied in vitro and in vivo. LDL effects on RPE lysosomal enzymes were assessed. Lysosomal enzyme activity was compared in RPE cells from monkeys fed diets rich in fish oil to those from control animals and in cultured RPE cells exposed to sera from these monkeys. RESULTS: RPE acid lipase activity was substantial and comparable to that of mononuclear phagocytes. Acid lipase activity increased significantly following phagocytic challenge with photoreceptor outer segment (POS) membranes. Receptor-mediated RPE uptake of labeled lipoproteins was determined in vitro. Distinctive uptake of labeled lipoproteins occurred in RPE cells and mononuclear phagocytes in vivo. Native LDL enhanced RPE lysosomal enzyme activity. RPE lysosomal enzymes increased significantly in RPE cells from monkeys fed fish oil-rich diets and in cultured RPE cells exposed to their sera. CONCLUSIONS: RPE cells contain substantial acid lipase for efficient metabolism of lipids imbibed by POS phagocytosis and LDL uptake. Diets rich in fish oil-derived omega-3 fatty acids, by enhancing acid lipase, may reduce RPE lipofuscin accumulation, RPE oxidative damage, and the development of ARMD. PMID:12545699

  19. Role of Nuclear Receptors in Lipid Dysfunction and Obesity-Related Diseases

    PubMed Central

    Wada, Taira; Xie, Wen; Renga, Barbara; Zampella, Angela; Distrutti, Eleonora; Fiorucci, Stefano; Kong, Bo; Thomas, Ann M.; Guo, Grace L.; Narayanan, Ramesh; Yepuru, Muralimohan; Dalton, James T.; Chiang, John Y. L.

    2013-01-01

    This article is a report on a symposium sponsored by the American Society for Pharmacology and Experimental Therapeutics and held at the Experimental Biology 12 meeting in San Diego, CA. The presentations discussed the roles of a number of nuclear receptors in regulating glucose and lipid homeostasis, the pathophysiology of obesity-related disease states, and the promise associated with targeting their activities to treat these diseases. While many of these receptors (in particular, constitutive androstane receptor and pregnane X receptor) and their target enzymes have been thought of as regulators of drug and xenobiotic metabolism, this symposium highlighted the advances made in our understanding of the endogenous functions of these receptors. Similarly, as we gain a better understanding of the mechanisms underlying bile acid signaling pathways in the regulation of body weight and glucose homeostasis, we see the importance of using complementary approaches to elucidate this fascinating network of pathways. The observation that some receptors, like the farnesoid X receptor, can function in a tissue-specific manner via well defined mechanisms has important clinical implications, particularly in the treatment of liver diseases. Finally, the novel findings that agents that selectively activate estrogen receptor β can effectively inhibit weight gain in a high-fat diet model of obesity identifies a new role for this member of the steroid superfamily. Taken together, the significant findings reported during this symposium illustrate the promise associated with targeting a number of nuclear receptors for the development of new therapies to treat obesity and other metabolic disorders. PMID:23043185

  20. Evolutionary and Functional Diversification of the Vitamin D Receptor-Lithocholic Acid Partnership.

    PubMed

    Kollitz, Erin M; Zhang, Guozhu; Hawkins, Mary Beth; Whitfield, G Kerr; Reif, David M; Kullman, Seth W

    2016-01-01

    The evolution, molecular behavior, and physiological function of nuclear receptors are of particular interest given their diverse roles in regulating essential biological processes. The vitamin D receptor (VDR) is well known for its canonical roles in calcium homeostasis and skeletal maintenance. Additionally, VDR has received an increased amount of attention due to the discovery of numerous non-calcemic functions, including the detoxification of lithocholic acid. Lithocholic acid is a toxic metabolite of chenodeoxycholic acid, a primary bile acid. The partnership between the VDR and lithocholic acid has been hypothesized to be a recent adaptation that evolved to mediate the detoxification and elimination of lithocholic acid from the gut. This partnership is speculated to be limited to higher vertebrates (birds and mammals), as lower vertebrates do not synthesize the parent compound of lithocholic acid. However, the molecular functions associated with the observed insensitivity of basal VDRs to lithocholic acid have not been explored. Here we characterize canonical nuclear receptor functions of VDRs from select species representing key nodes in vertebrate evolution and span a range of bile salt phenotypes. Competitive ligand binding assays revealed that the receptor's affinity for lithocholic acid is highly conserved across species, suggesting that lithocholic acid affinity is an ancient and non-adaptive trait. However, transient transactivation assays revealed that lithocholic acid-mediated VDR activation might have evolved more recently, as the non-mammalian receptors did not respond to lithocholic acid unless exogenous coactivator proteins were co-expressed. Subsequent functional assays indicated that differential lithocholic acid-mediated receptor activation is potentially driven by differential protein-protein interactions between VDR and nuclear receptor coregulator proteins. We hypothesize that the vitamin D receptor-lithocholic acid partnership evolved as a

  1. Free Fatty Acid Receptors and Cancer: From Nutrition to Pharmacology.

    PubMed

    Hopkins, Mandi M; Meier, Kathryn E

    2017-01-01

    The effects of fatty acids on cancer cells have been studied for decades. The roles of dietary long-chain n-3 polyunsaturated fatty acids, and of microbiome-generated short-chain butyric acid, have been of particular interest over the years. However, the roles of free fatty acid receptors (FFARs) in mediating effects of fatty acids in tumor cells have only recently been examined. In reviewing the literature, the data obtained to date indicate that the long-chain FFARs (FFA1 and FFA4) play different roles than the short-chain FFARs (FFA2 and FFA3). Moreover, FFA1 and FFA4 can in some cases mediate opposing actions in the same cell type. Another conclusion is that different types of cancer cells respond differently to FFAR activation. Currently, the best-studied models are prostate, breast, and colon cancer. FFA1 and FFA4 agonists can inhibit proliferation and migration of prostate and breast cancer cells, but enhance growth of colon cancer cells. In contrast, FFA2 activation can in some cases inhibit proliferation of colon cancer cells. Although the available data are sometimes contradictory, there are several examples in which FFAR agonists inhibit proliferation of cancer cells. This is a unique response to GPCR activation that will benefit from a mechanistic explanation as the field progresses. The development of more selective FFAR agonists and antagonists, combined with gene knockout approaches, will be important for unraveling FFAR-mediated inhibitory effects. These inhibitory actions, mediated by druggable GPCRs, hold promise for cancer prevention and/or therapy.

  2. FATTY ACIDS MODULATE TOLL-LIKE RECEPTOR 4 ACTIVATION THROUGH REGULATION OF RECEPTOR DIMERIZATION AND RECRUITMENT INTO LIPID RAFTS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The saturated fatty acids acylated on Lipid A of lipopolysaccharide (LPS) or bacterial lipoproteins play critical roles in ligand recognition and receptor activation for Toll-like Receptor 4 (TLR4) and TLR2. The results from our previous studies (J Biol Chem 2003, 2004) demonstrated that saturated ...

  3. Chromosomal Integration of Retinoic Acid Response Elements Prevents Cooperative Transcriptional Activation by Retinoic Acid Receptor and Retinoid X Receptor

    PubMed Central

    Lefebvre, Bruno; Brand, Céline; Lefebvre, Philippe; Ozato, Keiko

    2002-01-01

    All-trans-retinoic acid receptors (RAR) and 9-cis-retinoic acid receptors (RXR) are nuclear receptors known to cooperatively activate transcription from retinoid-regulated promoters. By comparing the transactivating properties of RAR and RXR in P19 cells using either plasmid or chromosomal reporter genes containing the mRARβ2 gene promoter, we found contrasting patterns of transcriptional regulation in each setting. Cooperativity between RXR and RAR occurred at all times with transiently introduced promoters, but was restricted to a very early stage (<3 h) for chromosomal promoters. This time-dependent loss of cooperativity was specific for chromosomal templates containing two copies of a retinoid-responsive element (RARE) and was not influenced by the spacing between the two RAREs. This loss of cooperativity suggested a delayed acquisition of RAR full transcriptional competence because (i) cooperativity was maintained at RAR ligand subsaturating concentrations, (ii) overexpression of SRC-1 led to loss of cooperativity and even to strong repression of chromosomal templates activity, and (iii) loss of cooperativity was observed when additional cis-acting response elements were activated. Surprisingly, histone deacetylase inhibitors counteracted this loss of cooperativity by repressing partially RAR-mediated activation of chromosomal promoters. Loss of cooperativity was not correlated to local histone hyperacetylation or to alteration of constitutive RNA polymerase II (RNAP) loading at the promoter region. Unexpectedly, RNAP binding to transcribed regions was correlated to the RAR activation state as well as to acetylation levels of histones H3 and H4, suggesting that RAR acts at the mRARβ promoter by triggering the switch from an RNA elongation-incompetent RNAP form towards an RNA elongation-competent RNAP. PMID:11839811

  4. Biosynthesis, biological effects, and receptors of hydroxyeicosatetraenoic acids (HETEs) and oxoeicosatetraenoic acids (oxo-ETEs) derived from arachidonic acid.

    PubMed

    Powell, William S; Rokach, Joshua

    2015-04-01

    Arachidonic acid can be oxygenated by a variety of different enzymes, including lipoxygenases, cyclooxygenases, and cytochrome P450s, and can be converted to a complex mixture of oxygenated products as a result of lipid peroxidation. The initial products in these reactions are hydroperoxyeicosatetraenoic acids (HpETEs) and hydroxyeicosatetraenoic acids (HETEs). Oxoeicosatetraenoic acids (oxo-ETEs) can be formed by the actions of various dehydrogenases on HETEs or by dehydration of HpETEs. Although a large number of different HETEs and oxo-ETEs have been identified, this review will focus principally on 5-oxo-ETE, 5S-HETE, 12S-HETE, and 15S-HETE. Other related arachidonic acid metabolites will also be discussed in less detail. 5-Oxo-ETE is synthesized by oxidation of the 5-lipoxygenase product 5S-HETE by the selective enzyme, 5-hydroxyeicosanoid dehydrogenase. It actions are mediated by the selective OXE receptor, which is highly expressed on eosinophils, suggesting that it may be important in eosinophilic diseases such as asthma. 5-Oxo-ETE also appears to stimulate tumor cell proliferation and may also be involved in cancer. Highly selective and potent OXE receptor antagonists have recently become available and could help to clarify its pathophysiological role. The 12-lipoxygenase product 12S-HETE acts by the GPR31 receptor and promotes tumor cell proliferation and metastasis and could therefore be a promising target in cancer therapy. It may also be involved as a proinflammatory mediator in diabetes. In contrast, 15S-HETE may have a protective effect in cancer. In addition to GPCRs, higher concentration of HETEs and oxo-ETEs can activate peroxisome proliferator-activated receptors (PPARs) and could potentially regulate a variety of processes by this mechanism. This article is part of a Special Issue entitled "Oxygenated metabolism of PUFA: analysis and biological relevance".

  5. Unsaturated phosphinic analogues of gamma-aminobutyric acid as GABA(C) receptor antagonists.

    PubMed

    Chebib, M; Vandenberg, R J; Froestl, W; Johnston, G A

    1997-06-25

    The phosphinic and methylphosphinic analogues of gamma-aminobutyric acid (GABA) are potent GABA(C) receptor antagonists but are even more potent as GABA(B) receptor agonists. Conformationally restricted unsaturated phosphinic and methylphosphinic analogues of GABA and some potent GABA(B) receptor phosphonoamino acid antagonists were tested on GABA(C) receptors in Xenopus oocytes expressing human retinal rho1 mRNA. 3-Aminopropyl-n-butyl-phosphinic acid (CGP36742), an orally active GABA(B) receptor antagonist, was found to be a moderately potent GABA(C) receptor antagonist (IC50 = 62 microM). The unsaturated methylphosphinic and phosphinic analogues of GABA were competitive antagonists of the GABA(C) receptors, the order of potency being [(E)-3-aminopropen-1-yl]methylphosphinic acid (CGP44530, IC50 = 5.53 microM) > [(E)-3-aminopropen-1-yl]phosphinic acid (CGP38593, IC50 = 7.68 microM) > [(Z)-3-aminopropen-1-yl]methylphosphinic acid (CGP70523, IC50 = 38.94 microM) > [(Z)-3-aminopropen-1-yl]phosphinic acid (CGP70522, IC50 > 100 microM). This order of potency differs from that reported for these compounds as GABA(B) receptor agonists, where the phosphinic acids are more potent than the corresponding methylphosphinic acids.

  6. Evolutionary and Functional Diversification of the Vitamin D Receptor-Lithocholic Acid Partnership

    PubMed Central

    Zhang, Guozhu; Hawkins, Mary Beth; Whitfield, G. Kerr; Reif, David M.; Kullman, Seth W.

    2016-01-01

    The evolution, molecular behavior, and physiological function of nuclear receptors are of particular interest given their diverse roles in regulating essential biological processes. The vitamin D receptor (VDR) is well known for its canonical roles in calcium homeostasis and skeletal maintenance. Additionally, VDR has received an increased amount of attention due to the discovery of numerous non-calcemic functions, including the detoxification of lithocholic acid. Lithocholic acid is a toxic metabolite of chenodeoxycholic acid, a primary bile acid. The partnership between the VDR and lithocholic acid has been hypothesized to be a recent adaptation that evolved to mediate the detoxification and elimination of lithocholic acid from the gut. This partnership is speculated to be limited to higher vertebrates (birds and mammals), as lower vertebrates do not synthesize the parent compound of lithocholic acid. However, the molecular functions associated with the observed insensitivity of basal VDRs to lithocholic acid have not been explored. Here we characterize canonical nuclear receptor functions of VDRs from select species representing key nodes in vertebrate evolution and span a range of bile salt phenotypes. Competitive ligand binding assays revealed that the receptor’s affinity for lithocholic acid is highly conserved across species, suggesting that lithocholic acid affinity is an ancient and non-adaptive trait. However, transient transactivation assays revealed that lithocholic acid-mediated VDR activation might have evolved more recently, as the non-mammalian receptors did not respond to lithocholic acid unless exogenous coactivator proteins were co-expressed. Subsequent functional assays indicated that differential lithocholic acid-mediated receptor activation is potentially driven by differential protein-protein interactions between VDR and nuclear receptor coregulator proteins. We hypothesize that the vitamin D receptor-lithocholic acid partnership evolved as

  7. Peroxisome proliferator-activated receptor-gamma coactivator-1alpha activation of CYP7A1 during food restriction and diabetes is still inhibited by small heterodimer partner.

    PubMed

    Shin, Dong-Ju; Osborne, Timothy F

    2008-05-30

    Cholesterol 7alpha-hydroxylase (CYP7A1) catalyzes the rate-limiting step in the classic pathway of hepatic bile acid biosynthesis from cholesterol. During fasting and in type I diabetes, elevated levels of peroxisome proliferator-activated receptor gamma-coactivator-1alpha (PGC-1alpha) induce expression of the Cyp7A1 gene and overexpression of PGC-1alpha in hepatoma cells stimulates bile acid synthesis. Using Ad-PGC-1alpha-RNA interference to induce acute disruption of PGC-1alpha in mice, here we show that PGC-1alpha is necessary for fasting-mediated induction of CYP7A1. Co-immunoprecipitation and promoter activation studies reveal that the induction of CYP7A1 is mediated by direct interaction between PGC-1alpha and the AF2 domain of liver receptor homolog-1 (LRH-1). In contrast, the very similar PGC-1beta could not substitute for PGC-1alpha. We also show that transactivation of PGC-1alpha and LRH-1 is repressed by the small heterodimer partner (SHP). Treatment of mice with GW4064, a synthetic agonist for farnesoid X receptor, induced SHP expression and decreased both the recruitment of PGC-1alpha to the Cyp7A1 promoter and the fasting-induced expression of CYP7A1 mRNA. These data suggest that PGC-1alpha is an important co-activator for LRH-1 and that SHP targets the interaction between LRH-1 and PGC-1alpha to inhibit CYP7A1 expression. Overall, these studies provide further evidence for the important role of PGC-1alpha in bile acid homeostasis and suggest that pharmacological targeting of farnesoid X receptor in vivo can be used to reverse the increase in CYP7A1 associated with adverse metabolic conditions.

  8. Structural Conservation of Ligand Binding Reveals a Bile Acid-like Signaling Pathway in Nematodes*

    PubMed Central

    Zhi, Xiaoyong; Zhou, X. Edward; Melcher, Karsten; Motola, Daniel L.; Gelmedin, Verena; Hawdon, John; Kliewer, Steven A.; Mangelsdorf, David J.; Xu, H. Eric

    2012-01-01

    Bile acid-like molecules named dafachronic acids (DAs) control the dauer formation program in Caenorhabditis elegans through the nuclear receptor DAF-12. This mechanism is conserved in parasitic nematodes to regulate their dauer-like infective larval stage, and as such, the DAF-12 ligand binding domain has been identified as an important therapeutic target in human parasitic hookworm species that infect more than 600 million people worldwide. Here, we report two x-ray crystal structures of the hookworm Ancylostoma ceylanicum DAF-12 ligand binding domain in complex with DA and cholestenoic acid (a bile acid-like metabolite), respectively. Structure analysis and functional studies reveal key residues responsible for species-specific ligand responses of DAF-12. Furthermore, DA binds to DAF-12 mechanistically and is structurally similar to bile acids binding to the mammalian bile acid receptor farnesoid X receptor. Activation of DAF-12 by cholestenoic acid and the cholestenoic acid complex structure suggest that bile acid-like signaling pathways have been conserved in nematodes and mammals. Together, these results reveal the molecular mechanism for the interplay between parasite and host, provide a structural framework for DAF-12 as a promising target in treating nematode parasitism, and provide insight into the evolution of gut parasite hormone-signaling pathways. PMID:22170062

  9. Bile Acids, FXR, and Metabolic Effects of Bariatric Surgery

    PubMed Central

    Noel, Olivier F.; Still, Christopher D.; Argyropoulos, George; Edwards, Michael; Gerhard, Glenn S.

    2016-01-01

    Overweight and obesity represent major risk factors for diabetes and related metabolic diseases. Obesity is associated with a chronic and progressive inflammatory response leading to the development of insulin resistance and type 2 diabetes (T2D) mellitus, although the precise mechanism mediating this inflammatory process remains poorly understood. The most effective intervention for the treatment of obesity, bariatric surgery, leads to glucose normalization and remission of T2D. Recent work in both clinical studies and animal models supports bile acids (BAs) as key mediators of these effects. BAs are involved in lipid and glucose homeostasis primarily via the farnesoid X receptor (FXR) transcription factor. BAs are also involved in regulating genes involved in inflammation, obesity, and lipid metabolism. Here, we review the novel role of BAs in bariatric surgery and the intersection between BAs and immune, obesity, weight loss, and lipid metabolism genes. PMID:27006824

  10. Enhancement of arachidonic acid signaling pathway by nicotinic acid receptor HM74A.

    PubMed

    Tang, Yuting; Zhou, Lubing; Gunnet, Joseph W; Wines, Pamela G; Cryan, Ellen V; Demarest, Keith T

    2006-06-23

    HM74A is a G protein-coupled receptor for nicotinic acid (niacin), which has been used clinically to treat dyslipidemia for decades. The molecular mechanisms whereby niacin exerts its pleiotropic effects on lipid metabolism remain largely unknown. In addition, the most common side effect in niacin therapy is skin flushing that is caused by prostaglandin release, suggesting that the phospholipase A(2) (PLA(2))/arachidonic acid (AA) pathway is involved. Various eicosanoids have been shown to activate peroxisome-proliferator activated receptors (PPAR) that play a diverse array of roles in lipid metabolism. To further elucidate the potential roles of HM74A in mediating the therapeutic effects and/or side effects of niacin, we sought to explore the signaling events upon HM74A activation. Here we demonstrated that HM74A synergistically enhanced UTP- and bradykinin-mediated AA release in a pertussis toxin-sensitive manner in A431 cells. Activation of HM74A also led to Ca(2+)-mobilization and enhanced bradykinin-promoted Ca(2+)-mobilization through Gi protein. While HM74A increased ERK1/2 activation by the bradykinin receptor, it had no effects on UTP-promoted ERK1/2 activation.Furthermore, UTP- and bradykinin-mediated AA release was significantly decreased in the presence of both MAPK kinase inhibitor PD 098059 and PKC inhibitor GF 109203X. However, the synergistic effects of HM74A were not dramatically affected by co-treatment with both inhibitors, indicating the cross-talk occurred at the receptor level. Finally, stimulation of A431 cells transiently transfected with PPRE-luciferase with AA significantly induced luciferase activity, mimicking the effects of PPARgamma agonist rosiglitazone, suggesting that alteration of AA signaling pathway can regulate gene expression via endogenous PPARs.

  11. Enhancement of arachidonic acid signaling pathway by nicotinic acid receptor HM74A

    SciTech Connect

    Tang, Yuting . E-mail: ytang@prdus.jnj.com; Zhou, Lubing; Gunnet, Joseph W.; Wines, Pamela G.; Cryan, Ellen V.; Demarest, Keith T.

    2006-06-23

    HM74A is a G protein-coupled receptor for nicotinic acid (niacin), which has been used clinically to treat dyslipidemia for decades. The molecular mechanisms whereby niacin exerts its pleiotropic effects on lipid metabolism remain largely unknown. In addition, the most common side effect in niacin therapy is skin flushing that is caused by prostaglandin release, suggesting that the phospholipase A{sub 2} (PLA{sub 2})/arachidonic acid (AA) pathway is involved. Various eicosanoids have been shown to activate peroxisome-proliferator activated receptors (PPAR) that play a diverse array of roles in lipid metabolism. To further elucidate the potential roles of HM74A in mediating the therapeutic effects and/or side effects of niacin, we sought to explore the signaling events upon HM74A activation. Here we demonstrated that HM74A synergistically enhanced UTP- and bradykinin-mediated AA release in a pertussis toxin-sensitive manner in A431 cells. Activation of HM74A also led to Ca{sup 2+}-mobilization and enhanced bradykinin-promoted Ca{sup 2+}-mobilization through Gi protein. While HM74A increased ERK1/2 activation by the bradykinin receptor, it had no effects on UTP-promoted ERK1/2 activation.Furthermore, UTP- and bradykinin-mediated AA release was significantly decreased in the presence of both MAPK kinase inhibitor PD 098059 and PKC inhibitor GF 109203X. However, the synergistic effects of HM74A were not dramatically affected by co-treatment with both inhibitors, indicating the cross-talk occurred at the receptor level. Finally, stimulation of A431 cells transiently transfected with PPRE-luciferase with AA significantly induced luciferase activity, mimicking the effects of PPAR{gamma} agonist rosiglitazone, suggesting that alteration of AA signaling pathway can regulate gene expression via endogenous PPARs.

  12. Nutrient-sensing nuclear receptors PPARα and FXR control liver energy balance.

    PubMed

    Preidis, Geoffrey A; Kim, Kang Ho; Moore, David D

    2017-04-03

    The nuclear receptors PPARα (encoded by NR1C1) and farnesoid X receptor (FXR, encoded by NR1H4) are activated in the liver in the fasted and fed state, respectively. PPARα activation induces fatty acid oxidation, while FXR controls bile acid homeostasis, but both nuclear receptors also regulate numerous other metabolic pathways relevant to liver energy balance. Here we review evidence that they function coordinately to control key nutrient pathways, including fatty acid oxidation and gluconeogenesis in the fasted state and lipogenesis and glycolysis in the fed state. We have also recently reported that these receptors have mutually antagonistic impacts on autophagy, which is induced by PPARα but suppressed by FXR. Secretion of multiple blood proteins is a major drain on liver energy and nutrient resources, and we present preliminary evidence that the liver secretome may be directly suppressed by PPARα, but induced by FXR. Finally, previous studies demonstrated a striking deficiency in bile acid levels in malnourished mice that is consistent with results in malnourished children. We present evidence that hepatic targets of PPARα and FXR are dysregulated in chronic undernutrition. We conclude that PPARα and FXR function coordinately to integrate liver energy balance.

  13. Lysophosphatidic Acid (LPA) Receptor 5 Inhibits B Cell Antigen Receptor Signaling and Antibody Response1

    PubMed Central

    Shotts, Kristin; Donovan, Erin E.; Strauch, Pamela; Pujanauski, Lindsey M.; Victorino, Francisco; Al-Shami, Amin; Fujiwara, Yuko; Tigyi, Gabor; Oravecz, Tamas; Pelanda, Roberta; Torres, Raul M.

    2014-01-01

    Lysophospholipids have emerged as biologically important chemoattractants capable of directing lymphocyte development, trafficking and localization. Lysophosphatidic acid (LPA) is a major lysophospholipid found systemically and whose levels are elevated in certain pathological settings such as cancer and infections. Here, we demonstrate that BCR signal transduction by mature murine B cells is inhibited upon LPA engagement of the LPA5 (GPR92) receptor via a Gα12/13 – Arhgef1 pathway. The inhibition of BCR signaling by LPA5 manifests by impaired intracellular calcium store release and most likely by interfering with inositol 1,4,5-trisphosphate receptor activity. We further show that LPA5 also limits antigen-specific induction of CD69 and CD86 expression and that LPA5-deficient B cells display enhanced antibody responses. Thus, these data show that LPA5 negatively regulates BCR signaling, B cell activation and immune response. Our findings extend the influence of lysophospholipids on immune function and suggest that alterations in LPA levels likely influence adaptive humoral immunity. PMID:24890721

  14. Bile acid sequestrants in type 2 diabetes: potential effects on GLP1 secretion.

    PubMed

    Sonne, David P; Hansen, Morten; Knop, Filip K

    2014-08-01

    Bile acid sequestrants have been used for decades for the treatment of hypercholesterolaemia. Sequestering of bile acids in the intestinal lumen interrupts enterohepatic recirculation of bile acids, which initiate feedback mechanisms on the conversion of cholesterol into bile acids in the liver, thereby lowering cholesterol concentrations in the circulation. In the early 1990s, it was observed that bile acid sequestrants improved glycaemic control in patients with type 2 diabetes. Subsequently, several studies confirmed the finding and recently - despite elusive mechanisms of action - bile acid sequestrants have been approved in the USA for the treatment of type 2 diabetes. Nowadays, bile acids are no longer labelled as simple detergents necessary for lipid digestion and absorption, but are increasingly recognised as metabolic regulators. They are potent hormones, work as signalling molecules on nuclear receptors and G protein-coupled receptors and trigger a myriad of signalling pathways in many target organs. The most described and well-known receptors activated by bile acids are the farnesoid X receptor (nuclear receptor) and the G protein-coupled cell membrane receptor TGR5. Besides controlling bile acid metabolism, these receptors are implicated in lipid, glucose and energy metabolism. Interestingly, activation of TGR5 on enteroendocrine L cells has been suggested to affect secretion of incretin hormones, particularly glucagon-like peptide 1 (GLP1 (GCG)). This review discusses the role of bile acid sequestrants in the treatment of type 2 diabetes, the possible mechanism of action and the role of bile acid-induced secretion of GLP1 via activation of TGR5.

  15. The ketogenic diet; fatty acids, fatty acid-activated receptors and neurological disorders.

    PubMed

    Cullingford, Tim E

    2004-03-01

    This review outlines the molecular sensors that reprogram cellular metabolism in response to the ketogenic diet (KD). Special emphasis is placed on the fasting-, fatty acid- and drug-activated transcription factor, peroxisome proliferator-activated receptor alpha (PPARalpha). The KD causes a switch to ketogenesis that is coordinated with an array of changes in cellular lipid, amino acid, carbohydrate and inflammatory pathways. The role of both liver and brain PPARalpha in mediating such changes will be examined, with special reference to the anti-epileptic effects not only of the KD but a range of synthetic anti-epileptic drugs such as valproate. Finally, the implications of the KD and activated brain PPARalpha will be discussed in the context of their potential involvement in a range of disorders of neuro-degeneration and neuro-inflammation.

  16. Characterization of bicuculline/baclofen-insensitive (rho-like) gamma-aminobutyric acid receptors expressed in Xenopus oocytes. II. Pharmacology of gamma-aminobutyric acidA and gamma-aminobutyric acidB receptor agonists and antagonists.

    PubMed

    Woodward, R M; Polenzani, L; Miledi, R

    1993-04-01

    Poly(A)+ RNA from mammalian retina expresses bicuculline/baclofen-insensitive gamma-aminobutyric acid (GABA) receptors in Xenopus oocytes with properties similar to those of homooligomeric GABA rho 1 receptors. The pharmacological profile of these rho-like receptors was extended by measuring sensitivities to various GABAA and GABAB receptor ligands. For direct comparison the same compounds were also assayed with GABAA receptors expressed by rat brain RNA. The potency sequence for heterocyclic GABA analogues at the GABA rho-like receptors was GABA (1.3) > muscimol (2.3) > isoguvacine (100) (approximate EC50 in parentheses; all EC50 and Kb values given in microM). Both muscimol and isoguvacine were partial agonists at the rho-like receptors. 4,5,6,7-Tetrahydroisoxazolo[5,4-c]pyridin-3-ol (Kb congruent to 32), piperidine-4-sulfonic acid (Kb congruent to 85), and isonipecotic acid (Kb congruent to 1000) acted primarily as competitive antagonists, showing little or no activity as agonists. The sulfonic acid GABA analogue 3-aminopropanesulfonic acid was also a competitive antagonist (Kb congruent to 20). Conformationally restricted GABA analogues trans- and cis-4-aminocrotonic acid (TACA and CACA) were agonists at the rho-like receptors. TACA (EC50 congruent to 0.6) had twice the potency of GABA and was 125 times more potent than CACA (EC50 congruent to 75). Z-3-(Amidinothio)propenoic acid, an isothiouronium analogue of GABA, had little activity as an agonist but instead acted as a competitive antagonist (Kb congruent to 20). At concentrations of > 100 microM, bicuculline did have some weak competitive inhibitory effects on the GABA rho-like receptors (Kb congruent to 6000), but it was at least 5000 times more potent at GABAA receptors. Strychnine (Kb congruent to 70) and SR-95531 (Kb congruent to 35) also were competitive inhibitors of the rho-like receptors but were, respectively, 20 and 240 times more potent at GABAA receptors. The GABAB receptor ligands baclofen

  17. Pharmacology of the inhibitory glycine receptor: agonist and antagonist actions of amino acids and piperidine carboxylic acid compounds.

    PubMed

    Schmieden, V; Betz, H

    1995-11-01

    To define structure-activity relations for ligands binding to the inhibitory glycine receptor (GlyR), the agonistic and antagonistic properties of alpha- and beta-amino acids were analyzed at the recombinant human alpha 1 GlyR expressed in Xenopus oocytes. The agonistic activity of alpha-amino acids exhibited a marked stereoselectivity and was highly susceptible to substitutions at the C alpha-atom. In contrast, alpha-amino acid antagonism was not enantiomer dependent and was influenced little by C alpha-atom substitutions. The beta-amino acids taurine, beta-aminobutyric acid (beta-ABA), and beta-aminoisobutyric acid (beta-AIBA) are partial agonists at the GlyR. Low concentrations of these compounds competitively inhibited glycine responses, whereas higher concentrations elicited a significant membrane current. Nipecotic acid, which contains a trans-beta-amino acid configuration, behaved as purely competitive GlyR antagonist. Our data are consistent with the existence of a common binding site for all amino acid agonists and antagonists, at which the functional consequences of binding depend on the particular conformation a given ligand adopts within the binding pocket. In the case of beta-amino acids, the trans conformation appears to mediate antagonistic receptor binding, and the cis conformation appears to mediate agonistic receptor binding. This led us to propose that the partial agonist activity of a given beta-amino acid is determined by the relative mole fractions of the respective cis/trans conformers.

  18. Molecular identification of high and low affinity receptors for nicotinic acid.

    PubMed

    Wise, Alan; Foord, Steven M; Fraser, Neil J; Barnes, Ashley A; Elshourbagy, Nabil; Eilert, Michelle; Ignar, Diane M; Murdock, Paul R; Steplewski, Klaudia; Green, Andrew; Brown, Andrew J; Dowell, Simon J; Szekeres, Philip G; Hassall, David G; Marshall, Fiona H; Wilson, Shelagh; Pike, Nicholas B

    2003-03-14

    Nicotinic acid has been used clinically for over 40 years in the treatment of dyslipidemia producing a desirable normalization of a range of cardiovascular risk factors, including a marked elevation of high density lipoprotein and a reduction in mortality. The precise mechanism of action of nicotinic acid is unknown, although it is believed that activation of a G(i)-G protein-coupled receptor may contribute. Utilizing available information on the tissue distribution of nicotinic acid receptors, we identified candidate orphan receptors. The selected orphan receptors were screened for responses to nicotinic acid, in an assay for activation of G(i)-G proteins. Here we describe the identification of the G protein-coupled receptor HM74 as a low affinity receptor for nicotinic acid. We then describe the subsequent identification of HM74A in follow-up bioinformatics searches and demonstrate that it acts as a high affinity receptor for nicotinic acid and other compounds with related pharmacology. The discovery of HM74A as a molecular target for nicotinic acid may facilitate the discovery of superior drug molecules to treat dyslipidemia.

  19. Structural basis and functions of abscisic acid receptors PYLs

    PubMed Central

    Zhang, Xing L.; Jiang, Lun; Xin, Qi; Liu, Yang; Tan, Jian X.; Chen, Zhong Z.

    2015-01-01

    Abscisic acid (ABA) plays a key role in many developmental processes and responses to adaptive stresses in plants. Recently, a new family of nucleocytoplasmic PYR/PYL/RCAR (PYLs) has been identified as bona fide ABA receptors. PYLs together with protein phosphatases type-2C (PP2Cs), Snf1 (Sucrose-non-fermentation 1)-related kinases subfamily 2 (SnRK2s) and downstream substrates constitute the core ABA signaling network. Generally, PP2Cs inactivate SnRK2s kinases by physical interaction and direct dephosphorylation. Upon ABA binding, PYLs change their conformations and then contact and inhibit PP2Cs, thus activating SnRK2s. Here, we reviewed the recent progress in research regarding the structures of the core signaling pathways of ABA, including the (+)-ABA, (−)-ABA and ABA analogs pyrabactin as well as 6AS perception by PYLs, SnRK2s mimicking PYLs in binding PP2Cs. PYLs inhibited PP2Cs in both the presence and absence of ABA and activated SnRK2s. The present review elucidates multiple ABA signal perception and transduction by PYLs, which might shed light on how to design small chemical compounds for improving plant performance in the future. PMID:25745428

  20. Altered food consumption in mice lacking lysophosphatidic acid receptor-1.

    PubMed

    Dusaulcy, R; Daviaud, D; Pradère, J P; Grès, S; Valet, Ph; Saulnier-Blache, J S

    2009-12-01

    The release of lysophosphatidic acid (LPA) by adipocytes has previously been proposed to play a role in obesity and associated pathologies such as insulin resistance and diabetes. In the present work, the sensitivity to diet-induced obesity was studied in mice lacking one of the LPA receptor subtype (LPA1R). Conversely to what was observed in wild type (WT) mice, LPA1R-KO-mice fed a high fat diet (HFD) showed no significant increase in body weight or fat mass when compared to low fat diet (LFD). In addition, in contrast to what was observed in WT mice, LPA1R-KO mice did not exhibit over-consumption of food associated with HFD. Surprisingly, when fed a LFD, LPA1R-KO mice exhibited significant higher plasma leptin concentration and higher level of adipocyte leptin mRNA than WT mice. In conclusion, LPA1R-KO mice were found to be resistant to diet-induced obesity consecutive to a resistance to fat-induced over-consumption of food that may result at least in part from alterations in leptin expression and production.

  1. In vivo blockade of thalamic GABA(B) receptors increases excitatory amino-acid levels.

    PubMed

    Nyitrai, G; Emri, Z; Crunelli, V; Kékesi, K A; Dobolyi, A; Juhász, G

    1996-12-30

    The effect of intrathalamic application of GABA(B) receptor antagonists on the basal excitatory amino-acid levels was studied using microdialysis probes implanted in the dorsal lateral geniculate nucleus and in the ventrobasal complex. In both nuclei, continuous perfusion of the GABA(B) receptor antagonist 3-aminopropyl-(diethoxymethyl)-phosphinic acid (CGP 35348) produced an increase in the extracellular concentration of aspartate and (to a lesser extent) glutamate, but no change was observed in the level of taurine, the main amino acid involved in the regulation of brain osmolarity processes. In contrast, 3-amino-2-hydroxy-2-(4-chlorophenyl)-propanesulphonic acid (2-hydroxy-saclofen), another GABA(B) receptor antagonist, failed to affect the extracellular concentration of aspartate, glutamate and taurine. Thus, the basal level of excitatory amino acids in the thalamus in vivo is under the control of CGP 35348-sensitive GABA(B) receptors.

  2. Conformationally constrained farnesoid X receptor (FXR) agonists: Heteroaryl replacements of the naphthalene

    SciTech Connect

    Bass, Jonathan Y.; Caravella, Justin A.; Chen, Lihong; Creech, Katrina L.; Deaton, David N.; Madauss, Kevin P.; Marr, Harry B.; McFadyen, Robert B.; Miller, Aaron B.; Mills, Wendy Y.; Navas, III, Frank; Parks, Derek J.; Smalley, Jr., Terrence L.; Spearing, Paul K.; Todd, Dan; Williams, Shawn P.; Wisely, G. Bruce

    2014-08-13

    To improve on the drug properties of GSK8062 1b, a series of heteroaryl bicyclic naphthalene replacements were prepared. The quinoline 1c was an equipotent FXR agonist with improved drug developability parameters relative to 1b. In addition, analog 1c lowered body weight gain and serum glucose in a DIO mouse model of diabetes.

  3. Med1 subunit of the mediator complex in nuclear receptor-regulated energy metabolism, liver regeneration, and hepatocarcinogenesis.

    PubMed

    Jia, Yuzhi; Viswakarma, Navin; Reddy, Janardan K

    2014-01-01

    Several nuclear receptors regulate diverse metabolic functions that impact on critical biological processes, such as development, differentiation, cellular regeneration, and neoplastic conversion. In the liver, some members of the nuclear receptor family, such as peroxisome proliferator-activated receptors (PPARs), constitutive androstane receptor (CAR), farnesoid X receptor (FXR), liver X receptor (LXR), pregnane X receptor (PXR), glucocorticoid receptor (GR), and others, regulate energy homeostasis, the formation and excretion of bile acids, and detoxification of xenobiotics. Excess energy burning resulting from increases in fatty acid oxidation systems in liver generates reactive oxygen species, and the resulting oxidative damage influences liver regeneration and liver tumor development. These nuclear receptors are important sensors of exogenous activators as well as receptor-specific endogenous ligands. In this regard, gene knockout mouse models revealed that some lipid-metabolizing enzymes generate PPARα-activating ligands, while others such as ACOX1 (fatty acyl-CoA oxidase1) inactivate these endogenous PPARα activators. In the absence of ACOX1, the unmetabolized ACOX1 substrates cause sustained activation of PPARα, and the resulting increase in energy burning leads to hepatocarcinogenesis. Ligand-activated nuclear receptors recruit the multisubunit Mediator complex for RNA polymerase II-dependent gene transcription. Evidence indicates that the Med1 subunit of the Mediator is essential for PPARα, PPARγ, CAR, and GR signaling in liver. Med1 null hepatocytes fail to respond to PPARα activators in that these cells do not show induction of peroxisome proliferation and increases in fatty acid oxidation enzymes. Med1-deficient hepatocytes show no increase in cell proliferation and do not give rise to liver tumors. Identification of nuclear receptor-specific coactivators and Mediator subunits should further our understanding of the complexities of metabolic

  4. Med1 Subunit of the Mediator Complex in Nuclear Receptor-Regulated Energy Metabolism, Liver Regeneration, and Hepatocarcinogenesis

    PubMed Central

    Jia, Yuzhi; Viswakarma, Navin; Reddy, Janardan K.

    2014-01-01

    Several nuclear receptors regulate diverse metabolic functions that impact on critical biological processes, such as development, differentiation, cellular regeneration, and neoplastic conversion. In the liver, some members of the nuclear receptor family, such as peroxisome proliferator-activated receptors (PPARs), constitutive androstane receptor (CAR), farnesoid X receptor (FXR), liver X receptor (LXR), pregnane X receptor (PXR), glucocorticoid receptor (GR), and others, regulate energy homeostasis, the formation and excretion of bile acids, and detoxification of xenobiotics. Excess energy burning resulting from increases in fatty acid oxidation systems in liver generates reactive oxygen species, and the resulting oxidative damage influences liver regeneration and liver tumor development. These nuclear receptors are important sensors of exogenous activators as well as receptor-specific endogenous ligands. In this regard, gene knockout mouse models revealed that some lipid-metabolizing enzymes generate PPARα-activating ligands, while others such as ACOX1 (fatty acyl-CoA oxidase1) inactivate these endogenous PPARα activators. In the absence of ACOX1, the unmetabolized ACOX1 substrates cause sustained activation of PPARα, and the resulting increase in energy burning leads to hepatocarcinogenesis. Ligand-activated nuclear receptors recruit the multisubunit Mediator complex for RNA polymerase II-dependent gene transcription. Evidence indicates that the Med1 subunit of the Mediator is essential for PPARα, PPARγ, CAR, and GR signaling in liver. Med1 null hepatocytes fail to respond to PPARα activators in that these cells do not show induction of peroxisome proliferation and increases in fatty acid oxidation enzymes. Med1-deficient hepatocytes show no increase in cell proliferation and do not give rise to liver tumors. Identification of nuclear receptor-specific coactivators and Mediator subunits should further our understanding of the complexities of metabolic

  5. Tannic acid, a potent inhibitor of epidermal growth factor receptor tyrosine kinase.

    PubMed

    Yang, Er Bin; Wei, Liu; Zhang, Kai; Chen, Yu Zong; Chen, Wei Ning

    2006-03-01

    Increasing evidence supports the hypothesis that tannic acid, a plant polyphenol, exerts anticarcinogenic activity in chemically induced cancers. In the present study, tannic acid was found to strongly inhibit tyrosine kinase activity of epidermal growth factor receptor (EGFr) in vitro (IC50 = 323 nM). In contrast, the inhibition by tannic acid of p60(c-src) tyrosine kinase (IC50 = 14 microM) and insulin receptor tyrosine kinase (IC50 = 5 microM) was much weaker. The inhibition of EGFr tyrosine kinase by tannic acid was competitive with respect to ATP and non-competitive with respect to peptide substrate. In cultured cells, growth factor-induced tyrosine phosphorylation of growth factor receptors, including EGFr, platelet-derived growth factor receptor, and basic fibroblast growth factor receptor, was inhibited by tannic acid. No inhibition of insulin-induced tyrosine phosphorylation of insulin receptor and insulin-receptor substrate-1 was observed. EGF-stimulated growth of HepG2 cells was inhibited in the presence of tannic acid. The inhibition of serine/threonine-specific protein kinases, including cAMP-dependent protein kinase, protein kinase C and mitogen-activated protein kinase, by tannic acid was only detected at relatively high concentration, IC50 being 3, 325 and 142 microM respectively. The molecular modeling study suggested that tannic acid could be docked into the ATP binding pockets of either EGFr or insulin receptor. These results demonstrate that tannic acid is an in vitro potent inhibitor of EGFr tyrosine kinase.

  6. Effects of beer and hop on ionotropic gamma-aminobutyric acid receptors.

    PubMed

    Aoshima, Hitoshi; Takeda, Katsuichi; Okita, Yoichi; Hossain, Sheikh Julfikar; Koda, Hirofumi; Kiso, Yoshinobu

    2006-04-05

    Beer induced the response of the ionotropic gamma-aminobutyric acid receptors (GABA(A) receptors) expressed in Xenopus oocytes, indicating the presence of gamma-aminobutyric acid (GABA)-like activity. Furthermore, the pentane extract of the beer, hop (Humulus lupulus L.) oil, and myrcenol potentiated the GABA(A) receptor response elicited by GABA. The GABA(A) receptor responses were also potentiated by the addition of aliphatic esters, most of which are reported to be present in beer flavor. Aliphatic esters showed the tendency to decrease in the potentiation of the GABA(A) receptor response with an increase in their carbon chain length. When myrcenol was injected to mice prior to intraperitoneal administration of pentobarbital, the pentobarbital-induced sleeping time of mice increased additionally. Therefore, the beer contained not only GABA-like activity but also the modulator(s) of the GABA(A) receptor response.

  7. OR-1, a member of the nuclear receptor superfamily that interacts with the 9-cis-retinoic acid receptor.

    PubMed Central

    Teboul, M; Enmark, E; Li, Q; Wikström, A C; Pelto-Huikko, M; Gustafsson, J A

    1995-01-01

    We have cloned a member of the nuclear receptor superfamily. The cDNA was isolated from a rat liver library and encodes a protein of 446 aa with a predicted mass of 50 kDa. This clone (OR-1) shows no striking homology to any known member of the steroid/thyroid hormone receptor superfamily. The most related receptor is the ecdysone receptor and the highest homologies represent < 10% in the amino-terminal domain, between 15-37% in the carboxyl-terminal domain and 50-62% in the DNA binding domain. The expression of OR-1 appears to be widespread in both fetal and adult rat tissues. Potential DNA response elements composed of a direct repeat of the hexameric motif AGGTCA spaced by 0-6 nt were tested in gel shift experiments. OR-1 was shown to interact with the 9-cis-retinoic acid receptor (retinoid X receptor, RXR) and the OR-1/RXR complex to bind to a direct repeat spaced by 4 nt (DR4). In transfection experiments, OR-1 appears to activate RXR-mediated function through the DR4. Therefore OR-1 might modulate 9-cis-retinoic acid signaling by interacting with RXR. Images Fig. 3 Fig. 4 PMID:7892230

  8. Potentiation of Gamma Aminobutyric Acid Receptors (GABAAR) by Ethanol: How Are Inhibitory Receptors Affected?

    PubMed Central

    Förstera, Benjamin; Castro, Patricio A.; Moraga-Cid, Gustavo; Aguayo, Luis G.

    2016-01-01

    In recent years there has been an increase in the understanding of ethanol actions on the type A γ-aminobutyric acid chloride channel (GABAAR), a member of the pentameric ligand gated ion channels (pLGICs). However, the mechanism by which ethanol potentiates the complex is still not fully understood and a number of publications have shown contradictory results. Thus many questions still remain unresolved requiring further studies for a better comprehension of this effect. The present review concentrates on the involvement of GABAAR in the acute actions of ethanol and specifically focuses on the immediate, direct or indirect, synaptic and extra-synaptic modulatory effects. To elaborate on the immediate, direct modulation of GABAAR by acute ethanol exposure, electrophysiological studies investigating the importance of different subunits, and data from receptor mutants will be examined. We will also discuss the nature of the putative binding sites for ethanol based on structural data obtained from other members of the pLGICs family. Finally, we will briefly highlight the glycine gated chloride channel (GlyR), another member of the pLGIC family, as a suitable target for the development of new pharmacological tools. PMID:27199667

  9. Data for amino acid alignment of Japanese stingray melanocortin receptors with other gnathostome melanocortin receptor sequences, and the ligand selectivity of Japanese stingray melanocortin receptors.

    PubMed

    Takahashi, Akiyoshi; Davis, Perry; Reinick, Christina; Mizusawa, Kanta; Sakamoto, Tatsuya; Dores, Robert M

    2016-06-01

    This article contains structure and pharmacological characteristics of melanocortin receptors (MCRs) related to research published in "Characterization of melanocortin receptors from stingray Dasyatis akajei, a cartilaginous fish" (Takahashi et al., 2016) [1]. The amino acid sequences of the stingray, D. akajei, MC1R, MC2R, MC3R, MC4R, and MC5R were aligned with the corresponding melanocortin receptor sequences from the elephant shark, Callorhinchus milii, the dogfish, Squalus acanthias, the goldfish, Carassius auratus, and the mouse, Mus musculus. These alignments provide the basis for phylogenetic analysis of these gnathostome melanocortin receptor sequences. In addition, the Japanese stingray melanocortin receptors were separately expressed in Chinese Hamster Ovary cells, and stimulated with stingray ACTH, α-MSH, β-MSH, γ-MSH, δ-MSH, and β-endorphin. The dose response curves reveal the order of ligand selectivity for each stingray MCR.

  10. Nicotinic Acid Receptor Abnormalities in Human Skin Cancer: Implications for a Role in Epidermal Differentiation

    PubMed Central

    Bermudez, Yira; Benavente, Claudia A.; Meyer, Ralph G.; Coyle, W. Russell; Jacobson, Myron K.; Jacobson, Elaine L.

    2011-01-01

    Background Chronic UV skin exposure leads to epidermal differentiation defects in humans that can be largely restored by pharmacological doses of nicotinic acid. Nicotinic acid has been identified as a ligand for the human G-protein-coupled receptors GPR109A and GPR109B that signal through Gi-mediated inhibition of adenylyl cyclase. We have examined the expression, cellular distribution, and functionality of GPR109A/B in human skin and skin derived epidermal cells. Results Nicotinic acid increases epidermal differentiation in photodamaged human skin as judged by the terminal differentiation markers caspase 14 and filaggrin. Both GPR109A and GPR109B genes are transcribed in human skin and in epidermal keratinocytes, but expression in dermal fibroblasts is below limits of detection. Receptor transcripts are greatly over-expressed in squamous cell cancers. Receptor protein in normal skin is prominent from the basal through granular layers of the epidermis, with cellular localization more dispersive in the basal layer but predominantly localized at the plasma membrane in more differentiated epidermal layers. In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional Gi-mediated signaling. In contrast, in a squamous cell carcinoma derived cell line, receptor protein shows a more diffuse cellular localization and the receptors are nearly non-functional. Conclusions The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis. PMID:21655214

  11. Unnatural agrochemical ligands for engineered abscisic acid receptors.

    PubMed

    Rodriguez, Pedro L; Lozano-Juste, Jorge

    2015-06-01

    Existing agrochemicals can be endowed with new applications through protein engineering of plant receptors. A recent study shows an engineered PYR1 ABA receptor can be activated by mandipropamid. Plants engineered with such PYR1 variant are responsive to this agrochemical, which confers protection against drought through activation of ABA signaling.

  12. Tranexamic acid induces kaolin intake stimulating a pathway involving tachykinin neurokinin 1 receptors in rats.

    PubMed

    Kakiuchi, Hitoshi; Kawarai-Shimamura, Asako; Kuwagata, Makiko; Orito, Kensuke

    2014-01-15

    Tranexamic acid suppresses post-partum haemorrhage and idiopathic menorrhagia through its anti-fibrinolytic action. Although it is clinically useful, it is associated with high risks of side effects such as emesis. Understanding the mechanisms underlying tranexamic acid-induced emesis is very important to explore appropriate anti-emetic drugs for the prevention and/or suppression of emesis. In this study, we examined the receptors involved in tranexamic acid-induced kaolin intake in rats, which reflects the drug's clinical emetogenic potential in humans. Further, we examined the brain regions activated by administration of tranexamic acid and elucidated pivotal pathways of tranexamic acid-induced kaolin intake. We examined the effects of ondansetron, a 5-hydroxytryptamine 3 receptor antagonist, domperidone, a dopamine 2 receptor antagonist, and aprepitant, a tachykinin neurokinin 1 (NK1) receptor antagonist, on tranexamic acid-induced kaolin intake in rats. Then, we determined the brain regions that showed increased numbers of c-Fos immunoreactive cells. Finally, we examined the effects of an antagonist(s) that reduced tranexamic acid-induced kaolin intake on the increase in c-Fos immunoreactive cells. Aprepitant significantly decreased tranexamic acid-induced kaolin intake. However, neither ondansetron nor domperidone decreased kaolin intake. Tranexamic acid significantly increased c-Fos immunoreactive cells by approximately 5.5-fold and 22-fold in the area postrema and nucleus of solitary tract, respectively. Aprepitant decreased the number of c-Fos immunoreactive cells in both areas. Tranexamic acid induced kaolin intake possibly via stimulation of tachykinin NK1 receptors in rats. The tachykinin NK1 receptor could be targeted to prevent and/or suppress emesis in patients receiving tranexamic acid.

  13. Castor oil induces laxation and uterus contraction via ricinoleic acid activating prostaglandin EP3 receptors.

    PubMed

    Tunaru, Sorin; Althoff, Till F; Nüsing, Rolf M; Diener, Martin; Offermanns, Stefan

    2012-06-05

    Castor oil is one of the oldest drugs. When given orally, it has a laxative effect and induces labor in pregnant females. The effects of castor oil are mediated by ricinoleic acid, a hydroxylated fatty acid released from castor oil by intestinal lipases. Despite the wide-spread use of castor oil in conventional and folk medicine, the molecular mechanism by which ricinoleic acid acts remains unknown. Here we show that the EP(3) prostanoid receptor is specifically activated by ricinoleic acid and that it mediates the pharmacological effects of castor oil. In mice lacking EP(3) receptors, the laxative effect and the uterus contraction induced via ricinoleic acid are absent. Although a conditional deletion of the EP(3) receptor gene in intestinal epithelial cells did not affect castor oil-induced diarrhea, mice lacking EP(3) receptors only in smooth-muscle cells were unresponsive to this drug. Thus, the castor oil metabolite ricinoleic acid activates intestinal and uterine smooth-muscle cells via EP(3) prostanoid receptors. These findings identify the cellular and molecular mechanism underlying the pharmacological effects of castor oil and indicate a role of the EP(3) receptor as a target to induce laxative effects.

  14. Coexpression of striatal dopamine receptor subtypes and excitatory amino acid subunits.

    PubMed

    Ariano, M A; Larson, E R; Noblett, K L; Sibley, D R; Levine, M S

    1997-08-01

    The striatal cellular coexpression patterns for the D(1A) and D2 dopamine (DA) receptor subtypes and the ionotropic excitatory amino acid (EAA) subunits of the N-methyl-D-aspartate (NMDA-R1) and the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) (GluR1 and GluR2/3) receptor subunits were examined morphologically. Their coincidence was assessed by visualization of mRNA transcripts, localization of encoded receptor proteins, and binding analysis using concurrently paired methods of fluorescence detection. The findings indicated that 1) mRNA transcripts for both receptor systems were detected in the medium-sized neuron population, and the distribution of receptor message closely reflected protein and binding patterns, with the exception of the GluR1 subunit; 2) both DA receptor mRNA transcripts were coexpressed with each ionotropic EAA receptor subunit examined and with each other, and NMDA and AMPA receptor subunits also showed coincident expression; 3) D(1A) DA receptor protein was detected in neurons which coexpressed EAA subunit proteins; and 4) GluR2/3 and NMDA-R1 subunit proteins were coexpressed in medium-sized neurons which also demonstrated D2 DA receptor binding sites. These findings suggest morphological receptor "promiscuity" since the coexpression patterns between DA and EAA receptors were found in all permutations. The results provide a spatial framework for physiological findings describing functional interactions between the two DA receptor types and between specific DA and EAA receptors in the striatum.

  15. Urinary metabolomics in Fxr-null mice reveals activated adaptive metabolic pathways upon bile acid challenge.

    PubMed

    Cho, Joo-Youn; Matsubara, Tsutomu; Kang, Dong Wook; Ahn, Sung-Hoon; Krausz, Kristopher W; Idle, Jeffrey R; Luecke, Hans; Gonzalez, Frank J

    2010-05-01

    Farnesoid X receptor (FXR) is a nuclear receptor that regulates genes involved in synthesis, metabolism, and transport of bile acids and thus plays a major role in maintaining bile acid homeostasis. In this study, metabolomic responses were investigated in urine of wild-type and Fxr-null mice fed cholic acid, an FXR ligand, using ultra-performance liquid chromatography (UPLC) coupled with electrospray time-of-flight mass spectrometry (TOFMS). Multivariate data analysis between wild-type and Fxr-null mice on a cholic acid diet revealed that the most increased ions were metabolites of p-cresol (4-methylphenol), corticosterone, and cholic acid in Fxr-null mice. The structural identities of the above metabolites were confirmed by chemical synthesis and by comparing retention time (RT) and/or tandem mass fragmentation patterns of the urinary metabolites with the authentic standards. Tauro-3alpha,6,7alpha,12alpha-tetrol (3alpha,6,7alpha,12alpha-tetrahydroxy-5beta-cholestan-26-oyltaurine), one of the most increased metabolites in Fxr-null mice on a CA diet, is a marker for efficient hydroxylation of toxic bile acids possibly through induction of Cyp3a11. A cholestatic model induced by lithocholic acid revealed that enhanced expression of Cyp3a11 is the major defense mechanism to detoxify cholestatic bile acids in Fxr-null mice. These results will be useful for identification of biomarkers for cholestasis and for determination of adaptive molecular mechanisms in cholestasis.

  16. Amino Acid- vs. Peptide-Odorants: Responses of Individual Olfactory Receptor Neurons in an Aquatic Species

    PubMed Central

    Hassenklöver, Thomas; Pallesen, Lars P.; Schild, Detlev; Manzini, Ivan

    2012-01-01

    Amino acids are widely used waterborne olfactory stimuli proposed to serve as cues in the search for food. In natural waters the main source of amino acids is the decomposition of proteins. But this process also produces a variety of small peptides as intermediate cleavage products. In the present study we tested whether amino acids actually are the natural and adequate stimuli for the olfactory receptors they bind to. Alternatively, these olfactory receptors could be peptide receptors which also bind amino acids though at lower affinity. Employing calcium imaging in acute slices of the main olfactory epithelium of the fully aquatic larvae of Xenopus laevis we show that amino acids, and not peptides, are more effective waterborne odorants. PMID:23300867

  17. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells

    PubMed Central

    Hopkins, Mandi M.; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E.

    2016-01-01

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor. PMID:26821052

  18. Eicosopentaneoic Acid and Other Free Fatty Acid Receptor Agonists Inhibit Lysophosphatidic Acid- and Epidermal Growth Factor-Induced Proliferation of Human Breast Cancer Cells.

    PubMed

    Hopkins, Mandi M; Zhang, Zhihong; Liu, Ze; Meier, Kathryn E

    2016-01-26

    Many key actions of ω-3 (n-3) fatty acids have recently been shown to be mediated by two G protein-coupled receptors (GPCRs) in the free fatty acid receptor (FFAR) family, FFA1 (GPR40) and FFA4 (GPR120). n-3 Fatty acids inhibit proliferation of human breast cancer cells in culture and in animals. In the current study, the roles of FFA1 and FFA4 were investigated. In addition, the role of cross-talk between GPCRs activated by lysophosphatidic acid (LPA), and the tyrosine kinase receptor activated by epidermal growth factor (EGF), was examined. In MCF-7 and MDA-MB-231 human breast cancer cell lines, both LPA and EGF stimulated proliferation, Erk activation, Akt activation, and CCN1 induction. LPA antagonists blocked effects of LPA and EGF on proliferation in MCF-7 and MDA-MB-231, and on cell migration in MCF-7. The n-3 fatty acid eicosopentaneoic acid inhibited LPA- and EGF-induced proliferation in both cell lines. Two synthetic FFAR agonists, GW9508 and TUG-891, likewise inhibited LPA- and EGF-induced proliferation. The data suggest a major role for FFA1, which was expressed by both cell lines. The results indicate that n-3 fatty acids inhibit breast cancer cell proliferation via FFARs, and suggest a mechanism involving negative cross-talk between FFARS, LPA receptors, and EGF receptor.

  19. Mechanisms of triglyceride metabolism in patients with bile acid diarrhea

    PubMed Central

    Sagar, Nidhi Midhu; McFarlane, Michael; Nwokolo, Chuka; Bardhan, Karna Dev; Arasaradnam, Ramesh Pulendran

    2016-01-01

    Bile acids (BAs) are essential for the absorption of lipids. BA synthesis is inhibited through intestinal farnesoid X receptor (FXR) activity. BA sequestration is known to influence BA metabolism and control serum lipid concentrations. Animal data has demonstrated a regulatory role for the FXR in triglyceride metabolism. FXR inhibits hepatic lipogenesis by inhibiting the expression of sterol regulatory element binding protein 1c via small heterodimer primer activity. Conversely, FXR promotes free fatty acids oxidation by inducing the expression of peroxisome proliferator-activated receptor α. FXR can reduce the expression of microsomal triglyceride transfer protein, which regulates the assembly of very low-density lipoproteins (VLDL). FXR activation in turn promotes the clearance of circulating triglycerides by inducing apolipoprotein C-II, very low-density lipoproteins receptor (VLDL-R) and the expression of Syndecan-1 together with the repression of apolipoprotein C-III, which increases lipoprotein lipase activity. There is currently minimal clinical data on triglyceride metabolism in patients with bile acid diarrhoea (BAD). Emerging data suggests that a third of patients with BAD have hypertriglyceridemia. Further research is required to establish the risk of hypertriglyceridaemia in patients with BAD and elicit the mechanisms behind this, allowing for targeted treatment. PMID:27570415

  20. G-protein-coupled receptors for free fatty acids: nutritional and therapeutic targets.

    PubMed

    Milligan, Graeme; Ulven, Trond; Murdoch, Hannah; Hudson, Brian D

    2014-06-01

    It is becoming evident that nutrients and metabolic intermediates derived from such nutrients regulate cellular function by activating a number of cell-surface G-protein coupled receptors (GPCRs). Until now, members of the GPCR family have largely been considered as the molecular targets that communicate cellular signals initiated by hormones and neurotransmitters. Recently, based on tissue expression patterns of these receptors and the concept that they may elicit the production of a range of appetite- and hunger-regulating peptides, such nutrient sensing GPCRs are attracting considerable attention due to their potential to modulate satiety, improve glucose homeostasis and supress the production of various pro-inflammatory mediators. Despite the developing interests in these nutrients sensing GPCR both as sensors of nutritional status, and targets for limiting the development of metabolic diseases, major challenges remain to exploit their potential for therapeutic purposes. Mostly, this is due to limited characterisation and validation of these receptors because of paucity of selective and high-potency/affinity pharmacological agents to define the detailed function and regulation of these receptors. However, ongoing clinical trials of agonists of free fatty acid receptor 1 suggest that this receptor and other receptors for free fatty acids may provide a successful strategy for controlling hyperglycaemia and providing novel approaches to treat diabetes. Receptors responsive to free fatty acid have been of particular interest, and some aspects of these are considered herein.

  1. Activity of 2-substituted lysophosphatidic acid (LPA) analogs at LPA receptors: discovery of a LPA1/LPA3 receptor antagonist.

    PubMed

    Heise, C E; Santos, W L; Schreihofer, A M; Heasley, B H; Mukhin, Y V; Macdonald, T L; Lynch, K R

    2001-12-01

    The physiological implications of lysophosphatidic acid occupancy of individual receptors are largely unknown because selective agonists/antagonists are unavailable currently. The molecular cloning of three high-affinity lysophosphatidic acid receptors, LPA1, LPA2, and LPA3, provides a platform for developing receptor type-selective ligands. Starting with an N-acyl ethanolamide phosphate LPA analog, we made a series of substitutions at the second carbon to generate compounds with varying spatial, stereochemical, and electronic characteristics. Analysis of this series at each recombinant LPA receptor using a guanosine 5'-O-(3-[35S]thio)triphosphate (GTP[gamma35S]) binding assay revealed sharp differences in activity. Our results suggest that these receptors have one spatially restrictive binding pocket that interacts with the 2-substituted moieties and prefers small hydrophobic groups and hydrogen bonding functionalities. The agonist activity predicted by the GTP[gamma35S] binding assay was reflected in the activity of a subset of compounds in increasing arterial pressure in anesthetized rats. One compound with a bulky hydrophobic group (VPC12249) was a dual LPA1/LPA3 competitive antagonist. Several compounds that had smaller side chains were found to be LPA1-selective agonists.

  2. Betulin binds to gamma-aminobutyric acid receptors and exerts anticonvulsant action in mice.

    PubMed

    Muceniece, Ruta; Saleniece, Kristine; Rumaks, Juris; Krigere, Liga; Dzirkale, Zane; Mezhapuke, Rudolfs; Zharkova, Olga; Klusa, Vija

    2008-10-01

    The lupane type pentacyclic triterpenes: lupeol, betulin, and betulinic acid are widely distributed natural compounds. Recently, pharmaceutical compositions from plant extracts (family Marcgraviaceae) containing betulinic acid, have been patented as anxiolytic remedies. To extend our knowledge of the CNS effects of the triterpenes, we suggest here that the chemically related lupeol, betulin and betulinic acid may interact with the brain neurotransmitter gamma-aminobutyric acid (GABA) receptors in vitro and in vivo. Using radioligand receptor-binding assay, we showed that only betulin bound to the GABA(A)-receptor sites in mice brain in vitro and antagonised the GABA(A)-receptor antagonist bicuculline-induced seizures in mice after intracisternal and intraperitoneal administration. Neither betulinic acid nor lupeol bound to GABA(A) receptor nor did they inhibit bicuculline-induced seizures in vivo. These findings demonstrate for the first time the CNS effects of betulin in vivo, and they also show distinct GABA(A)-receptor-related properties of lupane type triterpenes. These findings may open new avenues in understanding the central effects of betulin, and they also indicate possibilities for novel drug design on the basis of betulin structure.

  3. γ-Hydroxybutyric acid (GHB) is not an agonist of extrasynaptic GABAA receptors.

    PubMed

    Connelly, William M; Errington, Adam C; Crunelli, Vincenzo

    2013-01-01

    γ-Hydroxybutyric acid (GHB) is an endogenous compound and a drug used clinically to treat the symptoms of narcolepsy. GHB is known to be an agonist of GABAB receptors with millimolar affinity, but also binds with much higher affinity to another site, known as the GHB receptor. While a body of evidence has shown that GHB does not bind to GABAA receptors widely, recent evidence has suggested that the GHB receptor is in fact on extrasynaptic α4β1δ GABAA receptors, where GHB acts as an agonist with an EC50 of 140 nM. We investigated three neuronal cell types that express a tonic GABAA receptor current mediated by extrasynaptic receptors: ventrobasal (VB) thalamic neurons, dentate gyrus granule cells and striatal medium spiny neurons. Using whole-cell voltage clamp in brain slices, we found no evidence that GHB (10 µM) induced any GABAA receptor mediated current in these cell types, nor that it modulated inhibitory synaptic currents. Furthermore, a high concentration of GHB (3 mM) was able to produce a GABAB receptor mediated current, but did not induce any other currents. These results suggest either that GHB is not a high affinity agonist at native α4β1δ receptors, or that these receptors do not exist in classical areas associated with extrasynaptic currents.

  4. AMPA receptor subunits expression and phosphorylation in cingulate cortex in rats following esophageal acid exposure

    PubMed Central

    BANERJEE, B.; MEDDA, B. K.; POCHIRAJU, S.; KANNAMPALLI, P.; LANG, I. M.; SENGUPTA, J. N.; SHAKER, R.

    2014-01-01

    Background We recently reported an increase in N-methyl-d-aspartate (NMDA) receptor subunit expression and CaMKII-dependent phosphorylation of NR2B in the rostral cingulate cortical (rCC) neurons following esophageal acid exposure in rats. As α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors mediate the fast excitatory transmission and play a critical role in synaptic plasticity, in this study, we investigated the effect of esophageal acid exposure in rats on the expression of AMPA receptor subunits and the involvement of these molecular alterations in acid-induced sensitization of neurons in the anterior cingulate (ACC) and midcingulate (MCC) cortices. Methods In molecular study, we examined GluA1 and GluA2 expression and phosphorylation in membrane preparations and in the isolated postsynaptic densities (PSDs) from rats receiving acute esophageal exposure of either saline (control group) or 0.1 NHCl (experimental group). In electrophysiological study, the effect of selective AMPA receptor (Ca2+ permeable) antagonist IEM-1460 and CaMKII inhibitor KN-93 was tested on responses of cortical neurons during acid infusion to address the underlying molecular mechanism of acid-induced sensitization. Key Results The acid exposure significantly increased expression of GluA1, pGluA1Ser831, and phosphorylated CaMKIIThr286, in the cortical membrane preparations. In isolated PSDs, a significant increase in pGluA1Ser831 was observed in acid-treated rats compared with controls. Microinjection of IEM-1460 or KN-93 near the recording site significantly attenuated acid-induced sensitization of cortical neurons. Conclusions & Inferences The underlying mechanism of acid-induced cortical sensitization involves upregulation and CaMKII-mediated phosphorylation of GluA1. These molecular changes of AMPA receptors subunit GluA1 in the cortical neurons might play an important role in acid-induced esophageal hypersensitivity. PMID:24118589

  5. Concomitant Action of Structural Elements and Receptor Phosphorylation Determines Arrestin-3 Interaction with the Free Fatty Acid Receptor FFA4*

    PubMed Central

    Butcher, Adrian J.; Hudson, Brian D.; Shimpukade, Bharat; Alvarez-Curto, Elisa; Prihandoko, Rudi; Ulven, Trond; Milligan, Graeme; Tobin, Andrew B.

    2014-01-01

    In addition to being nutrients, free fatty acids act as signaling molecules by activating a family of G protein-coupled receptors. Among these is FFA4, previously called GPR120, which responds to medium and long chain fatty acids, including health-promoting ω-3 fatty acids, which have been implicated in the regulation of metabolic and inflammatory responses. Here we show, using mass spectrometry, mutagenesis, and phosphospecific antibodies, that agonist-regulated phosphorylation of the human FFA4 receptor occurred primarily at five residues (Thr347, Thr349, Ser350, Ser357, and Ser360) in the C-terminal tail. Mutation of these residues reduced both the efficacy and potency of ligand-mediated arrestin-3 recruitment as well as affecting recruitment kinetics. Combined mutagenesis of all five of these residues was insufficient to fully abrogate interaction with arrestin-3, but further mutagenesis of negatively charged residues revealed additional structural components for the interaction with arrestin-3 within the C-terminal tail of the receptor. These elements consist of the acidic residues Glu341, Asp348, and Asp355 located close to the phosphorylation sites. Receptor phosphorylation thus operates in concert with structural elements within the C-terminal tail of FFA4 to allow for the recruitment of arrestin-3. Importantly, these mechanisms of arrestin-3 recruitment operate independently from Gq/11 coupling, thereby offering the possibility that ligands showing stimulus bias could be developed that exploit these differential coupling mechanisms. Furthermore, this provides a strategy for the design of biased receptors to probe physiologically relevant signaling. PMID:24817122

  6. REACTIVITY PROFILE OF LIGANDS OF MAMMALIAN RETINOIC ACID RECEPTORS: A PRELIMINARY COREPA ANALYSIS

    EPA Science Inventory

    Retinoic acid and associated derivatives comprise a class of endogenous hormones that bind to and activate different families of retinoic acid receptors (RARs, RXRs), and control many aspects of vertebrate development. Identification of potential RAR and RXR ligands is of interes...

  7. Developmental toxicity of perfluorononanoic acid is dependent on peroxisome proliferator activated receptor-alpha.

    EPA Science Inventory

    Perfluorononanoic acid (PFNA) is one of the predominant perfluoroalkyl acids in the environment and in tissues of humans and wildlife. PFNA strongly activates the mouse and human peroxisome proliferator-activated receptor-alpha (PPARα) in vitro and negatively impacts development ...

  8. The effects of avermectin on amino acid neurotransmitters and their receptors in the pigeon brain.

    PubMed

    Chen, Li-Jie; Sun, Bao-Hong; Cao, Ye; Yao, Hai-Dong; Qu, Jian-Ping; Liu, Ci; Xu, Shi-Wen; Li, Shu

    2014-03-01

    The objective of this study was to examine the effects of avermectin (AVM) on amino acid neurotransmitters and their receptors in the pigeon brain. Four groups two-month-old American king pigeons (n=20/group) were fed either a commercial diet or an AVM-supplemented diet (20mg/kg·diet, 40 mg/kg·diet, or 60 mg/kg·diet) for 30, 60, or 90 days. The contents of aspartic acid (ASP), glutamate (GLU), glycine (GLY), and γ-aminobutyric acid (GABA) in the brain tissues were determined using ultraviolet high-performance liquid chromatography (HPLC). The expression levels of the GLU and GABA receptor genes were analyzed using real-time quantitative polymerase chain reaction (qPCR). The results indicate that AVM exposure significantly enhances the contents of GABA, GLY, GLU, and ASP in the cerebrum, cerebellum, and optic lobe. In addition, AVM exposure increases the mRNA expression levels of γ-aminobutyric acid type A receptor (GABAAR), γ-aminobutyric acid type B receptor (GABABR), N-methyl-d-aspartate 1 receptor (NR1), N-methyl-d-aspartate 2A receptor (NR2A), and N-methyl-d-aspartate 2B receptor (NR2B) in a dose- and time-dependent manner. Moreover, we found that the most damaged organ was the cerebrum, followed by the cerebellum, and then the optic lobe. These results show that the AVM-induced neurotoxicity may be associated with its effects on amino acid neurotransmitters and their receptors. The information presented in this study will help supplement the available data for future AVM toxicity studies.

  9. Effects of arachidonic acid on FFA4 receptor: Signaling, phosphorylation and internalization.

    PubMed

    Villegas-Comonfort, S; Takei, Y; Tsujimoto, G; Hirasawa, A; García-Sáinz, J A

    2017-02-01

    Arachidonic acid increased intracellular calcium, in cells expressing green fluorescent protein-tagged human FFA4 receptors, with an EC50 of ~40µM. This action was not blocked by cyclooxygenase or lipoxigenase inhibitors but it was inhibited by AH7614, a FFA4 antagonist. Arachidonic acid induced ERK activation accompanied by EGF receptor transactivation. However, EGF transactivation was not the major mechanism through which the fatty acid induced ERK phosphorylation, as evidenced by the inability of AG1478 to block it. Arachidonic acid increased FFA4 receptor phosphorylation that reached its maximum within 15min with an EC50 of ~30µM; inhibitors of protein kinase C partially diminish this effect and AH7614 blocked it. Arachidonic acid induced rapid and sustained Akt/PKB phosphorylation and FFA4 - β-arrestin interaction. Confocal microscopy evidenced that FFA4 receptor activation and phosphorylation were associated to internalization. In conclusion, arachidonic acid is a bona fide FFA4 receptor agonist.

  10. Bile acid diarrhoea and FGF19: new views on diagnosis, pathogenesis and therapy.

    PubMed

    Walters, Julian R F

    2014-07-01

    Chronic diarrhoea induced by bile acids is common and the underlying mechanisms are linked to homeostatic regulation of hepatic bile acid synthesis by fibroblast growth factor 19 (FGF19). Increasing evidence, including that from several large case series using SeHCAT (selenium homocholic acid taurine) tests for diagnosis, indicates that bile acid diarrhoea (BAD) accounts for a sizeable proportion of patients who would otherwise be diagnosed with IBS. Studies of other approaches for diagnosis of BAD have shown increased bile acid synthesis, increased faecal levels of primary bile acids, dysbiosis and different urinary volatile organic compounds when compared with healthy controls or with other diseases. The role of the ileal hormone FGF19 in BAD has been strengthened: a prospective clinical study has confirmed low FGF19 levels in BAD, and so a test to measure these levels could be developed for diagnosis. In animal models, FGF19 depletion by antibodies produces severe diarrhoea. Bile acids affect colonic function through farnesoid X receptor (FXR) and TGR5 receptors. As well as these effects in the colon, FXR-dependent stimulation of ileal FGF19 production could be a logical mechanism to provide therapeutic benefit in BAD. Further studies of FGF19 in humans hold promise in providing novel treatments for this cause of chronic diarrhoea.

  11. Classification of inhibitory amino acid receptors in the mammalian nervous system.

    PubMed

    Simmonds, M A

    1986-01-01

    Electrophysiological and pharmacological evidence is summarized for the existence of an inhibitory receptor system operated by glycine and another two separate systems operated by gamma-aminobutyric acid (GABA) through GABA-A and GABA-B receptors, respectively. Claims for subclasses of GABA-A receptor are critically reviewed and found not-proven. A quantitative pharmacological profile of the GABA-A receptor and associated regulatory sites for picrotoxin, barbiturates and benzodiazepines on the dorsal funiculus of the rat cuneate nucleus is described. When compared with this profile and the pharmacological properties of the glycine receptor complex, the effects of taurine cannot be entirely explained by actions on these two receptor systems.

  12. Bile Acid Signaling Is Involved in the Neurological Decline in a Murine Model of Acute Liver Failure

    PubMed Central

    McMillin, Matthew; Frampton, Gabriel; Quinn, Matthew; Ashfaq, Samir; de los Santos, Mario; Grant, Stephanie; DeMorrow, Sharon

    2017-01-01

    Hepatic encephalopathy is a serious neurological complication of liver failure. Serum bile acids are elevated after liver damage and may disrupt the blood-brain barrier and enter the brain. Our aim was to assess the role of serum bile acids in the neurological complications after acute liver failure. C57Bl/6 or cytochrome p450 7A1 knockout (Cyp7A1−/−) mice were fed a control, cholestyramine-containing, or bile acid–containing diet before azoxymethane (AOM)-induced acute liver failure. In parallel, mice were given an intracerebroventricular infusion of farnesoid X receptor (FXR) Vivo-morpholino before AOM injection. Liver damage, neurological decline, and molecular analyses of bile acid signaling were performed. Total bile acid levels were increased in the cortex of AOM-treated mice. Reducing serum bile acids via cholestyramine feeding or using Cyp7A1−/− mice reduced bile acid levels and delayed AOM-induced neurological decline, whereas cholic acid or deoxycholic acid feeding worsened AOM-induced neurological decline. The expression of bile acid signaling machinery apical sodium-dependent bile acid transporter, FXR, and small heterodimer partner increased in the frontal cortex, and blocking FXR signaling delayed AOM-induced neurological decline. In conclusion, circulating bile acids may play a pathological role during hepatic encephalopathy, although precisely how they dysregulate normal brain function is unknown. Strategies to minimize serum bile acid concentrations may reduce the severity of neurological complications associated with liver failure. PMID:26683664

  13. RAGE is a nucleic acid receptor that promotes inflammatory responses to DNA

    PubMed Central

    Sirois, Cherilyn M.; Jin, Tengchuan; Miller, Allison L.; Bertheloot, Damien; Nakamura, Hirotaka; Horvath, Gabor L.; Mian, Abubakar; Jiang, Jiansheng; Schrum, Jacob; Bossaller, Lukas; Pelka, Karin; Garbi, Natalio; Brewah, Yambasu; Tian, Jane; Chang, ChewShun; Chowdhury, Partha S.; Sims, Gary P.; Kolbeck, Roland; Coyle, Anthony J.; Humbles, Alison A.

    2013-01-01

    Recognition of DNA and RNA molecules derived from pathogens or self-antigen is one way the mammalian immune system senses infection and tissue damage. Activation of immune signaling receptors by nucleic acids is controlled by limiting the access of DNA and RNA to intracellular receptors, but the mechanisms by which endosome-resident receptors encounter nucleic acids from the extracellular space are largely undefined. In this study, we show that the receptor for advanced glycation end-products (RAGE) promoted DNA uptake into endosomes and lowered the immune recognition threshold for the activation of Toll-like receptor 9, the principal DNA-recognizing transmembrane signaling receptor. Structural analysis of RAGE–DNA complexes indicated that DNA interacted with dimers of the outermost RAGE extracellular domains, and could induce formation of higher-order receptor complexes. Furthermore, mice deficient in RAGE were unable to mount a typical inflammatory response to DNA in the lung, indicating that RAGE is important for the detection of nucleic acids in vivo. PMID:24081950

  14. Modulatory effects of unsaturated fatty acids on the binding of glucocorticoids to rat liver glucocorticoid receptors.

    PubMed

    Vallette, G; Vanet, A; Sumida, C; Nunez, E A

    1991-09-01

    Binding of the synthetic glucocorticoid dexamethasone to the rat liver cytosol glucocorticoid receptor was inhibited by physiological concentrations of nonesterified fatty acids as a function of increasing dose, degree of unsaturation, and chain length of the fatty acid. Polyunsaturated fatty acids were the most potent inhibitors. Scatchard analysis and Line-weaver-Burk plots of the binding data revealed that both the association constants and number of binding sites decreased and that polyunsaturated fatty acids inhibition was of a mixed non-competitive type. The dissociation rate constant of [3H]dexamethasone from glucocorticoid receptors was increased by up to 10 times in the presence of docosahexaenoic acid, whereas a competitive inhibitor like the glucocorticoid antagonist RU 38486 had no effect. Moreover, sucrose density gradient analysis showed that docosahexaenoic acid inhibited the binding of [3H] dexamethasone to both the 8.8S and 4S forms. The results strongly suggest that unsaturated fatty acids are interacting at a site on the receptor different from the hormone binding site and the heat shock protein and that by binding to a second site unsaturated fatty acids greatly change the conformation of the hormone binding site to reduce its affinity for the hormone, either partially or completely depending on the concentration and the class of the fatty acid.

  15. Stimulation of acid secretion and phosphoinositol production by rat parietal cell muscarinic M sub 2 receptors

    SciTech Connect

    Pfeiffer, A.; Rochlitz, H.; Herz, A.; Paumgartner, G. )

    1988-04-01

    The muscarinic receptor system involved in hydrogen production by enriched rat gastric parietal cells was investigated. Muscarinic receptor density determined by (N-methyl-{sup 3}H)scopolamine binding was 8,100/cell. The receptor appeared to be of the M{sub 2} muscarinic receptor subtype, since it had a low affinity (K{sub d} 189 nM) for the M{sub 1} receptor antagonist pirenzepine compared with atropine. Receptor activation by carbachol rapidly augmented levels of polyphosphoinositides, indicating an activation of phospholipase C. The dose-response relations for the increase in inositol phosphates closely paralleled the binding of carbachol to muscarinic receptors. The inositol phosphate response was antagonized by pirenzepine with a K{sub i} of 177 nM. the stimulation of inositol phosphate levels by carbachol correlated well with the stimulation of ({sup 14}C)aminopyrine uptake, determine as an index of acid secretion. The muscarinic agonists oxotremorine, pilocarpine, and bethanechol elicited partial increases in inositol phosphates at maximal drug concentrations, and these partial increases correlated with their ability to stimulate ({sup 14}C)aminopyrine uptake. These data indicate that inositolpolyphosphates may be a second messenger of M{sub 2} receptors stimulating acid secretion.

  16. Free fatty acid receptors: emerging targets for treatment of diabetes and its complications

    PubMed Central

    Vangaveti, Venkat; Shashidhar, Venkatesh; Jarrod, Ghassan; Baune, Bernhard T.; Kennedy, R. Lee

    2010-01-01

    Fatty acids (FAs) are important as metabolic substrates and as structural components of biological membranes. However, they also function as signalling molecules. Recently, a series of G protein-coupled receptors (GPRs) for FAs has been described and characterized. These receptors have differing specificities for FAs of differing chain length and degree of saturation, for FA derivatives such as oleoylethanolamide, and for oxidized FAs. They are a critical component of the body's nutrient sensing apparatus, and small molecule agonists and antagonists of these receptors show considerable promise in the management of diabetes and its complications. Agonists of the long-chain free fatty acid receptors FFAR1 and GPR119 act as insulin secretagogues, both directly and by increasing incretins. Although, drugs acting at short-chain FFA receptors (FFAR2 and FFAR3) have not yet been developed, they are attractive targets as they regulate nutrient balance through effects in the intestine and adipose tissue. These include regulation of the secretion of cholecystokinin, peptide YY and leptin. Finally, GPR132 is a receptor for oxidized FAs, which may be a sensor of lipid overload and oxidative stress, and which is involved in atherosclerosis. Regulation of its signalling pathways with drugs may decrease the macrovascular risk experienced by diabetic patients. In summary, FA receptors are emerging drug targets that are involved in the regulation of nutrient status and carbohydrate tolerance, and modulators of these receptors may well figure prominently in the next generation of antidiabetic drugs. PMID:23148161

  17. Topoisomerase IIβ Negatively Modulates Retinoic Acid Receptor α Function: a Novel Mechanism of Retinoic Acid Resistance▿

    PubMed Central

    McNamara, Suzan; Wang, Hongling; Hanna, Nessrine; Miller, Wilson H.

    2008-01-01

    Interactions between retinoic acid (RA) receptor α (RARα) and coregulators play a key role in coordinating gene transcription and myeloid differentiation. In patients with acute promyelocytic leukemia (APL), the RARα gene is fused with the promyelocytic leukemia (PML) gene via the t(15;17) translocation, resulting in the expression of a PML/RARα fusion protein. Here, we report that topoisomerase II beta (TopoIIβ) associates with and negatively modulates RARα transcriptional activity and that increased levels of and association with TopoIIβ cause resistance to RA in APL cell lines. Knockdown of TopoIIβ was able to overcome resistance by permitting RA-induced differentiation and increased RA gene expression. Overexpression of TopoIIβ in clones from an RA-sensitive cell line conferred resistance by a reduction in RA-induced expression of target genes and differentiation. Chromatin immunoprecipitation assays indicated that TopoIIβ is bound to an RA response element and that inhibition of TopoIIβ causes hyperacetylation of histone 3 at lysine 9 and activation of transcription. Our results identify a novel mechanism of resistance in APL and provide further insight to the role of TopoIIβ in gene regulation and differentiation. PMID:18212063

  18. Modulation of Retinoic Acid Receptor-related Orphan Receptor α and γ Activity by 7-Oxygenated Sterol Ligands*

    PubMed Central

    Wang, Yongjun; Kumar, Naresh; Solt, Laura A.; Richardson, Timothy I.; Helvering, Leah M.; Crumbley, Christine; Garcia-Ordonez, Ruben D.; Stayrook, Keith R.; Zhang, Xi; Novick, Scott; Chalmers, Michael J.; Griffin, Patrick R.; Burris, Thomas P.

    2010-01-01

    The retinoic acid receptor-related orphan receptors α and γ (RORα (NR1F1) and RORγ (NR1F3)) are orphan nuclear receptors and perform critical roles in regulation of development, metabolism, and immune function. Cholesterol and cholesterol sulfate have been suggested to be RORα ligands, but the physiological significance is unclear. To date, no endogenous RORγ ligands have been described. Here, we demonstrate that 7-oxygenated sterols function as high affinity ligands for both RORα and RORγ by directly binding to their ligand-binding domains (Ki ∼20 nm), modulating coactivator binding, and suppressing the transcriptional activity of the receptors. One of the 7-oxygenated sterols, 7α-hydroxycholesterol (7α-OHC), serves as a key intermediate in bile acid metabolism, and we show that 7α-OHC modulates the expression of ROR target genes, including Glc-6-Pase and phosphoenolpyruvate carboxykinase, in an ROR-dependent manner. Furthermore, glucose output from hepatocytes is suppressed by 7α-OHC functioning as an RORα/γ ligand. Thus, RORα and RORγ are ligand-regulated members of the NR superfamily and may serve as sensors for 7-oxygenated sterols. PMID:19965867

  19. Mechanisms for the activation of Toll-like receptor 2/4 by saturated fatty acids and inhibition by docosahexaenoic acid.

    PubMed

    Hwang, Daniel H; Kim, Jeong-A; Lee, Joo Young

    2016-08-15

    Saturated fatty acids can activate Toll-like receptor 2 (TLR2) and TLR4 but polyunsaturated fatty acids, particularly docosahexaenoic acid (DHA) inhibit the activation. Lipopolysaccharides (LPS) and lipopetides, ligands for TLR4 and TLR2, respectively, are acylated by saturated fatty acids. Removal of these fatty acids results in loss of their ligand activity suggesting that the saturated fatty acyl moieties are required for the receptor activation. X-ray crystallographic studies revealed that these saturated fatty acyl groups of the ligands directly occupy hydrophobic lipid binding domains of the receptors (or co-receptor) and induce the dimerization which is prerequisite for the receptor activation. Saturated fatty acids also induce the dimerization and translocation of TLR4 and TLR2 into lipid rafts in plasma membrane and this process is inhibited by DHA. Whether saturated fatty acids induce the dimerization of the receptors by interacting with these lipid binding domains is not known. Many experimental results suggest that saturated fatty acids promote the formation of lipid rafts and recruitment of TLRs into lipid rafts leading to ligand independent dimerization of the receptors. Such a mode of ligand independent receptor activation defies the conventional concept of ligand induced receptor activation; however, this may enable diverse non-microbial molecules with endogenous and dietary origins to modulate TLR-mediated immune responses. Emerging experimental evidence reveals that TLRs play a key role in bridging diet-induced endocrine and metabolic changes to immune responses.

  20. Identification of the Orphan G Protein-coupled Receptor GPR31 as a Receptor for 12-(S)-Hydroxyeicosatetraenoic Acid*

    PubMed Central

    Guo, Yande; Zhang, Wenliang; Giroux, Craig; Cai, Yinlong; Ekambaram, Prasanna; Dilly, Ashok-kumar; Hsu, Andrew; Zhou, Senlin; Maddipati, Krishna Rao; Liu, Jingjing; Joshi, Sangeeta; Tucker, Stephanie C.; Lee, Menq-Jer; Honn, Kenneth V.

    2011-01-01

    Hydroxy fatty acids are critical lipid mediators involved in various pathophysiologic functions. We cloned and identified GPR31, a plasma membrane orphan G protein-coupled receptor that displays high affinity for the human 12-lipoxygenase-derived product 12-(S)-hydroxy-5,8,10,14-eicosatetraenoic acid (HETE). Thus, GPR31 is named 12-(S)-HETE receptor (12-HETER) in this study. The cloned 12-HETER demonstrated high affinity binding for 12-(S)-[3H]HETE (Kd = 4.8 ± 0.12 nm). Also, 12-(S)-HETE efficiently and selectively stimulated GTPγS coupling in the membranes of 12-HETER-transfected cells (EC50 = 0.28 ± 1.26 nm). Activating GTPγS coupling with 12-(S)-HETE proved to be both regio- and stereospecific. Also, 12-(S)-HETE/12-HETER interactions lead to activation of ERK1/2, MEK, and NFκB. Moreover, knocking down 12-HRTER specifically inhibited 12-(S)-HETE-stimulated cell invasion. Thus, 12-HETER represents the first identified high affinity receptor for the 12-(S)-HETE hydroxyl fatty acids. PMID:21712392

  1. Expression of functional receptors by the human gamma-aminobutyric acid A gamma 2 subunit.

    PubMed

    Martínez-Torres, Ataúlfo; Miledi, Ricardo

    2004-03-02

    gamma-Aminobutyric acid A (GABA(A)) receptors are heteromeric membrane proteins formed mainly by various combinations of alpha, beta, and gamma subunits; and it is commonly thought that the gamma 2 subunit alone does not form functional receptors. In contrast, we found that cDNA encoding the gamma 2L subunit of the human GABA(A) receptor, injected alone into Xenopus oocytes, expressed functional GABA receptors whose properties were investigated by using the two-microelectrode voltage-clamp technique. GABA elicited desensitizing membrane currents that recovered after a few minutes' wash. Repetitive applications of GABA induced a "run-up" of GABA currents that nearly doubled the amplitude of the first response. The GABA currents inverted direction at about -30 mV, indicating that they are carried mainly by Cl(-) ions. The homomeric gamma 2L receptors were also activated by beta-alanine > taurine > glycine, and, like some types of heteromeric GABA(A) receptors, the gamma 2L receptors were blocked by bicuculline and were potentiated by pentobarbital and flunitrazepam. These results indicate that the human gamma 2L subunit is capable of forming fully functional GABA receptors by itself in Xenopus oocytes and suggest that the roles proposed for the various subunits that make up the heteromeric GABA(A) receptors in situ require further clarification.

  2. Expression of functional receptors by the human γ-aminobutyric acid A γ2 subunit

    PubMed Central

    Martínez-Torres, Ataúlfo; Miledi, Ricardo

    2004-01-01

    γ-Aminobutyric acid A (GABAA) receptors are heteromeric membrane proteins formed mainly by various combinations of α, β, and γ subunits; and it is commonly thought that the γ2 subunit alone does not form functional receptors. In contrast, we found that cDNA encoding the γ2L subunit of the human GABAA receptor, injected alone into Xenopus oocytes, expressed functional GABA receptors whose properties were investigated by using the two-microelectrode voltage-clamp technique. GABA elicited desensitizing membrane currents that recovered after a few minutes' wash. Repetitive applications of GABA induced a “run-up” of GABA currents that nearly doubled the amplitude of the first response. The GABA currents inverted direction at about -30 mV, indicating that they are carried mainly by Cl- ions. The homomeric γ2L receptors were also activated by β-alanine > taurine > glycine, and, like some types of heteromeric GABAA receptors, the γ2L receptors were blocked by bicuculline and were potentiated by pentobarbital and flunitrazepam. These results indicate that the human γ2L subunit is capable of forming fully functional GABA receptors by itself in Xenopus oocytes and suggest that the roles proposed for the various subunits that make up the heteromeric GABAA receptors in situ require further clarification. PMID:14981251

  3. Optical control of trimeric P2X receptors and acid-sensing ion channels.

    PubMed

    Browne, Liam E; Nunes, João P M; Sim, Joan A; Chudasama, Vijay; Bragg, Laricia; Caddick, Stephen; North, R Alan

    2014-01-07

    P2X receptors are trimeric membrane proteins that function as ion channels gated by extracellular ATP. We have engineered a P2X2 receptor that opens within milliseconds by irradiation at 440 nm, and rapidly closes at 360 nm. This requires bridging receptor subunits via covalent attachment of 4,4'-bis(maleimido)azobenzene to a cysteine residue (P329C) introduced into each second transmembrane domain. The cis-trans isomerization of the azobenzene pushes apart the outer ends of the transmembrane helices and opens the channel in a light-dependent manner. Light-activated channels exhibited similar unitary currents, rectification, calcium permeability, and dye uptake as P2X2 receptors activated by ATP. P2X3 receptors with an equivalent mutation (P320C) were also light sensitive after chemical modification. They showed typical rapid desensitization, and they could coassemble with native P2X2 subunits in pheochromocytoma cells to form light-activated heteromeric P2X2/3 receptors. A similar approach was used to open and close human acid-sensing ion channels (ASICs), which are also trimers but are unrelated in sequence to P2X receptors. The experiments indicate that the opening of the permeation pathway requires similar and substantial movements of the transmembrane helices in both P2X receptors and ASICs, and the method will allow precise optical control of P2X receptors or ASICs in intact tissues.

  4. Docking simulations suggest that all- trans retinoic acid could bind to retinoid X receptors

    NASA Astrophysics Data System (ADS)

    Tsuji, Motonori; Shudo, Koichi; Kagechika, Hiroyuki

    2015-10-01

    Retinoid X receptors (RXRs) are ligand-controlled transcription factors which heterodimerize with other nuclear receptors to regulate gene transcriptions associated with crucial biological events. 9- cis retinoic acid (9cRA), which transactivates RXRs, is believed to be an endogenous RXR ligand. All- trans retinoic acid (ATRA) is a natural ligand for retinoic acid receptors (RARs), which heterodimerize with RXRs. Although the concentration of 9cRA in tissues is very low, ATRA is relatively abundant and some reports show that ATRA activates RXRs. We computationally studied the possibility of ATRA binding to RXRs using two different docking methods with our developed programs to assess the binding affinities of naturally occurring retinoids. The simulations showed good correlations to the reported binding affinities of these molecules for RXRs and RARs.

  5. Role of transient receptor potential and acid-sensing ion channels in peripheral inflammatory pain.

    PubMed

    White, John P M; Cibelli, Mario; Rei Fidalgo, Antonio; Paule, Cleoper C; Noormohamed, Faruq; Urban, Laszlo; Maze, Mervyn; Nagy, Istvan

    2010-03-01

    Pain originating in inflammation is the most common pathologic pain condition encountered by the anesthesiologist whether in the context of surgery, its aftermath, or in the practice of pain medicine. Inflammatory agents, released as components of the body's response to peripheral tissue damage or disease, are now known to be collectively capable of activating transient receptor potential vanilloid type 1, transient receptor potential vanilloid type 4, transient receptor potential ankyrin type 1, and acid-sensing ion channels, whereas individual agents may activate only certain of these ion channels. These ionotropic receptors serve many physiologic functions-as, indeed, do many of the inflammagens released in the inflammatory process. Here, we introduce the reader to the role of these ionotropic receptors in mediating peripheral pain in response to inflammation.

  6. Glutamate receptor-like channels in plants: a role as amino acid sensors in plant defence?

    PubMed Central

    Roberts, Michael R.

    2014-01-01

    Plant glutamate receptor-like genes (GLRs) are homologous to the genes for mammalian ionotropic glutamate receptors (iGluRs), after which they were named, but in the 16 years since their existence was first revealed, progress in elucidating their biological role has been disappointingly slow. Recently, however, studies from a number of laboratories focusing on the model plant species Arabidopsis thaliana (L.) have thrown new light on the functional properties of some members of the GLR gene family. One important finding has been that plant GLR receptors have a much broader ligand specificity than their mammalian iGluR counterparts, with evidence that some individual GLR receptors can be gated by as many as seven amino acids. These results, together with the ubiquity of their expression throughout the plant, open up the possibility that GLR receptors could have a pervasive role in plants as non-specific amino acid sensors in diverse biological processes. Addressing what one of these roles could be, recent studies examining the wound response and disease susceptibility in GLR knockout mutants have provided evidence that some members of clade 3 of the GLR gene family encode important components of the plant's defence response. Ways in which this family of amino acid receptors might contribute to the plant's ability to respond to an attack from pests and pathogens are discussed. PMID:24991414

  7. Lysophosphatidic Acid Signaling through the Lysophosphatidic Acid-1 Receptor Is Required for Alveolarization.

    PubMed

    Funke, Manuela; Knudsen, Lars; Lagares, David; Ebener, Simone; Probst, Clemens K; Fontaine, Benjamin A; Franklin, Alicia; Kellner, Manuela; Kühnel, Mark; Matthieu, Stephanie; Grothausmann, Roman; Chun, Jerold; Roberts, Jesse D; Ochs, Matthias; Tager, Andrew M

    2016-07-01

    Lysophosphatidic acid (LPA) signaling through one of its receptors, LPA1, contributes to both the development and the pathological remodeling after injury of many organs. Because we found previously that LPA-LPA1 signaling contributes to pulmonary fibrosis, here we investigated whether this pathway is also involved in lung development. Quantitative assessment of lung architecture of LPA1-deficient knock-out (KO) and wild-type (WT) mice at 3, 12, and 24 weeks of age using design-based stereology suggested the presence of an alveolarization defect in LPA1 KO mice at 3 weeks, which persisted as alveolar numbers increased in WT mice into adulthood. Across the ages examined, the lungs of LPA1 KO mice exhibited decreased alveolar numbers, septal tissue volumes, and surface areas, and increased volumes of the distal airspaces. Elastic fibers, critical to the development of alveolar septa, appeared less organized and condensed and more discontinuous in KO alveoli starting at P4. Tropoelastin messenger RNA expression was decreased in KO lungs, whereas expression of matrix metalloproteinases degrading elastic fibers was either decreased or unchanged. These results are consistent with the abnormal lung phenotype of LPA1 KO mice, being attributable to reduced alveolar septal formation during development, rather than to increased septal destruction as occurs in the emphysema of chronic obstructive pulmonary disease. Peripheral septal fibroblasts and myofibroblasts, which direct septation in late alveolarization, demonstrated reduced production of tropoelastin and matrix metalloproteinases, and diminished LPA-induced migration, when isolated from LPA1 KO mice. Taken together, our data suggest that LPA-LPA1 signaling is critically required for septation during alveolarization.

  8. Boronic acids as probes for investigation of allosteric modulation of the chemokine receptor CXCR3.

    PubMed

    Bernat, Viachaslau; Admas, Tizita Haimanot; Brox, Regine; Heinemann, Frank W; Tschammer, Nuska

    2014-11-21

    The chemokine receptor CXCR3 is a G protein-coupled receptor, which conveys extracellular signals into cells by changing its conformation upon agonist binding. To facilitate the mechanistic understanding of allosteric modulation of CXCR3, we combined computational modeling with the synthesis of novel chemical tools containing boronic acid moiety, site-directed mutagenesis, and detailed functional characterization. The design of boronic acid derivatives was based on the predictions from homology modeling and docking. The choice of the boronic acid moiety was dictated by its unique ability to interact with proteins in a reversible covalent way, thereby influencing conformational dynamics of target biomolecules. During the synthesis of the library we have developed a novel approach for the purification of drug-like boronic acids. To validate the predicted binding mode and to identify amino acid residues responsible for the transduction of signal through CXCR3, we conducted a site-directed mutagenesis study. With the use of allosteric radioligand RAMX3 we were able to establish the existence of a second allosteric binding pocket in CXCR3, which enables different binding modes of structurally closely related allosteric modulators of CXCR3. We have also identified residues Trp109(2.60) and Lys300(7.35) inside the transmembrane bundle of the receptor as crucial for the regulation of the G protein activation. Furthermore, we report the boronic acid 14 as the first biased negative allosteric modulator of the receptor. Overall, our data demonstrate that boronic acid derivatives represent an outstanding tool for determination of key receptor-ligand interactions and induction of ligand-biased signaling.

  9. Evolution of the bile salt nuclear receptor FXR in vertebrates*s⃞

    PubMed Central

    Reschly, Erica J.; Ai, Ni; Ekins, Sean; Welsh, William J.; Hagey, Lee R.; Hofmann, Alan F.; Krasowski, Matthew D.

    2008-01-01

    Bile salts, the major end metabolites of cholesterol, vary significantly in structure across vertebrate species, suggesting that nuclear receptors binding these molecules may show adaptive evolutionary changes. We compared across species the bile salt specificity of the major transcriptional regulator of bile salt synthesis, the farnesoid X receptor (FXR). We found that FXRs have changed specificity for primary bile salts across species by altering the shape and size of the ligand binding pocket. In particular, the ligand binding pockets of sea lamprey (Petromyzon marinus) and zebrafish (Danio rerio) FXRs, as predicted by homology models, are flat and ideal for binding planar, evolutionarily early bile alcohols. In contrast, human FXR has a curved binding pocket best suited for the bent steroid ring configuration typical of evolutionarily more recent bile acids. We also found that the putative FXR from the sea squirt Ciona intestinalis, a chordate invertebrate, was completely insensitive to activation by bile salts but was activated by sulfated pregnane steroids, suggesting that the endogenous ligands of this receptor may be steroidal in nature. Our observations present an integrated picture of the coevolution of bile salt structure and of the binding pocket of their target nuclear receptor FXR. PMID:18362391

  10. Bile acids reduce endocytosis of high-density lipoprotein (HDL) in HepG2 cells.

    PubMed

    Röhrl, Clemens; Eigner, Karin; Fruhwürth, Stefanie; Stangl, Herbert

    2014-01-01

    High-density lipoprotein (HDL) transports lipids to hepatic cells and the majority of HDL-associated cholesterol is destined for biliary excretion. Cholesterol is excreted into the bile directly or after conversion to bile acids, which are also present in the plasma as they are effectively reabsorbed through the enterohepatic cycle. Here, we provide evidence that bile acids affect HDL endocytosis. Using fluorescent and radiolabeled HDL, we show that HDL endocytosis was reduced in the presence of high concentrations of taurocholate, a natural non-cell-permeable bile acid, in human hepatic HepG2 and HuH7 cells. In contrast, selective cholesteryl-ester (CE) uptake was increased. Taurocholate exerted these effects extracellularly and independently of HDL modification, cell membrane perturbation or blocking of endocytic trafficking. Instead, this reduction of endocytosis and increase in selective uptake was dependent on SR-BI. In addition, cell-permeable bile acids reduced HDL endocytosis by farnesoid X receptor (FXR) activation: chenodeoxycholate and the non-steroidal FXR agonist GW4064 reduced HDL endocytosis, whereas selective CE uptake was unaltered. Reduced HDL endocytosis by FXR activation was independent of SR-BI and was likely mediated by impaired expression of the scavenger receptor cluster of differentiation 36 (CD36). Taken together we have shown that bile acids reduce HDL endocytosis by transcriptional and non-transcriptional mechanisms. Further, we suggest that HDL endocytosis and selective lipid uptake are not necessarily tightly linked to each other.

  11. Rho/ROCK acts downstream of lysophosphatidic acid receptor 1 in modulating P2X3 receptor-mediated bone cancer pain in rats

    PubMed Central

    Wu, Jing-xiang; Yuan, Xiao-min; Wang, Qiong; Wei, Wang

    2016-01-01

    Background Lysophosphatidic acid receptor 1 and Rho/ROCK signaling is implicated in bone cancer pain development. However, it remains unknown whether the two signaling pathways function together in P2X3 receptor-mediated bone cancer pain. Results In this study, using a rat model of bone cancer, we examined the expression of P2X3 and lysophosphatidic acid receptor 1 in rat dorsal root ganglion neurons and further dissected whether lysophosphatidic acid receptor 1 and Rho/ROCK-mediated pathways interacted in modulating rat pain behavior. Bone cancer was established by inoculating Walker 256 cells into the left tibia of female Wistar rats. We observed a gradual and yet significant decline in mean paw withdrawal threshold in rats with bone cancer, but not in control rats. Our immunohistochemical staining revealed that the number of P2X3- and lysophosphatidic acid receptor 1-positive dorsal root ganglion neurons was significantly greater in rats with bone cancer than control rats. Lysophosphatidic acid receptor 1 blockade with VPC32183 significantly attenuated decline in mean paw withdrawal threshold. Flinching behavior test further showed that lysophosphatidic acid receptor 1 inhibition with VPC32183 transiently but significantly attenuated α,β-meATP-induced increase in paw lift time per minute. Rho inhibition by intrathecal BoTXC3 caused a rapid reversal in decline in mean paw withdrawal threshold of rats with bone cancer. Flinching behavior test showed that BoTXC3 transiently and significantly attenuated α,β-meATP-induced increase in paw lift time per minute. Similar findings were observed with ROCK inhibition by intrathecal Y27632. Furthermore, VPC32183 and BoTXC3 effectively aborted the appearance of lysophosphatidic acid-induced calcium influx peak. Conclusions Lysophosphatidic acid and its receptor LPAR1, acting through the Rho-ROCK pathway, regulate P2X3 receptor in the development of both mechanical and spontaneous pain in bone cancer. PMID:27094551

  12. FGF15/FGFR4 integrates growth factor signaling with hepatic bile acid metabolism and insulin action.

    PubMed

    Shin, Dong-Ju; Osborne, Timothy F

    2009-04-24

    The current studies show FGF15 signaling decreases hepatic forkhead transcription factor 1 (FoxO1) activity through phosphatidylinositol (PI) 3-kinase-dependent phosphorylation. The bile acid receptor FXR (farnesoid X receptor) activates expression of fibroblast growth factor (FGF) 15 in the intestine, which acts through hepatic FGFR4 to suppress cholesterol-7alpha hydroxylase (CYP7A1) and limit bile acid production. Because FoxO1 activity and CYP7A1 gene expression are both increased by fasting, we hypothesized CYP7A1 might be a FoxO1 target gene. Consistent with recently reported results, we show CYP7A1 is a direct target of FoxO1. Additionally, we show that the PI 3-kinase pathway is key for both the induction of CYP7A1 by fasting and the suppression by FGF15. FGFR4 is the major hepatic FGF receptor isoform and is responsible for the hepatic effects of FGF15. We also show that expression of FGFR4 in liver was decreased by fasting, increased by insulin, and reduced by streptozotocin-induced diabetes, implicating FGFR4 as a primary target of insulin regulation. Because insulin and FGF both target the PI 3-kinase pathway, these observations suggest FoxO1 is a key node in the convergence of FGF and insulin signaling pathways and functions as a key integrator for the regulation of glucose and bile acid metabolism.

  13. Ascorbic acid enables reversible dopamine receptor /sup 3/H-agonist binding

    SciTech Connect

    Leff, S.; Sibley, D.R.; Hamblin, M.; Creese, I.

    1981-11-16

    The effects of ascorbic acid on dopaminergic /sup 3/H-agonist receptor binding were studied in membrane homogenates of bovine anterior pituitary and caudate, and rat striatum. In all tissues virtually no stereospecific binding (defined using 1uM (+)butaclamol) of the /sup 3/H-agonists N-propylnorapomorphine (NPA), apomorphine, or dopamine could be demonstrated in the absence of ascorbic acid. Although levels of total /sup 3/H-agonist binding were three to five times greater in the absence than in the presence of 0.1% ascorbic acid, the increased binding was entirely non-stereospecific. Greater amounts of dopamine-inhibitable /sup 3/H-NPA binding could be demonstrated in the absence of 0.1% ascorbic acid, but this measure of ''specific binding'' was demonstrated not to represent dopamine receptor binding since several other catecholamines and catechol were equipotent with dopamine and more potent than the dopamine agonist (+/-)amino-6,7-dihydroxy-1,2,3,4-tetrahydronapthalene (ADTN) in inhibiting this binding. High levels of dopamine-displaceable /sup 3/H-agonist binding were detected in fresh and boiled homogenates of cerebellum, an area of brain which receives no dopaminergic innervation, further demonstrating the non-specific nature of /sup 3/H-agonist binding in the absence of ascorbic acid. These studies emphasize that under typical assay conditions ascorbic acid is required in order to demonstrate reversible and specific /sup 3/H-agonist binding to dopamine receptors.

  14. Ligand specificities of recombinant retinoic acid receptors RAR alpha and RAR beta.

    PubMed Central

    Crettaz, M; Baron, A; Siegenthaler, G; Hunziker, W

    1990-01-01

    Binding of retinoic acid (RA) to specific RA receptors alpha and beta (RAR alpha and RAR beta) was studied. Receptors were obtained in two ways: (1) full-length receptors were produced by transient expression of the respective human cDNAs in COS 1 cells; and (2) the ligand-binding domains of RAR alpha and RAR beta were produced in Escherichia coli. RA binding to the wild-type and truncated forms of the receptor was identical for both RAR alpha and RAR beta, indicating that the ligand-binding domains have retained the binding characteristics of the intact receptors. Furthermore, RA bound with the same affinity to both RAR alpha and RAR beta. Only retinoid analogues with an acidic end-group were able to actively bind to both receptors. On measuring the binding of various retinoids, we have found that the properties of the ligand-binding sites of RAR alpha and RAR beta were rather similar. Two retinoid analogues were capable of binding preferentially to either RAR alpha or RAR beta, suggesting that it may be possible to synthesize specific ligands for RAR alpha and RAR beta. PMID:2176462

  15. Oleic acid stimulates system A amino acid transport in primary human trophoblast cells mediated by toll-like receptor 4.

    PubMed

    Lager, Susanne; Gaccioli, Francesca; Ramirez, Vanessa I; Jones, Helen N; Jansson, Thomas; Powell, Theresa L

    2013-03-01

    Obese women have an increased risk to deliver large babies. However, the mechanisms underlying fetal overgrowth in these pregnancies are not well understood. Obese pregnant women typically have elevated circulating lipid levels. We tested the hypothesis that fatty acids stimulate placental amino acid transport, mediated via toll-like receptor 4 (TLR4) and mammalian target of rapamycin (mTOR) signaling pathways. Circulating NEFA levels and placental TLR4 expression were assessed in women with varying prepregnancy body mass index (BMI). The effects of oleic acid on system A and system L amino acid transport, and on the activation of the mTOR (4EBP1, S6K1, rpS6), TLR4 (IĸB, JNK, p38 MAPK), and STAT3 signaling pathways were determined in cultured primary human trophoblast cells. Maternal circulating NEFAs (n = 33), but not placental TLR4 mRNA expression (n = 16), correlated positively with BMI (P < 0.05). Oleic acid increased trophoblast JNK and STAT3 phosphorylation (P < 0.05), whereas mTOR activity was unaffected. Furthermore, oleic acid doubled trophoblast system A activity (P < 0.05), without affecting system L activity. siRNA-mediated silencing of TLR4 expression prevented the stimulatory effect of oleic acid on system A activity. Our data suggest that maternal fatty acids can increase placental nutrient transport via TLR4, thereby potentially affecting fetal growth.

  16. Identification of dehydroabietc acid from Boswellia thurifera resin as a positive GABAA receptor modulator.

    PubMed

    Rueda, Diana C; Raith, Melanie; De Mieri, Maria; Schöffmann, Angela; Hering, Steffen; Hamburger, Matthias

    2014-12-01

    In a two-microelectrode voltage clamp assay with Xenopus laevis oocytes, a petroleum ether extract (100 μg/mL) of the resin of Boswellia thurifera (Burseraceae) potentiated GABA-induced chloride currents (IGABA) through receptors of the subtype α₁β₂γ₂s by 319.8% ± 79.8%. With the aid of HPLC-based activity profiling, three known terpenoids, dehydroabietic acid (1), incensole (2), and AKBA (3), were identified in the active fractions of the extract. Structure elucidation was achieved by means of HR-MS and microprobe 1D/2D NMR spectroscopy. Compound 1 induced significant receptor modulation in the oocyte assay, with a maximal potentiation of IGABA of 397.5% ± 34.0%, and EC₅₀ of 8.7 μM ± 1.3 μM. This is the first report of dehydroabietic acid as a positive GABAA receptor modulator.

  17. Retinoic acids up-regulate functional eosinophil-driving receptor CCR3.

    PubMed

    Ueki, S; Nishikawa, J; Yamauchi, Y; Konno, Y; Tamaki, M; Itoga, M; Kobayashi, Y; Takeda, M; Moritoki, Y; Ito, W; Chihara, J

    2013-07-01

    Eotaxins and their receptor CCR3 have a definitive role for tissue accumulation of eosinophils both under homeostatic and pathologic conditions. However, physiological stimuli that can up-regulate CCR3 in blood-derived human eosinophils have not been recognized. As a prior gene microarray study revealed up-regulation of CCR3 in eosinophils stimulated with retinoic acids (RAs), the expression of functional CCR3 was examined. We found that 9-cis RA and all-trans RA (ATRA) significantly induced surface CCR3 expression regardless of the presence of IL-3 or IL-5. Pharmacological manipulations with receptor-specific agonists and antagonists indicated that retinoic acid receptor-α activation is critical for CCR3 up-regulation. RA-induced CCR3 was associated with its functional capacity, in terms of the calcium mobilization and chemotactic response to eotaxin-1 (CCL11). Our study suggests an important role of vitamin A derivatives in the tissue accumulation of eosinophils.

  18. Treatment of Type 2 Diabetes by Free Fatty Acid Receptor Agonists

    PubMed Central

    Watterson, Kenneth R.; Hudson, Brian D.; Ulven, Trond; Milligan, Graeme

    2014-01-01

    Dietary free fatty acids (FFAs), such as ω-3 fatty acids, regulate metabolic and anti-inflammatory processes, with many of these effects attributed to FFAs interacting with a family of G protein-coupled receptors. Selective synthetic ligands for free fatty acid receptors (FFA1-4) have consequently been developed as potential treatments for type 2 diabetes (T2D). In particular, clinical studies show that Fasiglifam, an agonist of the long-chain FFA receptor, FFA1, improved glycemic control and reduced HbA1c levels in T2D patients, with a reduced risk of hypoglycemia. However, this ligand was removed from clinical trials due to potential liver toxicity and determining if this is a target or a ligand-specific feature is now of major importance. Pre-clinical studies also show that FFA4 agonism increases insulin sensitivity, induces weight loss, and reduces inflammation and the metabolic and anti-inflammatory effects of short chain fatty acids (SCFAs) are linked with FFA2 and FFA3 activation. In this review, we therefore show that FFA receptor agonism is a potential clinical target for T2D treatment and discuss ongoing drug development programs within industry and academia aimed at improving the safety and effectiveness of these potential treatments. PMID:25221541

  19. Binding of retinoic acid receptor heterodimers to DNA. A role for histones NH2 termini.

    PubMed

    Lefebvre, P; Mouchon, A; Lefebvre, B; Formstecher, P

    1998-05-15

    The retinoic acid signaling pathway is controlled essentially through two types of nuclear receptors, RARs and RXRs. Ligand dependent activation or repression of retinoid-regulated genes is dependent on the binding of retinoic acid receptor (RAR)/9-cis-retinoic acid receptor (RXR) heterodimers to retinoic acid response element (RARE). Although unliganded RXR/RAR heterodimers bind constitutively to DNA in vitro, a clear in vivo ligand-dependent occupancy of the RARE present in the RARbeta2 gene promoter has been reported (Dey, A., Minucci, S., and Ozato, K. (1994) Mol. Cell. Biol. 14, 8191-8201). Nucleosomes are viewed as general repressors of the transcriptional machinery, in part by preventing the access of transcription factors to DNA. The ability of hRXRalpha/hRARalpha heterodimers to bind to a nucleosomal template in vitro has therefore been examined. The assembly of a fragment from the RARbeta2 gene promoter, which contains a canonical DR5 RARE, into a nucleosome core prevented hRXRalpha/hRARalpha binding to this DNA, in conditions where a strong interaction is observed with a linear DNA template. However, histone tails removal by limited proteolysis and histone hyperacetylation yielded nucleosomal RAREs able to bind to hRXRalpha/hRARalpha heterodimers. These data establish therefore the role of histones NH2 termini as a major impediment to retinoid receptors access to DNA, and identify histone hyperacetylation as a potential physiological regulator of retinoid-induced transcription.

  20. Identification of diarylsulfonamides as agonists of the free fatty acid receptor 4 (FFA4/GPR120).

    PubMed

    Sparks, Steven M; Chen, Grace; Collins, Jon L; Danger, Dana; Dock, Steven T; Jayawickreme, Channa; Jenkinson, Stephen; Laudeman, Christopher; Leesnitzer, M Anthony; Liang, Xi; Maloney, Patrick; McCoy, David C; Moncol, David; Rash, Vincent; Rimele, Thomas; Vulimiri, Padmaja; Way, James M; Ross, Sean

    2014-07-15

    The exploration of a diarylsulfonamide series of free fatty acid receptor 4 (FFA4/GPR120) agonists is described. This work led to the identification of selective FFA4 agonist 8 (GSK137647A) and selective FFA4 antagonist 39. The in vitro profile of compounds 8 and 39 is presented herein.

  1. Cross-talk between lysophosphatidic acid receptor 1 and tropomyosin receptor kinase A promotes lung epithelial cell migration.

    PubMed

    Nan, Ling; Wei, Jianxin; Jacko, Anastasia M; Culley, Miranda K; Zhao, Jing; Natarajan, Viswanathan; Ma, Haichun; Zhao, Yutong

    2016-02-01

    Lysophosphatidic acid (LPA) is a bioactive lysophospholipid, which plays a crucial role in the regulation of cell proliferation, migration, and differentiation. LPA exerts its biological effects mainly through binding to cell-surface LPA receptors (LPA1-6), which belong to the G protein-coupled receptor (GPCR) family. Recent studies suggest that cross-talk between receptor tyrosine kinases (RTKs) and GPCRs modulates GPCRs-mediated signaling. Tropomyosin receptor kinase A (TrkA) is a RTK, which mediates nerve growth factor (NGF)-induced biological functions including cell migration in neuronal and non-neuronal cells. Here, we show LPA1 transactivation of TrkA in murine lung epithelial cells (MLE12). LPA induced tyrosine phosphorylation of TrkA in both time- and dose-dependent manners. Down-regulation of LPA1 by siRNA transfection attenuated LPA-induced phosphorylation of TrkA, suggesting a cross-talk between LPA1 and TrkA. To investigate the molecular regulation of the cross-talk, we focused on the interaction between LPA1 and TrkA. We found that LPA induced interaction between LPA1 and TrkA. The LPA1/TrkA complex was localized on the plasma membrane and in the cytoplasm. The C-terminus of LPA1 was identified as the binding site for TrkA. Inhibition of TrkA attenuated LPA-induced phosphorylation of TrkA and LPA1 internalization, as well as lung epithelial cell migration. These studies provide a molecular mechanism for the transactivation of TrkA by LPA, and suggest that the cross-talk between LPA1 and TrkA regulates LPA-induced receptor internalization and lung epithelial cell migration.

  2. Amino acid coevolution reveals three-dimensional structure and functional domains of insect odorant receptors.

    PubMed

    Hopf, Thomas A; Morinaga, Satoshi; Ihara, Sayoko; Touhara, Kazushige; Marks, Debora S; Benton, Richard

    2015-01-13

    Insect odorant receptors (ORs) comprise an enormous protein family that translates environmental chemical signals into neuronal electrical activity. These heptahelical receptors are proposed to function as ligand-gated ion channels and/or to act metabotropically as G protein-coupled receptors (GPCRs). Resolving their signalling mechanism has been hampered by the lack of tertiary structural information and primary sequence similarity to other proteins. We use amino acid evolutionary covariation across these ORs to define restraints on structural proximity of residue pairs, which permit de novo generation of three-dimensional models. The validity of our analysis is supported by the location of functionally important residues in highly constrained regions of the protein. Importantly, insect OR models exhibit a distinct transmembrane domain packing arrangement to that of canonical GPCRs, establishing the structural unrelatedness of these receptor families. The evolutionary couplings and models predict odour binding and ion conduction domains, and provide a template for rationale structure-activity dissection.

  3. Insulin receptor aggregation and autophosphorylation in the presence of cationic polyamino acids

    SciTech Connect

    Kohanski, R.A. )

    1989-12-15

    Aggregation and autophosphorylation of the insulin receptor-protein kinase, from cultured 3T3-L1 adipocytes, were studied in the presence of cationic polyamino acids. Poly-L-lysine and poly-L-arginine produced the following effects with the purified receptor: first, the autophosphorylation rate was increased by polycations. Half-maximal stimulation was proportional to polymer length. The rate enhancement was greater at lower ATP concentrations. Second, near-endpoint (equilibrium) autophosphorylation was greater in the presence of the polycations. Polycations inhibited the reverse reaction: ADP + phosphoreceptor yielding ATP + aporeceptor. Third, the (32P)phosphopeptides generated by trypsin digestion of the 32P-beta-subunit, showed that no new autophosphorylation sites resulted from the presence of polycations. Fourth, the polycations, but not insulin, promoted receptor aggregation, and phosphoreceptor aggregated more readily than aporeceptor. Insulin receptor enriched through the wheat germ agglutinin eluate step was compared with purified receptor. Higher concentrations of poly-L-arginine were required to stimulate autophosphorylation and to promote aggregation. Finally, several polycation-dependent substrates present in the wheat germ agglutinin eluate co-aggregated with the insulin receptor. Polycation-stimulated receptor autophosphorylation is linked to a lower KM,app for ATP, but substrate phosphorylation may require the aggregation.

  4. Transport of cationic amino acids by the mouse ecotropic retrovirus receptor.

    PubMed

    Kim, J W; Closs, E I; Albritton, L M; Cunningham, J M

    1991-08-22

    Susceptibility of rodent cells to infection by ecotropic murine leukaemia viruses (MuLV) is determined by binding of the virus envelope to a membrane receptor that has multiple membrane-spanning domains. Cells infected by ecotropic MuLV synthesize envelope protein, gp70, which binds to this receptor, thereby preventing additional infections. The consequences of envelope-MuLV receptor binding for the infected host cell have not been directly determined, partly because the cellular function of the MuLV receptor protein is unknown. Here we report a coincidence in the positions of the first eight putative membrane-spanning domains found in the virus receptor and in two related proteins, the arginine and histidine permeases of Saccharomyces cerevisiae (Fig. 1), but not in any other proteins identified by computer-based sequence comparison of the GenBank data base. Xenopus oocytes injected with receptor-encoding messenger RNA show increased uptake of L-arginine, L-lysine and L-ornithine. The transport properties and the expression pattern of the virus receptor behave in ways previously attributed to y+, the principal transporter of cationic L-amino acids in mammalian cells.

  5. Identification of a Novel Non-retinoid Pan Inverse Agonist of the Retinoic Acid Receptors

    PubMed Central

    Busby, Scott A.; Kumar, Naresh; Kuruvilla, Dana S.; Istrate, Monica A.; Conkright, Juliana J.; Wang, Yongjun; Kamenecka, Theodore M.; Cameron, Michael D.; Roush, William R.; Burris, Thomas P.; Griffin, Patrick R.

    2011-01-01

    Retinoids are potent forms of vitamin A and are involved in a broad range of physiological processes and the pharmacological effects of retinoids are primarily mediated by the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs). Several natural and synthetic RAR modulators have proven to be clinically useful for a number of therapeutic indications including cancer, psoriasis, and diabetes. Unfortunately, these agents lead to a number of significant side effects. Most synthetic retinoid ligands are based on the retinoid scaffold and thus have similarities to the natural ligand with all previously disclosed RAR ligands having a carboxylic acid that makes a critical ionic bridge within the ligand binding domain of the receptors. The potential therapeutic value offered from RAR modulation provides the impetus to identify novel ligands based on unique scaffolds that may offer improved toxicity and pharmacokinetic profiles. Here we describe the identification of an atypical RAR inverse agonist that represents the first non-acid, non-retinoid direct modulator of RAR receptor subfamily. SR-0065 functions as a pan-RAR inverse agonist suppressing the basal activity of RARα, RARβ, and RARγ as well as inhibiting agonist induced RAR activity. SR-0065 treatment enhanced receptor interaction with a peptide representative of the corepressor SMRT and in cells SR-0065 enhances recruitment of SMRT to RARγ. The acid form of SR-0065, SR-1758, was inactive in all assays. Thus, SR-0065 represents a new class of non-acid, non-retinoid RAR modulator that may be used as a point to initiate development of improved RAR-targeted drugs. PMID:21381756

  6. Bile acid dysregulation, gut dysbiosis, and gastrointestinal cancer.

    PubMed

    Tsuei, Jessica; Chau, Thinh; Mills, David; Wan, Yu-Jui Yvonne

    2014-11-01

    Because of increasingly widespread sedentary lifestyles and diets high in fat and sugar, the global diabetes and obesity epidemic continues to grow unabated. A substantial body of evidence has been accumulated which associates diabetes and obesity to dramatically higher risk of cancer development, particularly in the liver and gastrointestinal tract. Additionally, diabetic and obese individuals have been shown to suffer from dysregulation of bile acid (BA) homeostasis and dysbiosis of the intestinal microbiome. Abnormally elevated levels of cytotoxic secondary BAs and a pro-inflammatory shift in gut microbial profile have individually been linked to numerous enterohepatic diseases including cancer. However, recent findings have implicated a detrimental interplay between BA dysregulation and intestinal dysbiosis that promotes carcinogenesis along the gut-liver axis. This review seeks to examine the currently investigated interactions between the regulation of BA metabolism and activity of the intestinal microbiota and how these interactions can drive cancer formation in the context of diabesity. The precarcinogenic effects of BA dysregulation and gut dysbiosis including excessive inflammation, heightened oxidative DNA damage, and increased cell proliferation are discussed. Furthermore, by focusing on the mediatory roles of BA nuclear receptor farnesoid x receptor, ileal transporter apical sodium dependent BA transporter, and G-coupled protein receptor TGR5, this review attempts to connect BA dysregulation, gut dysbiosis, and enterohepatic carcinogenesis at a mechanistic level. A better understanding of the intricate interplay between BA homeostasis and gut microbiome can yield novel avenues to combat the impending rise in diabesity-related cancers.

  7. Bile acid dysregulation, gut dysbiosis, and gastrointestinal cancer

    PubMed Central

    Tsuei, Jessica; Chau, Thinh; Mills, David; Wan, Yu-Jui Yvonne

    2015-01-01

    Because of increasingly widespread sedentary lifestyles and diets high in fat and sugar, the global diabetes and obesity epidemic continues to grow unabated. A substantial body of evidence has been accumulated which associates diabetes and obesity to dramatically higher risk of cancer development, particularly in the liver and gastrointestinal tract. Additionally, diabetic and obese individuals have been shown to suffer from dysregulation of bile acid (BA) homeostasis and dysbiosis of the intestinal microbiome. Abnormally elevated levels of cytotoxic secondary BAs and a pro-inflammatory shift in gut microbial profile have individually been linked to numerous enterohepatic diseases including cancer. However, recent findings have implicated a detrimental interplay between BA dysregulation and intestinal dysbiosis that promotes carcinogenesis along the gut–liver axis. This review seeks to examine the currently investigated interactions between the regulation of BA metabolism and activity of the intestinal microbiota and how these interactions can drive cancer formation in the context of diabesity. The precarcinogenic effects of BA dysregulation and gut dysbiosis including excessive inflammation, heightened oxidative DNA damage, and increased cell proliferation are discussed. Furthermore, by focusing on the mediatory roles of BA nuclear receptor farnesoid x receptor, ileal transporter apical sodium dependent BA transporter, and G-coupled protein receptor TGR5, this review attempts to connect BA dysregulation, gut dysbiosis, and enterohepatic carcinogenesis at a mechanistic level. A better understanding of the intricate interplay between BA homeostasis and gut microbiome can yield novel avenues to combat the impending rise in diabesity-related cancers. PMID:24951470

  8. Acidic stimuli activates two distinct pathways in taste receptor cells from rat fungiform papillae.

    PubMed

    Liu, L; Simon, S A

    2001-12-27

    A sour taste sensation may be produced when acidic stimuli interact with taste receptor cells (TRCs) on the dorsal surface of the tongue. We have searched for pathways in TRCs that may be activated by acidic stimuli using RT-PCR and changes in intracellular calcium (Ca(2+)(I)) induced by acidic stimuli in rat fungiform papillae. RT-PCR revealed the presence of proton-gated subunits ASIC-beta and VR1. Ca(2+) imaging measurements of the TRCs revealed two distinct responses to acidic stimuli: Ca(2+)(i) was increased in 9% (28/308; Type I) and was decreased in 39% (121/308; Type II). Neither of these responses was affected by the removal of extracellular Ca(2+), indicating that the changes arise from the release and sequestration of Ca(2+) from intracellular stores. These responses were also not inhibited by the vanilloid receptor antagonist, capsazepine, suggesting they do not arise from the activation of vanilloid receptors. The Type I, but not the Type II response was inhibited by amiloride. Dose-response measurements for Types I and II responses yielded pH(50%) of 4.8 and 4.9, respectively. Type II responses were inhibited by pertussis toxin, suggesting G-protein involvement. TRCs that exhibit Type II responses could also be activated by quinine (which increased Ca(2+)(I)) thus suggesting a mechanism by which the addition of acid may be suppressive to other chemical stimuli.

  9. Tetrahydro-iso-alpha Acids Antagonize Estrogen Receptor Alpha Activity in MCF-7 Breast Cancer Cells.

    PubMed

    Lempereur, Maëlle; Majewska, Claire; Brunquers, Amandine; Wongpramud, Sumalee; Valet, Bénédicte; Janssens, Philippe; Dillemans, Monique; Van Nedervelde, Laurence; Gallo, Dominique

    2016-01-01

    Tetrahydro-iso-alpha acids commonly called THIAA or Tetra are modified hop acids extracted from hop (Humulus lupulus L.) which are frequently used in brewing industry mainly in order to provide beer bitterness and foam stability. Interestingly, molecular structure of tetrahydro-iso-alpha acids is close to a new type of estrogen receptor alpha (ERα) antagonists aimed at disrupting the binding of coactivators containing an LxxLL motif (NR-box). In this work we show that THIAA decreases estradiol-stimulated proliferation of MCF-7 (ERα-positive breast cancer cells). Besides, we show that it inhibits ERα transcriptional activity. Interestingly, this extract fails to compete with estradiol for ERα binding and does not significantly impact the receptor turnover rate in MCF-7 cells, suggesting that it does not act like classical antiestrogens. Hence, we demonstrate that THIAA is able to antagonize ERα estradiol-induced recruitment of the LxxLL binding motif.

  10. Tetrahydro-iso-alpha Acids Antagonize Estrogen Receptor Alpha Activity in MCF-7 Breast Cancer Cells

    PubMed Central

    Lempereur, Maëlle; Majewska, Claire; Brunquers, Amandine; Wongpramud, Sumalee; Valet, Bénédicte; Janssens, Philippe; Dillemans, Monique; Van Nedervelde, Laurence; Gallo, Dominique

    2016-01-01

    Tetrahydro-iso-alpha acids commonly called THIAA or Tetra are modified hop acids extracted from hop (Humulus lupulus L.) which are frequently used in brewing industry mainly in order to provide beer bitterness and foam stability. Interestingly, molecular structure of tetrahydro-iso-alpha acids is close to a new type of estrogen receptor alpha (ERα) antagonists aimed at disrupting the binding of coactivators containing an LxxLL motif (NR-box). In this work we show that THIAA decreases estradiol-stimulated proliferation of MCF-7 (ERα-positive breast cancer cells). Besides, we show that it inhibits ERα transcriptional activity. Interestingly, this extract fails to compete with estradiol for ERα binding and does not significantly impact the receptor turnover rate in MCF-7 cells, suggesting that it does not act like classical antiestrogens. Hence, we demonstrate that THIAA is able to antagonize ERα estradiol-induced recruitment of the LxxLL binding motif. PMID:27190515

  11. Metformin interferes with bile acid homeostasis through AMPK-FXR crosstalk

    PubMed Central

    Lien, Fleur; Berthier, Alexandre; Bouchaert, Emmanuel; Gheeraert, Céline; Alexandre, Jeremy; Porez, Geoffrey; Prawitt, Janne; Dehondt, Hélène; Ploton, Maheul; Colin, Sophie; Lucas, Anthony; Patrice, Alexandre; Pattou, François; Diemer, Hélène; Van Dorsselaer, Alain; Rachez, Christophe; Kamilic, Jelena; Groen, Albert K.; Staels, Bart; Lefebvre, Philippe

    2014-01-01

    The nuclear bile acid receptor farnesoid X receptor (FXR) is an important transcriptional regulator of bile acid, lipid, and glucose metabolism. FXR is highly expressed in the liver and intestine and controls the synthesis and enterohepatic circulation of bile acids. However, little is known about FXR-associated proteins that contribute to metabolic regulation. Here, we performed a mass spectrometry–based search for FXR-interacting proteins in human hepatoma cells and identified AMPK as a coregulator of FXR. FXR interacted with the nutrient-sensitive kinase AMPK in the cytoplasm of target cells and was phosphorylated in its hinge domain. In cultured human and murine hepatocytes and enterocytes, pharmacological activation of AMPK inhibited FXR transcriptional activity and prevented FXR coactivator recruitment to promoters of FXR-regulated genes. Furthermore, treatment with AMPK activators, including the antidiabetic biguanide metformin, inhibited FXR agonist induction of FXR target genes in mouse liver and intestine. In a mouse model of intrahepatic cholestasis, metformin treatment induced FXR phosphorylation, perturbed bile acid homeostasis, and worsened liver injury. Together, our data indicate that AMPK directly phosphorylates and regulates FXR transcriptional activity to precipitate liver injury under conditions favoring cholestasis. PMID:24531544

  12. Bile acids and nonalcoholic fatty liver disease: Molecular insights and therapeutic perspectives

    PubMed Central

    Arab, Juan P.; Karpen, Saul J.; Dawson, Paul A.

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) is a burgeoning health problem worldwide and an important risk factor for both hepatic and cardiometabolic mortality. The rapidly increasing prevalence of this disease and of its aggressive form nonalcoholic steatohepatitis (NASH) will require novel therapeutic approaches to prevent disease progression to advanced fibrosis or cirrhosis and cancer. In recent years, bile acids have emerged as relevant signaling molecules that act at both hepatic and extrahepatic tissues to regulate lipid and carbohydrate metabolic pathways as well as energy homeostasis. Activation or modulation of bile acid receptors, such as the farnesoid X receptor and TGR5, and transporters, such as the ileal apical sodium‐dependent bile acid transporter, appear to affect both insulin sensitivity and NAFLD/NASH pathogenesis at multiple levels, and these approaches hold promise as novel therapies. In the present review, we summarize current available data on the relationships of bile acids to NAFLD and the potential for therapeutically targeting bile‐acid‐related pathways to address this growing world‐wide disease. (Hepatology 2017;65:350‐362) PMID:27358174

  13. The lactate receptor, G-protein-coupled receptor 81/hydroxycarboxylic acid receptor 1: Expression and action in brain.

    PubMed

    Morland, Cecilie; Lauritzen, Knut Husø; Puchades, Maja; Holm-Hansen, Signe; Andersson, Krister; Gjedde, Albert; Attramadal, Håvard; Storm-Mathisen, Jon; Bergersen, Linda Hildegard

    2015-07-01

    We have proposed that lactate is a "volume transmitter" in the brain and underpinned this by showing that the lactate receptor, G-protein-coupled receptor 81 (GPR81, also known as HCA1 or HCAR1), which promotes lipid storage in adipocytes, is also active in the mammalian brain. This includes the cerebral neocortex and the hippocampus, where it can be stimulated by physiological concentrations of lactate and by the HCAR1 agonist 3,5-dihydroxybenzoate to reduce cAMP levels. Cerebral HCAR1 is concentrated on the postsynaptic membranes of excitatory synapses and also is enriched at the blood-brain barrier. In synaptic spines and in adipocytes, HCAR1 immunoreactivity is also located on subplasmalemmal vesicular organelles, suggesting trafficking to and from the plasma membrane. Through activation of HCAR1, lactate can act as a volume transmitter that links neuronal activity, cerebral blood flow, energy metabolism, and energy substrate availability, including a glucose- and glycogen-saving response. HCAR1 may contribute to optimizing the cAMP concentration. For instance, in the prefrontal cortex, excessively high cAMP levels are implicated in impaired cognition in old age, fatigue, stress, and schizophrenia and in the deposition of phosphorylated tau protein in Alzheimer's disease. HCAR1 could serve to ameliorate these conditions and might also act through downstream mechanisms other than cAMP. Lactate exits cells through monocarboxylate transporters in an equilibrating manner and through astrocyte anion channels activated by depolarization. In addition to locally produced lactate, lactate produced by exercising muscle as well as exogenous HCAR1 agonists, e.g., from fruits and berries, might activate the receptor on cerebral blood vessels and brain cells.

  14. Novel protective role of the circadian nuclear receptor retinoic acid-related orphan receptor-α in diabetic cardiomyopathy.

    PubMed

    Zhao, Yichao; Xu, Longwei; Ding, Song; Lin, Nan; Ji, Qingqi; Gao, Lingchen; Su, Yuanyuan; He, Ben; Pu, Jun

    2017-04-01

    Diabetic cardiomyopathy is a major complication that significantly contributes to morbidity and mortality in diabetics with few therapies. Moreover, antidiabetic drugs reported inconsistent or even adverse cardiovascular effects, suggesting that it is important to exploit novel therapeutic targets against diabetic cardiomyopathy. Here, we observed that the nuclear melatonin receptor, the retinoic acid-related orphan receptor-α (RORα), was downregulated in diabetic hearts. By utilizing a mouse line with RORα disruption, we demonstrated that RORα deficiency led to significantly augmented diastolic dysfunction and cardiac remodeling induced by diabetes. Microscopic and molecular analyses further indicated that the detrimental effects of RORα deficiency were associated with aggravated myocardial apoptosis, autophagy dysfunction, and oxidative stress by disrupting antioxidant gene expression. By contrast, restoration of cardiac RORα levels in transgenic mice significantly improved cardiac functional and structural parameters at 8 weeks after diabetes induction. Consistent with genetic manipulation, pharmacological activation of RORα by melatonin and SR1078 (a synthetic agonist) showed beneficial effects against diabetic cardiomyopathy, while the RORα inhibitor SR3335 significantly exacerbated cardiac impairments in diabetic mice. Collectively, these findings suggest that cardiac-targeted manipulation of nuclear melatonin receptor RORα may hold promise for delaying diabetic cardiomyopathy development.

  15. Evidence for impaired retinoic acid receptor-thyroid hormone receptor AF-2 cofactor activity in human lung cancer.

    PubMed Central

    Moghal, N; Neel, B G

    1995-01-01

    Retinoic acid (RA) is required for normal airway epithelial cell growth and differentiation both in vivo and in vitro. One of the earliest events following the exposure of bronchial epithelial cells to RA is the strong induction of RA receptor beta (RAR beta) mRNA. Previous work established that many lung cancer cell lines and primary tumors display abnormal RAR beta mRNA expression, most often absence or weak expression of the RAR beta 2 isoform, even after RA treatment. Restoration of RAR beta 2 into RAR beta-negative lung cancer cell lines has been reported to inhibit tumorigenicity. Since RAR beta 2 inactivation may contribute to lung cancer, we have investigated the molecular mechanism of defective RAR beta 2 expression. Nuclear run-on assays and transient transfections with RAR beta 2 promoter constructs indicate the presence of trans-acting transcriptional defects in most lung cancer cell lines, which map to the RA response element (RARE). These defects cannot be complemented by RAR-retinoid X receptor cotransfection and can be separated into two types: (i) one affecting transcription from direct repeat RAREs, but not palindromic RAREs, and (ii) another affecting transcription from both types of RARE. Studies using chimeras between RAR alpha, TR alpha, and other transcription factors suggest the existence of novel RAR-thyroid hormone receptor AF-2-specific cofactors, which are necessary for high levels of transcription. Furthermore, these factors may be frequently inactivated in human lung cancer. PMID:7791800

  16. (3-Aminocyclopentyl)methylphosphinic acids: novel GABA(C) receptor antagonists.

    PubMed

    Chebib, Mary; Hanrahan, Jane R; Kumar, Rohan J; Mewett, Kenneth N; Morriss, Gwendolyn; Wooller, Soraya; Johnston, Graham A R

    2007-03-01

    Our understanding of the role GABA(C) receptors play in the central nervous system is limited due to a lack of specific ligands. Here we describe the pharmacological effects of (+/-)-cis-3- and (+/-)-trans-3-(aminocyclopentyl)methylphosphinic acids ((+/-)-cis- and (+/-)-trans-3-ACPMPA) as novel ligands for the GABA(C) receptor showing little activity at GABA(A) or GABA(B) receptors. (+/-)-cis-3-ACPMPA has similar potency to (1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid (TPMPA) at human recombinant rho1 (K(B)=1.0+/-0.2microM) and rat rho3 (K(B)=5.4+/-0.8microM) but is 15 times more potent than TPMPA on human recombinant rho2 (K(B)=1.0+/-0.3microM) GABA(C) receptors expressed in Xenopus oocytes. (+/-)-cis- and (+/-)-trans-3-ACPMPA are novel lead compounds for developing into more potent and selective GABA(C) receptor antagonists with increased lipophilicity for in vivo studies.

  17. Signaling through retinoic acid receptors in cardiac development: Doing the right things at the right times.

    PubMed

    Xavier-Neto, José; Sousa Costa, Ângela M; Figueira, Ana Carolina M; Caiaffa, Carlo Donato; Amaral, Fabio Neves do; Peres, Lara Maldanis Cerqueira; da Silva, Bárbara Santos Pires; Santos, Luana Nunes; Moise, Alexander R; Castillo, Hozana Andrade

    2015-02-01

    Retinoic acid (RA) is a terpenoid that is synthesized from vitamin A/retinol (ROL) and binds to the nuclear receptors retinoic acid receptor (RAR)/retinoid X receptor (RXR) to control multiple developmental processes in vertebrates. The available clinical and experimental data provide uncontested evidence for the pleiotropic roles of RA signaling in development of multiple embryonic structures and organs such eyes, central nervous system, gonads, lungs and heart. The development of any of these above-mentioned embryonic organ systems can be effectively utilized to showcase the many strategies utilized by RA signaling. However, it is very likely that the strategies employed to transfer RA signals during cardiac development comprise the majority of the relevant and sophisticated ways through which retinoid signals can be conveyed in a complex biological system. Here, we provide the reader with arguments indicating that RA signaling is exquisitely regulated according to specific phases of cardiac development and that RA signaling itself is one of the major regulators of the timing of cardiac morphogenesis and differentiation. We will focus on the role of signaling by RA receptors (RARs) in early phases of heart development. This article is part of a Special Issue entitled: Nuclear receptors in animal development.

  18. Neurochemical, pharmacological, and developmental studies on cerebellar receptors for dicarboxylic amino acids

    SciTech Connect

    Sharif, N.A.; Roberts, P.J.

    1984-01-01

    Specific binding of L-(/sup 3/H)glutamate ((/sup 3/H)Glu) and L(/sup 3/H)Asp) to cerebellar membranes represented a time-, temperature-, pH- and protein-dependent interaction which was both saturable and reversible. Binding sites for both radioligands appeared maximally enriched in synaptosomal fractions isolated by gradient centrifugation. Kinetically derived dissociation constant (K/sub off//K/sub on/ . K/sub d/) for (/sup 3/H)Glu binding to this fraction indicated high-affinity (433 nM). Competition experiments employing analogs of excitatory amino acids, including new antagonists, helped identify binding sites for (/sup 3/H)Glu and (/sup 3/H)Asp as receptors with differential pharmacological specificities. Membrane freezing reduced numbers of both receptor types, but binding activity could be recovered partially by incubation at 37 degrees C. Glu receptors exhibited a pronounced deleterious sensitivity to thiol modifying reagents and L-Glu (50-1000 microM) provided protection against these compounds during co-incubation with cerebellar membranes. It is suggested that cold storage may induce partially reversible receptor inactivation by promoting sulfhydryl group/bond modification. Rat cerebellar glutamatergic function (endogenous Glu content, Glu uptake and receptor sites) exhibited an apparent ontogenetic peak between days 8-12 postpartum with a plateauing profile from day 30 to adulthood. The accelerated development (days 8-12) coincides with the first demonstrable Glu release and kainic acid neurotoxicity, as described previously.

  19. Endocytosis of lysosomal acid phosphatase; involvement of mannose receptor and effect of lectins.

    PubMed

    Imai, K; Yoshimura, T

    1994-08-01

    Acid phosphatase and beta-glucosidase are unique among lysosomal enzymes in that they have both high mannose and complex type sugasr chains, whereas oligosaccharide chains of lysosomal enzymes in matrix are of high mannose type. We have previously shown that beta-glucosidase was endocytosed into macrophages via an unidentified receptor different from a mannose/fucose receptor (K. Imai, Cell Struct. Funct. 13, 325-332, 1988). Here, we show that uptake of acid phosphatase purified from rat liver lysosomes into rat macrophages was inhibited by ligands for a mannose/fucose receptor and was mediated via an apparently single binding site with Kuptake of 24.7 nM. These results indicate that acid phosphatase and beta-glucosidase recognize different types of receptors even if they have similar sugar chains. Polyvalent concanavalin A which binds both to the enzyme and to macrophages specifically stimulated the uptake in a dose dependent manner, whereas wheat germ agglutinin and phytohaemagglutinin did not.

  20. Guanidino acids act as rho1 GABA(C) receptor antagonists.

    PubMed

    Chebib, Mary; Gavande, Navnath; Wong, Kit Yee; Park, Anna; Premoli, Isabella; Mewett, Kenneth N; Allan, Robin D; Duke, Rujee K; Johnston, Graham A R; Hanrahan, Jane R

    2009-10-01

    GABA(C) receptors play a role in myopia, memory-related disorders and circadian rhythms signifying a need to develop potent and selective agents for this class of receptors. Guanidino analogs related to glycine, beta-alanine and taurine were evaluated at human rho(1)GABA(C) receptors expressed in Xenopus oocytes using 2-electrode voltage clamp methods. Of the 12 analogs tested, 8 analogs were active as antagonists and the remaining were inactive. (S)-2-guanidinopropionic acid (IC(50) = 2.2 microM) and guanidinoacetic acid (IC(50) = 5.4 microM; K (B) = 7.75 microM [pK (B) = 5.11 +/- 0.06]) were the most potent being competitive antagonists at this receptor. In contrast, the beta-alanine and GABA guanidino analogs showed reduced activity, indicating the distance between the carboxyl carbon and terminal nitrogen of the guanidino group is critical for activity. Substituting the C2-position of guanidinoacetic acid with various alkyl groups reduced activity indicating that steric effects may impact on activity. The results of this study contribute to the structure-activity-relationship profile required in developing novel therapeutic agents.

  1. Computer-aided design of a novel ligand for retinoic acid receptor in cancer chemotherapy

    NASA Astrophysics Data System (ADS)

    Silva, Carlos H. T. P.; Leopoldino, Andreia M.; Silva, Eloiza H. T.; Espinoza, V. A. A.; Taft, C. A.

    The isotypes of RAR and RXR are retinoic acid and retinoid X acid receptors, respectively, whose ligand-binding domain contains the ligand-dependent activation function, with distinct pharmacological targets for retinoids, involved in the treatment of various cancers and skin diseases. Due to the major challenge which cancer treatment and cure still imposes after many decades to the international scientific community, there is actually considerable interest in new ligands with increased bioactivity. We have focused on the retinoid acid receptor, which is considered an interesting target for drug design. In this work, we carried out density functional geometry optimizations and different docking procedures. We performed screening in a large database (hundreds of thousands of molecules which we optimized at the AM1 level) yielding a set of potential bioactive ligands. A new ligand was selected and optimized at the B3LYP/6-31G* level. A flexible docking program was used to investigate the interactions between the receptor and the new ligand. The result of this work is compared with several crystallographic ligands of RAR. Our theoretically more bioactive new ligand indicates stronger and more hydrogen bonds as well as hydrophobic interactions with the receptor.

  2. Conformations, energies, and intramolecular hydrogen bonds in dicarboxylic acids: implications for the design of synthetic dicarboxylic acid receptors.

    PubMed

    Nguyen, Thanh Ha; Hibbs, David E; Howard, Siân T

    2005-09-01

    The various conformers of the dicarboxylic acids HO2C--(CH2)n--CO2H, n = 1-4, were obtained using density functional methods (DFT), both in the gas phase and in the aqueous phase using a polarized continuum model (PCM). Several new conformers were identified, particularly for the two larger molecules glutaric (n = 3) and adipic acid (n =4). The PCM results show that the stability of most conformers were affected, many becoming unstable in the aqueous phase; and the energy ordering of conformers is also different. The results suggest that conformational preferences could be important in determining the design and stability of appropriate synthetic receptors for glutaric and adipic acid. Geometry changes between gas and aqueous phases were most marked in those conformers containing an intramolecular hydrogen bond. Additional calculations have probed the strength of intramolecular hydrogen bonds in these dicarboxylic acids. In the cases of glutaric and adipic acid, the strength of the intramolecular hydrogen bond were estimated to be around 28-29 kJ/mol, without any vibrational energy correction. The intramolecular hydrogen bond energies in malonic and succinic acid were also estimated from the calculated H-bond distances using an empirical relationship. Intramolecular H-bond redshifts of 170-250 cm(-1) have been estimated from the results of the harmonic frequency analyses.

  3. PLZF is a negative regulator of retinoic acid receptor transcriptional activity.

    PubMed

    Martin, Perrine J; Delmotte, Marie-Hélène; Formstecher, Pierre; Lefebvre, Philippe

    2003-09-06

    BACKGROUND: Retinoic acid receptors (RARs) are ligand-regulated transcription factors controlling cellular proliferation and differentiation. Receptor-interacting proteins such as corepressors and coactivators play a crucial role in specifying the overall transcriptional activity of the receptor in response to ligand treatment. Little is known however on how receptor activity is controlled by intermediary factors which interact with RARs in a ligand-independent manner. RESULTS: We have identified the promyelocytic leukemia zinc finger protein (PLZF), a transcriptional corepressor, to be a RAR-interacting protein using the yeast two-hybrid assay. We confirmed this interaction by GST-pull down assays and show that the PLZF N-terminal zinc finger domain is necessary and sufficient for PLZF to bind RAR. The RAR ligand binding domain displayed the highest affinity for PLZF, but corepressor and coactivator binding interfaces did not contribute to PLZF recruitment. The interaction was ligand-independent and correlated to a decreased transcriptional activity of the RXR-RAR heterodimer upon overexpression of PLZF. A similar transcriptional interference could be observed with the estrogen receptor alpha and the glucocorticoid receptor. We further show that PLZF is likely to act by preventing RXR-RAR heterodimerization, both in-vitro and in intact cells. CONCLUSION: Thus RAR and PLZF interact physically and functionally. Intriguingly, these two transcription factors play a determining role in hematopoiesis and regionalization of the hindbrain and may, upon chromosomal translocation, form fusion proteins. Our observations therefore define a novel mechanism by which RARs activity may be controlled.

  4. Synthesis of C5-tetrazole derivatives of 2-amino-adipic acid displaying NMDA glutamate receptor antagonism.

    PubMed

    Lenda, Fatimazohra; Crouzin, Nadine; Cavalier, Mélanie; Guiramand, Janique; Lanté, Fabien; Barbanel, Gérard; Cohen-Solal, Catherine; Martinez, Jean; Guenoun, Farhate; Lamaty, Frédéric; Vignes, Michel

    2011-03-01

    Five derivatives of 2-amino-adipic acid bearing a tetrazole-substituted in C5 position were synthesized. These compounds displayed selective antagonism towards N-methyl-D: -aspartate (NMDA) receptors compared with AMPA receptors, and they were devoid of any neurotoxicity. Among these five analogues, one exhibited a higher affinity for synaptic NMDA responses than the other four. Therefore, C5 tetrazole-substituted of 2-amino-adipic acid represent an interesting series of new NMDA receptor antagonists. This approach may be considered as a new strategy to develop ligands specifically targeted to synaptic or extra-synaptic NMDA receptors.

  5. Lysergic acid diethylamide-induced Fos expression in rat brain: role of serotonin-2A receptors.

    PubMed

    Gresch, P J; Strickland, L V; Sanders-Bush, E

    2002-01-01

    Lysergic acid diethylamide (LSD) produces altered mood and hallucinations in humans and binds with high affinity to serotonin-2A (5-HT(2A)) receptors. Although LSD interacts with other receptors, the activation of 5-HT(2A) receptors is thought to mediate the hallucinogenic properties of LSD. The goal of this study was to identify the brain sites activated by LSD and to determine the influence of 5-HT(2A) receptors in this activation. Rats were pretreated with the 5-HT(2A) receptor antagonist MDL 100907 (0.3 mg/kg, i.p.) or vehicle 30 min prior to LSD (500 microg/kg, i.p.) administration and killed 3 h later. Brain tissue was examined for Fos protein expression by immunohistochemistry. LSD administration produced a five- to eight-fold increase in Fos-like immunoreactivity in medial prefrontal cortex, anterior cingulate cortex, and central nucleus of amygdala. However, in dorsal striatum and nucleus accumbens no increase in Fos-like immunoreactivity was observed. Pretreatment with MDL 100907 completely blocked LSD-induced Fos-like immunoreactivity in medial prefrontal cortex and anterior cingulate cortex, but only partially blocked LSD-induced Fos-like immunoreactivity in amygdala. Double-labeled immunohistochemistry revealed that LSD did not induce Fos-like immunoreactivity in cortical cells expressing 5-HT(2A) receptors, suggesting an indirect activation of cortical neurons. These results indicate that the LSD activation of medial prefrontal cortex and anterior cingulate cortex is mediated by 5-HT(2A) receptors, whereas in amygdala 5-HT(2A) receptor activation is a component of the response. These findings support the hypothesis that the medial prefrontal cortex, anterior cingulate cortex, and perhaps the amygdala, are important regions involved in the production of hallucinations.

  6. Novel bile acid therapeutics for the treatment of chronic liver diseases

    PubMed Central

    Hegade, Vinod S.; Speight, R. Alexander; Etherington, Rachel E.; Jones, David E. J.

    2016-01-01

    Recent developments in understanding the role of bile acids (BAs) as signalling molecules in human metabolism and inflammation have opened new avenues in the field of hepatology research. BAs are no longer considered as simple molecules helping in fat digestion but as agents with real therapeutic value in treating complex autoimmune and metabolic liver diseases. BAs and their receptors such as farnesoid X receptor, transmembrane G protein-coupled receptor 5 and peroxisome proliferator-activated receptor have been identified as novel targets for drug development. Some of these novel pharmaceuticals are already in clinical evaluation with the most advanced drugs having reached phase III trials. Chronic liver diseases such as primary biliary cholangitis, primary sclerosing cholangitis and nonalcoholic fatty liver disease, for which there is no or limited pharmacotherapy, are most likely to gain from these developments. In this review we discuss recent and the most relevant basic and clinical research findings related to BAs and their implications for novel therapy for chronic liver diseases. PMID:27134666

  7. Hydroxycarboxylic acid receptors are essential for breast cancer cells to control their lipid/fatty acid metabolism.

    PubMed

    Stäubert, Claudia; Broom, Oliver Jay; Nordström, Anders

    2015-08-14

    Cancer cells exhibit characteristic changes in their metabolism with efforts being made to address them therapeutically. However, targeting metabolic enzymes as such is a major challenge due to their essentiality for normal proliferating cells. The most successful pharmaceutical targets are G protein-coupled receptors (GPCRs), with more than 40% of all currently available drugs acting through them.We show that, a family of metabolite-sensing GPCRs, the Hydroxycarboxylic acid receptor family (HCAs), is crucial for breast cancer cells to control their metabolism and proliferation.We found HCA1 and HCA3 mRNA expression were significantly increased in breast cancer patient samples and detectable in primary human breast cancer patient cells. Furthermore, siRNA mediated knock-down of HCA3 induced considerable breast cancer cell death as did knock-down of HCA1, although to a lesser extent. Liquid Chromatography Mass Spectrometry based analyses of breast cancer cell medium revealed a role for HCA3 in controlling intracellular lipid/fatty acid metabolism. The presence of etomoxir or perhexiline, both inhibitors of fatty acid β-oxidation rescues breast cancer cells with knocked-down HCA3 from cell death.Our data encourages the development of drugs acting on cancer-specific metabolite-sensing GPCRs as novel anti-proliferative agents for cancer therapy.

  8. Mercaptoacetate blocks fatty acid-induced GLP-1 secretion in male rats by directly antagonizing GPR40 fatty acid receptors.

    PubMed

    Li, Ai-Jun; Wang, Qing; Dinh, Thu T; Simasko, Steve M; Ritter, Sue

    2016-04-15

    Mercaptoacetate (MA) is an orexigenic agent reported to block fatty acid (FA) oxidation. Recently, however, we reported evidence from isolated nodose ganglion neurons that MA antagonizes the G protein-coupled long- and medium-chain FA receptor GPR40. GPR40 mediates FA-induced secretion of the satietogenic incretin peptide glucagon-like peptide 1 (GLP-1), by enteroendocrine L cells, as well as FA-induced enhancement of glucose-stimulated insulin secretion. Our results in cultured nodose neurons suggest that MA would also block GPR40 in enteroendocrine cells controlling GLP-1 secretion. If so, this would suggest an alternative mechanism by which MA increases food intake. We tested the hypothesis that MA blocks FA-induced GLP-1 secretion in vitro using cultured STC-1 cells (a murine enteroendocrine cell line) and in vivo in adult male rats. In vitro, MA blocked the increase in both cytosolic Ca(2+)and GLP-1 release stimulated by FAs and also reduced (but less effectively) the response of STC-1 cells to grifolic acid, a partial agonist of the GPR120 FA receptor. In vivo, MA reduced GLP-1 secretion following olive oil gavage while also increasing glucose and decreasing insulin levels. The carnitine palmatoyltransferase 1 antagonist etomoxir did not alter these responses. Results indicate that MA's actions, including its orexigenic effect, are mediated by GPR40 (and possibly GPR120) receptor antagonism and not by blockade of fat oxidation, as previously believed. Analysis of MA's interaction with GPR40 may facilitate understanding of the multiple functions of this receptor and the manner in which FAs participate in the control of hunger and satiety.

  9. Taurine activates glycine and gamma-aminobutyric acid A receptors in rat substantia gelatinosa neurons.

    PubMed

    Wu, Jun; Kohno, Tatsuro; Georgiev, Stefan K; Ikoma, Miho; Ishii, Hideaki; Petrenko, Andrey B; Baba, Hiroshi

    2008-02-12

    Taurine has been suggested to modulate nociceptive information at the spinal cord level. In this study, the pharmacological properties of taurine were investigated in adult rat substantia gelatinosa (SG) neurons using whole-cell patch-clamp method. We found that taurine seemed to have higher efficacy than glycine on glycine receptors in SG neurons. An increase in chloride conductance was responsible for taurine-induced currents. Taurine at 0.3 mM activated glycine receptors, whereas at 3 mM activated both glycine and gamma-aminobutyric acid A receptors. The currents activated by coapplication of taurine and glycine are cross inhibitive. Altogether these results show that taurine might represent another important neurotransmitter or modulator in SG neurons, which may be involved in antinociception.

  10. Effect of niflumic acid on electromechanical coupling by tachykinin NK1 receptor activation in rabbit colon.

    PubMed

    Patacchini, R; Santicioli, P; Maggi, C A

    1996-05-15

    We have investigated the effect of the Cl- channel blocker, niflumic acid, on the contractile response and electromechanical coupling activated by stimulation of the tachykinin NK1 receptor in the longitudinal muscle of rabbit proximal colon, in the presence of indomethacin (5 microM). The application of submaximal equieffective concentrations of the tachykinin NK1 receptor-selective agonist [Sar9]substance P sulfone (30 nM), of carbachol (300 nM) and KCl (40 mM), produced distinct phasic and tonic components of contraction. Niflumic acid (10-100 microM) preferentially and markedly inhibited the tonic component of the response to [Sar9]substance P sulfone and to carbachol, without affecting the response to KCl. Nifedipine (1 microM) abolished the response to KCl and greatly reduced the response to [Sar9]substance P sulfone and carbachol. The nifedipine-resistant response to [Sar9]substance P sulfone was attenuated by niflumic acid (100 microM), while that to carbachol was unaffected. In sucrose gap experiments, superfusion with niflumic acid (100 microM), in the presence of nifedipine (3 microM), produced membrane hyperpolarization, which was totally blocked by tetraethylammonium (10 mM). Niflumic acid inhibited both depolarization and contraction induced by [Sar9]substance P sulfone, both in the absence or in the presence of tetraethylammonium. The present findings support the idea that a niflumic acid-sensitive mechanism, probably an effect on Cl- channels, takes part in the post-receptorial events activated by tachykinin NK1 receptor stimulation in the longitudinal muscle of rabbit colon, and suggest that this mechanism would be more important for generating the sustained tonic than the phasic component of contraction.

  11. SIGNALLING THROUGH RETINOIC ACID RECEPTORS IN CARDIAC DEVELOPMENT: DOING THE RIGHT THINGS AT THE RIGHT TIMES

    PubMed Central

    Xavier-Neto, José; Costa, Ângela M. Sousa; Figueira, Ana Carolina M.; Caiaffa, Carlo Donato; do Amaral, Fabio Neves; Peres, Lara Maldanis Cerqueira; da Silva, Bárbara Santos Pires; Santos, Luana Nunes; Moise, Alexander R.; Castillo, Hozana Andrade

    2015-01-01

    Retinoic acid (RA) is a terpenoid that is synthesized from Vitamin A/retinol (ROL) and binds to the nuclear receptors retinoic acid receptor (RAR)/retinoid X receptor (RXR) to control multiple developmental processes in vertebrates. The available clinic and experimental data provide uncontested evidence for the pleiotropic roles of RA signalling in development of multiple embryonic structures and organs such eyes, central nervous system, gonads, lungs and heart. The development of any of these above-mentioned embryonic organ systems can be effectively utilized to showcase the many strategies utilized by RA signalling. However, it is very likely that the strategies employed to transfer RA signals during cardiac development comprise the majority of the relevant and sophisticated ways through which retinoid signals can be conveyed in a complex biological system. Here, we provide the reader with arguments indicating that RA signalling is exquisitely regulated according to specific phases of cardiac development and that RA signalling itself is one of the major regulators of the timing of cardiac morphogenesis and differentiation. We will focus on the role of signalling by RA receptors (RARs) in early phases of heart development. PMID:25134739

  12. The evolution of bat nucleic acid-sensing Toll-like receptors.

    PubMed

    Escalera-Zamudio, Marina; Zepeda-Mendoza, M Lisandra; Loza-Rubio, Elizabeth; Rojas-Anaya, Edith; Méndez-Ojeda, Maria L; Arias, Carlos F; Greenwood, Alex D

    2015-12-01

    We characterized the nucleic acid-sensing Toll-like receptors (TLR) of a New World bat species, the common vampire bat (Desmodus rotundus), and through a comparative molecular evolutionary approach searched for general adaptation patterns among the nucleic acid-sensing TLRs of eight different bats species belonging to three families (Pteropodidae, Vespertilionidae and Phyllostomidae). We found that the bat TLRs are evolving slowly and mostly under purifying selection and that the divergence pattern of such receptors is overall congruent with the species tree, consistent with the evolution of many other mammalian nuclear genes. However, the chiropteran TLRs exhibited unique mutations fixed in ligand-binding sites, some of which involved nonconservative amino acid changes and/or targets of positive selection. Such changes could potentially modify protein function and ligand-binding properties, as some changes were predicted to alter nucleic acid binding motifs in TLR 9. Moreover, evidence for episodic diversifying selection acting specifically upon the bat lineage and sublineages was detected. Thus, the long-term adaptation of chiropterans to a wide variety of environments and ecological niches with different pathogen profiles is likely to have shaped the evolution of the bat TLRs in an order-specific manner. The observed evolutionary patterns provide evidence for potential functional differences between bat and other mammalian TLRs in terms of resistance to specific pathogens or recognition of nucleic acids in general.

  13. Citric acid cycle intermediates as ligands for orphan G-protein-coupled receptors.

    PubMed

    He, Weihai; Miao, Frederick J-P; Lin, Daniel C-H; Schwandner, Ralf T; Wang, Zhulun; Gao, Jinhai; Chen, Jin-Long; Tian, Hui; Ling, Lei

    2004-05-13

    The citric acid cycle is central to the regulation of energy homeostasis and cell metabolism. Mutations in enzymes that catalyse steps in the citric acid cycle result in human diseases with various clinical presentations. The intermediates of the citric acid cycle are present at micromolar concentration in blood and are regulated by respiration, metabolism and renal reabsorption/extrusion. Here we show that GPR91 (ref. 3), a previously orphan G-protein-coupled receptor (GPCR), functions as a receptor for the citric acid cycle intermediate succinate. We also report that GPR99 (ref. 4), a close relative of GPR91, responds to alpha-ketoglutarate, another intermediate in the citric acid cycle. Thus by acting as ligands for GPCRs, succinate and alpha-ketoglutarate are found to have unexpected signalling functions beyond their traditional roles. Furthermore, we show that succinate increases blood pressure in animals. The succinate-induced hypertensive effect involves the renin-angiotensin system and is abolished in GPR91-deficient mice. Our results indicate a possible role for GPR91 in renovascular hypertension, a disease closely linked to atherosclerosis, diabetes and renal failure.

  14. Transcriptional upregulation of retinoic acid receptor beta (RAR beta) expression by phenylacetate in human neuroblastoma cells.

    PubMed

    Sidell, N; Chang, B; Yamashiro, J M; Wada, R K

    1998-02-25

    Sodium phenylacetate (NaPA) has been shown to synergize with retinoic acid (RA) in inducing the differentiation of human neuroblastoma cells. Our studies indicated that NaPA can impact on the RA differentiation program by upregulating nuclear retinoic acid receptor-beta (RAR beta) expression. We have found that NaPA does not alter the half-life of RAR beta mRNA; thus, increased stability of mRNA levels does not contribute to NaPA induction. In contrast, NaPA was able to specifically activate a reporter gene construct (delta SV beta RE-CAT) which contains a retinoic acid response element (RARE beta) that is located in the RAR beta promoter. Activation of delta SV beta RE-CAT by NaPA also occurred in neuroblastoma cells cotransfected with a nuclear retinoic acid receptor expression vector, demonstrating the independence of this activation on cellular RAR levels. Taken together, our findings suggest that induction of RAR beta by NaPA is regulated at the level of transcription and mediated through the retinoic acid response element, RARE beta. This effect may account, at least in part, for the strong synergy between NaPA and RA in promoting neuroblastoma differentiation.

  15. Arylpiperazines with N-acylated amino acids as 5-HT1A receptor ligands.

    PubMed

    Zajdel, Paweł; Subra, Gilles; Bojarski, Andrzej J; Duszyńska, Beata; Pawłowski, Maciej; Martinez, Jean

    2006-07-01

    A library consisting of 60 arylpiperazines modified with N-acylated amino acids was prepared on BAL linker SynPhasetrade mark Lanterns and evaluated in vitro for 5-HT(1A) receptor affinity. Biological screening, followed by a simple Fujita-Ban analysis, enabled the description of structure-activity relationships and allowed the selection of some potent, high-affinity ligands for in vivo pharmacological investigations.

  16. Bile acids regulate intestinal cell proliferation by modulating EGFR and FXR signaling

    PubMed Central

    Dossa, Avafia Y.; Escobar, Oswaldo; Golden, Jamie; Frey, Mark R.; Ford, Henri R.

    2015-01-01

    Bile acids (BAs) are synthesized in the liver and secreted into the intestine. In the lumen, enteric bacteria metabolize BAs from conjugated, primary forms into more toxic unconjugated, secondary metabolites. Secondary BAs can be injurious to the intestine and may contribute to disease. The epidermal growth factor receptor (EGFR) and the nuclear farnesoid X receptor (FXR) are known to interact with BAs. In this study we examined the effects of BAs on intestinal epithelial cell proliferation and investigated the possible roles for EGFR and FXR in these effects. We report that taurine-conjugated cholic acid (TCA) induced proliferation, while its unconjugated secondary counterpart deoxycholic acid (DCA) inhibited proliferation. TCA stimulated phosphorylation of Src, EGFR, and ERK 1/2. Pharmacological blockade of any of these pathways or genetic ablation of EGFR abrogated TCA-stimulated proliferation. Interestingly, Src or EGFR inhibitors eliminated TCA-induced phosphorylation of both molecules, suggesting that their activation is interdependent. In contrast to TCA, DCA exposure diminished EGFR phosphorylation, and pharmacological or siRNA blockade of FXR abolished DCA-induced inhibition of proliferation. Taken together, these results suggest that TCA induces intestinal cell proliferation via Src, EGFR, and ERK activation. In contrast, DCA inhibits proliferation via an FXR-dependent mechanism that may include downstream inactivation of the EGFR/Src/ERK pathway. Since elevated secondary BA levels are the result of specific bacterial modification, this may provide a mechanism through which an altered microbiota contributes to normal or abnormal intestinal epithelial cell proliferation. PMID:26608185

  17. Synthetic FXR agonist GW4064 is a modulator of multiple G protein-coupled receptors.

    PubMed

    Singh, Nidhi; Yadav, Manisha; Singh, Abhishek Kumar; Kumar, Harish; Dwivedi, Shailendra Kumar Dhar; Mishra, Jay Sharan; Gurjar, Anagha; Manhas, Amit; Chandra, Sharat; Yadav, Prem Narayan; Jagavelu, Kumaravelu; Siddiqi, Mohammad Imran; Trivedi, Arun Kumar; Chattopadhyay, Naibedya; Sanyal, Sabyasachi

    2014-05-01

    The synthetic nuclear bile acid receptor (farnesoid X receptor [FXR]) agonist GW4064 is extensively used as a specific pharmacological tool to illustrate FXR functions. We noticed that GW4064 activated empty luciferase reporters in FXR-deficient HEK-293T cells. We postulated that this activity of GW4064 might be routed through as yet unknown cellular targets and undertook an unbiased exploratory approach to identify these targets. Investigations revealed that GW4064 activated cAMP and nuclear factor for activated T-cell response elements (CRE and NFAT-RE, respectively) present on these empty reporters. Whereas GW4064-induced NFAT-RE activation involved rapid intracellular Ca(2+) accumulation and NFAT nuclear translocation, CRE activation involved soluble adenylyl cyclase-dependent cAMP accumulation and Ca(2+)-calcineurin-dependent nuclear translocation of transducers of regulated CRE-binding protein 2. Use of dominant negative heterotrimeric G-protein minigenes revealed that GW4064 caused activation of Gαi/o and Gq/11 G proteins. Sequential pharmacological inhibitor-based screening and radioligand-binding studies revealed that GW4064 interacted with multiple G protein-coupled receptors. Functional studies demonstrated that GW4064 robustly activated H1 and H4 and inhibited H2 histamine receptor signaling events. We also found that MCF-7 breast cancer cells, reported to undergo GW4064-induced apoptosis in an FXR-dependent manner, did not express FXR, and the GW4064-mediated apoptosis, also apparent in HEK-293T cells, could be blocked by selective histamine receptor regulators. Taken together, our results demonstrate identification of histamine receptors as alternate targets for GW4064, which not only necessitates cautious interpretation of the biological functions attributed to FXR using GW4064 as a pharmacological tool but also provides a basis for the rational designing of new pharmacophores for histamine receptor modulation.

  18. The nuclear receptor PPARγ individually responds to serotonin- and fatty acid-metabolites

    PubMed Central

    Waku, Tsuyoshi; Shiraki, Takuma; Oyama, Takuji; Maebara, Kanako; Nakamori, Rinna; Morikawa, Kosuke

    2010-01-01

    The nuclear receptor, peroxisome proliferator-activated receptor γ (PPARγ), recognizes various synthetic and endogenous ligands by the ligand-binding domain. Fatty-acid metabolites reportedly activate PPARγ through conformational changes of the Ω loop. Here, we report that serotonin metabolites act as endogenous agonists for PPARγ to regulate macrophage function and adipogenesis by directly binding to helix H12. A cyclooxygenase inhibitor, indomethacin, is a mimetic agonist of these metabolites. Crystallographic analyses revealed that an indole acetate functions as a common moiety for the recognition by the sub-pocket near helix H12. Intriguingly, a serotonin metabolite and a fatty-acid metabolite each bind to distinct sub-pockets, and the PPARγ antagonist, T0070907, blocked the fatty-acid agonism, but not that of the serotonin metabolites. Mutational analyses on receptor-mediated transcription and coactivator binding revealed that each metabolite individually uses coregulator and/or heterodimer interfaces in a ligand-type-specific manner. Furthermore, the inhibition of the serotonin metabolism reduced the expression of the endogenous PPARγ-target gene. Collectively, these results suggest a novel agonism, in which PPARγ functions as a multiple sensor in response to distinct metabolites. PMID:20717101

  19. The Molecular Basis of Ligand Interaction at Free Fatty Acid Receptor 4 (FFA4/GPR120)*

    PubMed Central

    Hudson, Brian D.; Shimpukade, Bharat; Milligan, Graeme; Ulven, Trond

    2014-01-01

    The long-chain fatty acid receptor FFA4 (previously GPR120) is receiving substantial interest as a novel target for the treatment of metabolic and inflammatory disease. This study examines for the first time the detailed mode of binding of both long-chain fatty acid and synthetic agonist ligands at FFA4 by integrating molecular modeling, receptor mutagenesis, and ligand structure-activity relationship approaches in an iterative format. In doing so, residues required for binding of fatty acid and synthetic agonists to FFA4 have been identified. This has allowed for the refinement of a well validated model of the mode of ligand-FFA4 interaction that will be invaluable in the identification of novel ligands and the future development of this receptor as a therapeutic target. The model reliably predicted the effects of substituent variations on agonist potency, and it was also able to predict the qualitative effect of binding site mutations in the majority of cases. PMID:24860101

  20. Obeticholic acid for the treatment of primary biliary cholangitis in adult patients: clinical utility and patient selection

    PubMed Central

    Bowlus, Christopher L

    2016-01-01

    Primary biliary cholangitis (PBC), previously known as primary biliary “cirrhosis”, is a rare autoimmune liver disease characterized by the hallmark autoantibodies to mitochondrial antigens and immune-mediated destruction of small bile duct epithelial cells leading to cholestasis and cirrhosis. Surprisingly, while immune modulators have not been effective in the treatment of PBC, supplementation with the hydrophilic bile acid (BA) ursodeoxycholic acid (UDCA) has been demonstrated to slow the disease progression. However, a significant minority of PBC patients do not have a complete response to UDCA and remain at risk of continued disease progression. Although the mechanisms of action are not well understood, UDCA provided proof of concept for BA therapy in PBC. Obeticholic acid (OCA), a novel derivative of the human BA chenodeoxycholic acid, is a potent agonist of the nuclear hormone receptor farnesoid X receptor, which regulates BA synthesis and transport. A series of clinical trials of OCA in PBC, primarily in combination with UDCA, have established that OCA leads to significant reductions in serum alkaline phosphatase that are predicted to lead to improved clinical outcomes, while dose-dependent pruritus has been the most common adverse effect. On the basis of these studies, OCA was given conditional approval by the US Food and Drug Administration with plans to establish the long-term clinical efficacy of OCA in patients with advanced PBC. PMID:27621676

  1. Essential role for retinoic acid in the promotion of CD4+ T cell effector responses via retinoic acid receptor alpha

    PubMed Central

    Hall, J.A.; Cannons, J.L.; Grainger, J.R.; Santos, L.M. Dos; Hand, T.W.; Naik, S.; Wohlfert, E.A.; Chou, D.B.; Oldenhove, G.; Robinson, M.; Grigg, M.E.; Kastenmayer, R.; Schwartzberg, P.L.; Belkaid, Y.

    2012-01-01

    SUMMARY Vitamin A and its metabolite, retinoic acid (RA), have recently been implicated in the regulation of immune homeostasis via the peripheral induction of regulatory T cells. Here we show that RA is also required to elicit proinflammatory CD4+ helper T cell responses to infection and mucosal vaccination. Retinoic acid receptor alpha (RARα) is the critical mediator of these effects. Strikingly, antagonism of RAR signaling and deficiency in RARα(Rara−/−) results in a cell autonomous CD4+ T cell activation defect. Altogether, these findings reveal a fundamental role for the RA/RARα axis in the development of both regulatory and inflammatory arms of adaptive immunity and establish nutritional status as a broad regulator of adaptive T cell responses. PMID:21419664

  2. A gate-latch-lock mechanism for hormone signalling by abscisic acid receptors

    SciTech Connect

    Melcher, Karsten; Ng, Ley-Moy; Zhou, X Edward; Soon, Fen-Fen; Xu, Yong; Suino-Powell, Kelly M; Park, Sang-Youl; Weiner, Joshua J; Fujii, Hiroaki; Chinnusamy, Viswanathan; Kovach, Amanda; Li, Jun; Wang, Yonghong; Li, Jiayang; Peterson, Francis C; Jensen, Davin R; Yong, Eu-Leong; Volkman, Brian F; Cutler, Sean R; Zhu, Jian-Kang; Xu, H Eric

    2010-01-12

    Abscisic acid (ABA) is a ubiquitous hormone that regulates plant growth, development and responses to environmental stresses. Its action is mediated by the PYR/PYL/RCAR family of START proteins, but it remains unclear how these receptors bind ABA and, in turn, how hormone binding leads to inhibition of the downstream type 2C protein phosphatase (PP2C) effectors. Here we report crystal structures of apo and ABA-bound receptors as well as a ternary PYL2-ABA-PP2C complex. The apo receptors contain an open ligand-binding pocket flanked by a gate that closes in response to ABA by way of conformational changes in two highly conserved β-loops that serve as a gate and latch. Moreover, ABA-induced closure of the gate creates a surface that enables the receptor to dock into and competitively inhibit the PP2C active site. A conserved tryptophan in the PP2C inserts directly between the gate and latch, which functions to further lock the receptor in a closed conformation. Together, our results identify a conserved gate-latch-lock mechanism underlying ABA signalling.

  3. Characterization of gamma-aminobutyric acid receptors in the neurointermediate lobe of the amphibian Xenopus laevis.

    PubMed

    Verburg-van Kemenade, B M; Jenks, B G; Lenssen, F J; Vaudry, H

    1987-02-01

    The neurotransmitter gamma-aminobutyric acid (GABA) is involved in the regulation of secretion of MSH from the intermediate lobe of Xenopus laevis. The purpose of this study was to identify the GABA receptor(s) involved by determination of the effect of specific receptor agonists and antagonists on the release of immunoreactive MSH from superfused neurointermediate lobes of Xenopus. Exogenous GABA induces a rapid inhibition of MSH secretion. There was no evidence for a transitory stimulatory effect of GABA as reported for the rat melanotropes. Both the GABA agonists (GABAa) homotaurine and isoguvacine and the GABA agonist (GABAb) baclofen inhibited MSH release in a dose-dependent manner. In vivo, homotaurine and baclofen caused aggregation of pigment in dermal melanophores. The MSH release-inhibiting effect of homotaurine and isoguvacine could be antagonized by the specific GABAa receptor antagonist bicuculline. However, bicuculline and picrotoxin failed to block the effect of exogenous GABA. We conclude that in the neurointermediate lobe of Xenopus laevis both GABAa and GABAb receptors are present, suggesting a dual inhibitory regulation.

  4. N-Methyl-d-aspartate Receptor Antagonists Have Variable Affect in 3-Nitropropionic Acid Toxicity

    PubMed Central

    Carbery, Timothy; Geddes, James W.

    2013-01-01

    There is accumulating evidence that excitotoxicity and oxidative stress resulting from excessive activation of glutamate (N-methyl-d-aspartate) NMDA receptors are major participants in striatal degeneration associated with 3-nitropropionic acid (3NP) administration. Although excitotoxic and oxidative mechanisms are implicated in 3NP toxicity, there are conflicting reports as to whether NMDA receptor antagonists attenuate or exacerbate the 3NP-induced neurodegeneration. In the present study, we investigated the involvement of NMDA receptors in striatal degeneration, protein oxidation and motor impairment following systemic 3NP administration. We examined whether NMDA receptor antagonists, memantine and ifenprodil, influence the neurotoxicity of 3NP. The development of striatal lesion and protein oxidation following 3NP administration is delayed by memantine but not affected by ifenprodil. However, in behavioral experiments, memantine failed to improve and ifenprodil exacerbated the motor deficits associated with 3NP toxicity. Together, these findings suggest caution in the application of NMDA receptor antagonists as a neuroprotective agent in neurodegenerative disorders associated with metabolic impairment. PMID:18688711

  5. N-methyl-D-aspartate receptor antagonists have variable affect in 3-nitropropionic acid toxicity.

    PubMed

    Nasr, Payman; Carbery, Timothy; Geddes, James W

    2009-03-01

    There is accumulating evidence that excitotoxicity and oxidative stress resulting from excessive activation of glutamate (N-methyl-D-aspartate) NMDA receptors are major participants in striatal degeneration associated with 3-nitropropionic acid (3NP) administration. Although excitotoxic and oxidative mechanisms are implicated in 3NP toxicity, there are conflicting reports as to whether NMDA receptor antagonists attenuate or exacerbate the 3NP-induced neurodegeneration. In the present study, we investigated the involvement of NMDA receptors in striatal degeneration, protein oxidation and motor impairment following systemic 3NP administration. We examined whether NMDA receptor antagonists, memantine and ifenprodil, influence the neurotoxicity of 3NP. The development of striatal lesion and protein oxidation following 3NP administration is delayed by memantine but not affected by ifenprodil. However, in behavioral experiments, memantine failed to improve and ifenprodil exacerbated the motor deficits associated with 3NP toxicity. Together, these findings suggest caution in the application of NMDA receptor antagonists as a neuroprotective agent in neurodegenerative disorders associated with metabolic impairment.

  6. Agrochemical control of plant water use using engineered abscisic acid receptors.

    PubMed

    Park, Sang-Youl; Peterson, Francis C; Mosquna, Assaf; Yao, Jin; Volkman, Brian F; Cutler, Sean R

    2015-04-23

    Rising temperatures and lessening fresh water supplies are threatening agricultural productivity and have motivated efforts to improve plant water use and drought tolerance. During water deficit, plants produce elevated levels of abscisic acid (ABA), which improves water consumption and stress tolerance by controlling guard cell aperture and other protective responses. One attractive strategy for controlling water use is to develop compounds that activate ABA receptors, but agonists approved for use have yet to be developed. In principle, an engineered ABA receptor that can be activated by an existing agrochemical could achieve this goal. Here we describe a variant of the ABA receptor PYRABACTIN RESISTANCE 1 (PYR1) that possesses nanomolar sensitivity to the agrochemical mandipropamid and demonstrate its efficacy for controlling ABA responses and drought tolerance in transgenic plants. Furthermore, crystallographic studies provide a mechanistic basis for its activity and demonstrate the relative ease with which the PYR1 ligand-binding pocket can be altered to accommodate new ligands. Thus, we have successfully repurposed an agrochemical for a new application using receptor engineering. We anticipate that this strategy will be applied to other plant receptors and represents a new avenue for crop improvement.

  7. Dopamine D2High receptors stimulated by phencyclidines, lysergic acid diethylamide, salvinorin A, and modafinil.

    PubMed

    Seeman, Philip; Guan, Hong-Chang; Hirbec, Hélène

    2009-08-01

    Although it is commonly stated that phencyclidine is an antagonist at ionotropic glutamate receptors, there has been little measure of its potency on other receptors in brain tissue. Although we previously reported that phencyclidine stimulated cloned-dopamine D2Long and D2Short receptors, others reported that phencyclidine did not stimulate D2 receptors in homogenates of rat brain striatum. This study, therefore, examined whether phencyclidine and other hallucinogens and psychostimulants could stimulate the incorporation of [(35)S]GTP-gamma-S into D2 receptors in homogenates of rat brain striatum, using the same conditions as previously used to study the cloned D2 receptors. Using 10 microM dopamine to define 100% stimulation, phencyclidine elicited a maximum incorporation of 46% in rat striata, with a half-maximum concentration of 70 nM for phencyclidine, when compared with 80 nM for dopamine, 89 nM for salvinorin A (48 nM for D2Long), 105 nM for lysergic acid diethylamide (LSD), 120 nM for R-modafinil, 710 nM for dizocilpine, 1030 nM for ketamine, and >10,000 nM for S-modafinil. These compounds also inhibited the binding of the D2-selective ligand [(3)H]domperidone. The incorporation was inhibited by the presence of 200 microM guanylylimidodiphosphate and also by D2 blockade, using 10 microM S-sulpiride, but not by D1 blockade with 10 microM SCH23390. Hypertonic buffer containing 150 mM NaCl inhibited the stimulation by phencyclidine, which may explain negative results by others. It is concluded that phencyclidine and other psychostimulants and hallucinogens can stimulate dopamine D2 receptors at concentrations related to their behavioral actions.

  8. Ultraviolet irradiation selectively disrupts the gamma-aminobutyric acid/benzodiazepine receptor-linked chloride ionophore

    SciTech Connect

    Evoniuk, G.; Moody, E.J.; Skolnick, P. )

    1989-05-01

    The ability of UV light to affect radioligand binding and 36Cl-uptake at the gamma-aminobutyric acidA (GABAA) receptor-chloride channel complex was examined. Exposure to 302 nm UV light produced a rapid (t1/2 = 4 min) reduction in (35S)t-butylbicyclo-phosphorothionate binding (assayed in the presence of 200 mM chloride) to sites associated with the GABAA receptor-coupled chloride ionophore. Saturation analysis revealed that this effect could be attributed entirely to a decrease in the maximum number of binding sites. Exposure to UV irradiation at lower (254 nm) and higher (366 nm) wavelengths also inhibited (35S)t-butylbicy-clophosphorothionate binding, but the respective rates of inactivation were 8- and 27-fold slower, compared with 302 nm. Other anion-dependent interactions at the GABAA receptor complex were disrupted in a similar manner. In the absence of permeant anion, (3H)flunitrazepam binding to benzodiazepine receptors was unaffected by 302 nm UV irradiation, whereas chloride-enhanced (3H)flunitrazepam binding was inhibited markedly. In the presence of 250-500 mM chloride, (3H)methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate binding to benzodiazepine receptors was also inhibited after UV exposure. Basal 36Cl- uptake into synaptoneurosomes was nearly doubled after 15 min of exposure to 302 nm light, whereas pentobarbital- and muscimol-stimulated 36Cl- uptake were reduced significantly. UV irradiation at 302 nm appears to disrupt selectively the anion-dependent functional interactions at the GABAA receptor complex. The apparent wavelength specificity suggests that the gating structure (channel) may contain tryptophan and/or tyrosine residues vital to the regulation of anion movement through the ionophore portion of this supramolecular receptor-ion channel complex.

  9. Inhibition of lysophosphatidic acid receptors 1 and 3 attenuates atherosclerosis development in LDL-receptor deficient mice

    PubMed Central

    Kritikou, Eva; van Puijvelde, Gijs H. M.; van der Heijden, Thomas; van Santbrink, Peter J.; Swart, Maarten; Schaftenaar, Frank H.; Kröner, Mara J.; Kuiper, Johan; Bot, Ilze

    2016-01-01

    Lysophosphatidic acid (LPA) is a natural lysophospholipid present at high concentrations within lipid-rich atherosclerotic plaques. Upon local accumulation in the damaged vessels, LPA can act as a potent activator for various types of immune cells through its specific membrane receptors LPA1/3. LPA elicits chemotactic, pro-inflammatory and apoptotic effects that lead to atherosclerotic plaque progression. In this study we aimed to inhibit LPA signaling by means of LPA1/3 antagonism using the small molecule Ki16425. We show that LPA1/3 inhibition significantly impaired atherosclerosis progression. Treatment with Ki16425 also resulted in reduced CCL2 production and secretion, which led to less monocyte and neutrophil infiltration. Furthermore, we provide evidence that LPA1/3 blockade enhanced the percentage of non-inflammatory, Ly6Clow monocytes and CD4+ CD25+ FoxP3+ T-regulatory cells. Finally, we demonstrate that LPA1/3 antagonism mildly reduced plasma LDL cholesterol levels. Therefore, pharmacological inhibition of LPA1/3 receptors may prove a promising approach to diminish atherosclerosis development. PMID:27883026

  10. Inhibition of lysophosphatidic acid receptors 1 and 3 attenuates atherosclerosis development in LDL-receptor deficient mice.

    PubMed

    Kritikou, Eva; van Puijvelde, Gijs H M; van der Heijden, Thomas; van Santbrink, Peter J; Swart, Maarten; Schaftenaar, Frank H; Kröner, Mara J; Kuiper, Johan; Bot, Ilze

    2016-11-24

    Lysophosphatidic acid (LPA) is a natural lysophospholipid present at high concentrations within lipid-rich atherosclerotic plaques. Upon local accumulation in the damaged vessels, LPA can act as a potent activator for various types of immune cells through its specific membrane receptors LPA1/3. LPA elicits chemotactic, pro-inflammatory and apoptotic effects that lead to atherosclerotic plaque progression. In this study we aimed to inhibit LPA signaling by means of LPA1/3 antagonism using the small molecule Ki16425. We show that LPA1/3 inhibition significantly impaired atherosclerosis progression. Treatment with Ki16425 also resulted in reduced CCL2 production and secretion, which led to less monocyte and neutrophil infiltration. Furthermore, we provide evidence that LPA1/3 blockade enhanced the percentage of non-inflammatory, Ly6C(low) monocytes and CD4(+) CD25(+) FoxP3(+) T-regulatory cells. Finally, we demonstrate that LPA1/3 antagonism mildly reduced plasma LDL cholesterol levels. Therefore, pharmacological inhibition of LPA1/3 receptors may prove a promising approach to diminish atherosclerosis development.

  11. Structure-based drug design targeting the cell membrane receptor GPBAR1: exploiting the bile acid scaffold towards selective agonism

    NASA Astrophysics Data System (ADS)

    di Leva, Francesco Saverio; Festa, Carmen; Renga, Barbara; Sepe, Valentina; Novellino, Ettore; Fiorucci, Stefano; Zampella, Angela; Limongelli, Vittorio

    2015-11-01

    Bile acids can regulate nutrient metabolism through the activation of the cell membrane receptor GPBAR1 and the nuclear receptor FXR. Developing an exogenous control over these receptors represents an attractive strategy for the treatment of enterohepatic and metabolic disorders. A number of dual GPBAR1/FXR agonists are known, however their therapeutic use is limited by multiple unwanted effects due to activation of the diverse downstream signals controlled by the two receptors. On the other hand, designing selective GPBAR1 and FXR agonists is challenging since the two proteins share similar structural requisites for ligand binding. Here, taking advantage of our knowledge of the two targets, we have identified through a rational drug design study a series of amine lithocholic acid derivatives as selective GPBAR1 agonists. The presence of the 3α-NH2 group on the steroidal scaffold is responsible for the selectivity over FXR unveiling unprecedented structural insights into bile acid receptors activity modulation.

  12. Retinoic Acid Receptor β2 Agonists Restore Glycemic Control In Diabetes and Reduce Steatosis

    PubMed Central

    Trasino, Steven E.; Tang, Xiao-Han; Jessurun, Jose; Gudas, Lorraine J.

    2016-01-01

    Aims Retinoids (vitamin A (retinol), and structurally related molecules) possess metabolic modulating properties, prompting new interest in their role in the treatment of diabetes and fatty liver disease, but little is known about the effects of specific retinoic acid receptor (RAR) agonists in these diseases. Materials and Methods Synthetic agonists for retinoic acid receptor RARβ2 were administered to wild type (wt) mice in a model of high fat diet (HFD)-induced type 2 diabetes (T2D) and to ob/ob and db/db mice (genetic models of obesity-associated T2D). Results We demonstrate that administration of synthetic agonists for the retinoic acid receptor RARβ2 to either wild type (wt) mice in a model of high fat diet (HFD)-induced type 2 diabetes (T2D) or to ob/ob and db/db mice (genetic models of obesity-associated T2D) reduces hyperglycemia, peripheral insulin resistance, and body weight. Furthermore, RARβ2 agonists dramatically reduce steatosis, lipid peroxidation, and oxidative stress in the liver, pancreas, and kidneys of obese, diabetic mice. RARβ2 agonists also lower levels of mRNAs involved in lipogenesis, such as SREBP1 and FASN (fatty acid synthase), and increase mRNAs that mediate mitochondrial fatty acid β-oxidation, such as CPT1α, in these organs. RARβ2 agonists lower triglyceride levels in these organs, and in muscle. Conclusions Collectively, our data show that orally active, rapidly acting, high affinity pharmacological agonists for RARβ2 improve the diabetic phenotype while reducing lipid levels in key insulin target tissues. We suggest that RARβ2 agonists should be useful drugs for T2D therapy and for treatment of hepatic steatosis. PMID:26462866

  13. Positive modulation of the α9α10 nicotinic cholinergic receptor by ascorbic acid

    PubMed Central

    Boffi, JC; Wedemeyer, C; Lipovsek, M; Katz, E; Calvo, DJ; Elgoyhen, AB

    2013-01-01

    Background and Purpose The activation of α9α10 nicotinic cholinergic receptors (nAChRs) present at the synapse between efferent olivocochlear fibres and cochlear hair cells can prevent acoustic trauma. Hence, pharmacological potentiators of these receptors could be useful therapeutically. In this work, we characterize ascorbic acid as a positive modulator of recombinant α9α10 nAChRs. Experimental Approach ACh-evoked responses were analysed under two-electrode voltage-clamp recordings in Xenopus laevis oocytes injected with α9 and α10 cRNAs. Key Results Ascorbic acid potentiated ACh responses in X. laevis oocytes expressing α9α10 (but not α4β2 or α7) nAChRs, in a concentration-dependent manner, with an effective concentration range of 1–30 mM. The compound did not affect the receptor's current–voltage profile nor its apparent affinity for ACh, but it significantly enhanced the maximal evoked currents (percentage of ACh maximal response, 240 ± 20%). This effect was specific for the L form of reduced ascorbic acid. Substitution of the extracellular cysteine residues present in loop C of the ACh binding site did not affect the potentiation. Ascorbic acid turned into a partial agonist of α9α10 nAChRs bearing a point mutation at the pore domain of the channel (TM2 V13′T mutant). A positive allosteric mechanism of action rather than an antioxidant effect of ascorbic acid is proposed. Conclusions and Implications The present work describes one of the few agents that activates or potentiates α9α10 nAChRs and leads to new avenues for designing drugs with potential therapeutic use in inner ear disorders. PMID:22994414

  14. Skin Barrier Recovery by Protease-Activated Receptor-2 Antagonist Lobaric Acid

    PubMed Central

    Joo, Yeon Ah; Chung, Hyunjin; Yoon, Sohyun; Park, Jong Il; Lee, Ji Eun; Myung, Cheol Hwan; Hwang, Jae Sung

    2016-01-01

    Atopic dermatitis (AD) results from gene and environment interactions that lead to a range of immunological abnormalities and breakdown of the skin barrier. Protease-activated receptor 2 (PAR2) belongs to a family of G-protein coupled receptors and is expressed in suprabasal layers of the epidermis. PAR2 is activated by both trypsin and a specific agonist peptide, SLIGKV-NH2 and is involved in both epidermal permeability barrier homeostasis and epithelial inflammation. In this study, we investigated the effect of lobaric acid on inflammation, keratinocyte differentiation, and recovery of the skin barrier in hairless mice. Lobaric acid blocked trypsin-induced and SLIGKV-NH2-induced PAR2 activation resulting in decreased mobilization of intracellular Ca2+ in HaCaT keratinocytes. Lobaric acid reduced expression of interleukin-8 induced by SLIGKV-NH2 and thymus and activation regulated chemokine (TARC) induced by tumor necrosis factor-a (TNF-α) and IFN-γ in HaCaT keratinocytes. Lobaric acid also blocked SLIGKV-NH2-induced activation of ERK, which is a downstream signal of PAR2 in normal human keratinocytes (NHEKs). Treatment with SLIGKV-NH2 downregulated expression of involucrin, a differentiation marker protein in HaCaT keratinocytes, and upregulated expression of involucrin, transglutamase1 and filaggrin in NHEKs. However, lobaric acid antagonized the effect of SLIGKV-NH2 in HaCaT keratinocytes and NHEKs. Topical application of lobaric acid accelerated barrier recovery kinetics in a SKH-1 hairless mouse model. These results suggested that lobaric acid is a PAR2 antagonist and could be a possible therapeutic agent for atopic dermatitis. PMID:27169822

  15. Folic acid mediates activation of the pro-oncogene STAT3 via the Folate Receptor alpha.

    PubMed

    Hansen, Mariann F; Greibe, Eva; Skovbjerg, Signe; Rohde, Sarah; Kristensen, Anders C M; Jensen, Trine R; Stentoft, Charlotte; Kjær, Karina H; Kronborg, Camilla S; Martensen, Pia M

    2015-07-01

    The signal transducer and activator of transcription 3 (STAT3) is a well-described pro-oncogene found constitutively activated in several cancer types. Folates are B vitamins that, when taken up by cells through the Reduced Folate Carrier (RFC), are essential for normal cell growth and replication. Many cancer cells overexpress a glycophosphatidylinositol (GPI)-anchored Folate Receptor α (FRα). The function of FRα in cancer cells is still poorly described, and it has been suggested that transport of folate is not its primary function in these cells. We show here that folic acid and folinic acid can activate STAT3 through FRα in a Janus Kinase (JAK)-dependent manner, and we demonstrate that gp130 functions as a transducing receptor for this signalling. Moreover, folic acid can promote dose dependent cell proliferation in FRα-positive HeLa cells, but not in FRα-negative HEK293 cells. After folic acid treatment of HeLa cells, up-regulation of the STAT3 responsive genes Cyclin A2 and Vascular Endothelial Growth Factor (VEGF) were verified by qRT-PCR. The identification of this FRα-STAT3 signal transduction pathway activated by folic and folinic acid contributes to the understanding of the involvement of folic acid in preventing neural tube defects as well as in tumour growth. Previously, the role of folates in these diseases has been attributed to their roles as one-carbon unit donors following endocytosis into the cell. Our finding that folic acid can activate STAT3 via FRα adds complexity to the established roles of B9 vitamins in cancer and neural tube defects.

  16. Active transport of amino acids by a guanidiniocarbonyl-pyrrole receptor.

    PubMed

    Urban, Christian; Schmuck, Carsten

    2010-08-16

    Herein we report the synthesis and characterization of a transporter 9 for N-acetylated amino acids. Transporter 9 is a conjugate of a guanidiniocarbonyl pyrrole cation, one of the most efficient carboxylate binding motifs reported so far, and a hydrophobic tris(dodecylbenzyl) group, which ensures solubility in organic solvents. In its protonated form, 9 binds N-acetylated amino acid carboxylates in wet organic solvents with association constants in the range of 10(4) M(-1) as estimated by extraction experiments. Aromatic amino acids are preferred due to additional cation-pi-interactions of the amino acid side chain with the guanidiniocarbonyl pyrrole moiety. U-tube experiments established efficient transport across a bulk liquid chloroform phase with fluxes approaching 10(-6) mol m(-2) s(-1). In experiments with single substrates, the release rate of the amino acid from the receptor-substrate complex at the interface with the receiving phase is rate determining. In contrast to this, in competition experiments with several substrates, the thermodynamic affinity to 9 becomes decisive. As 9 can only transport anions in its protonated form and has a pK(a) value of approximately 7, pH-driven active transport of amino acids is also possible. Transport occurs as a symport of the amino acid carboxylate and a proton.

  17. Fatty Acid Amide Hydrolase (FAAH) Inhibition Enhances Memory Acquisition through Activation of PPAR-alpha Nuclear Receptors

    ERIC Educational Resources Information Center

    Mazzola, Carmen; Medalie, Julie; Scherma, Maria; Panlilio, Leigh V.; Solinas, Marcello; Tanda, Gianluigi; Drago, Filippo; Cadet, Jean Lud; Goldberg, Steven R.; Yasar, Sevil

    2009-01-01

    Inhibitors of fatty acid amide hydrolase (FAAH) increase endogenous levels of anandamide (a cannabinoid CB[subscript 1]-receptor ligand) and oleoylethanolamide and palmitoylethanolamide (OEA and PEA, ligands for alpha-type peroxisome proliferator-activated nuclear receptors, PPAR-alpha) when and where they are naturally released in the brain.…

  18. Amino acids in the COOH-terminal region of the oxytocin receptor third intracellular domain are important for receptor function.

    PubMed

    Zhong, Miao; Parish, Bridgette; Murtazina, Dilyara A; Ku, Chun-Ying; Sanborn, Barbara M

    2007-04-01

    Previously, residue K6.30 in the COOH-terminal region of the third intracellular domain (3iC) of the oxytocin (OT) receptor (OTR) was identified as important for receptor function leading to phospholipase C activation in both OTR and the vasopressin V(2) receptor (V(2)R) chimera V(2)ROTR3iC. Substitution of either A6.28K or V6.30K in wild-type V(2)R did not recapitulate the increase in phosphatidylinositide (PI) turnover observed in V(2)ROTR3iC. Hence, the role of K6.30 may be context-specific. Deletion of two NH(2)-terminal OTR3iC segments in the V(2)ROTR3iC chimera did not diminish vasopressin-stimulated PI turnover, whereas deletion of RVSSVKL (residues 6.19-6.25) reduced receptor expression. Deletion of this sequence in wild-type OTR reduced expression by 50% without affecting affinity for [(3)H]OT. This OTR mutant was unable to activate PI turnover or extracellular signal-regulated kinase 1/2 phosphorylation. The effects of alanine substitution for individual residues in RVSSVKL indicated differential importance for OTR function. The R6.19A substitution lost high-affinity sites for [(3)H]OT and the ability to stimulate PI turnover. Affinity for [(3)H]OT and membrane expression was not affected by any other substitutions. OTR-V6.20A and OTR-K6.24A mutants functioned as well as wild-type OTR, whereas OTR S6.21A, S6.22A, and V6.23A mutants exhibited impaired abilities to activate PI turnover (20-40% of OTR), and the OTR-L6.25A mutant exhibited constitutive activity. In conclusion, specific amino acids in the RVSSVKL segment in the COOH-terminal region of the third intracellular domain of OTR influence the ability of OTR to activate G protein-mediated actions.

  19. Correlation of phospholipid structure with functional effects on the nicotinic acetylcholine receptor. A modulatory role for phosphatidic acid.

    PubMed Central

    Bhushan, A; McNamee, M G

    1993-01-01

    Fourier transform infrared spectroscopy is used to characterize specific interactions between negatively charged lipids, such as phosphatidic acid, and the purified nicotinic acetylcholine receptor from Torpedo californica. The specific interaction of phosphatidic acid with acetylcholine receptor is demonstrated by the receptor-induced perturbation of the lipid ionization state, which is monitored using Fourier transform infrared bands arising from the phosphate head group. The acetylcholine receptor shifts the pKa of phosphatidic acid molecules adjacent to the receptor to a lower value by almost 2 pH units from 8.5 to 6.6. Decreased pH also leads to changes in ion channel function and to changes in the secondary structure of the acetylcholine receptor in membranes containing ionizable phospholipids. Phospholipase D restores functional activity of acetylcholine receptor reconstituted in an unfavorable environment containing phosphatidylcholine by generating phosphatidic acid. Lipids such as phosphatidic acid may serve as allosteric effectors for membrane protein function and the lipid-protein interface could be a site for activity-dependent changes that lead to modulation of synaptic efficacy. PMID:8471723

  20. Three Dimensional Structure Prediction of Fatty Acid Binding Site on Human Transmembrane Receptor CD36.

    PubMed

    Tarhda, Zineb; Semlali, Oussama; Kettani, Anas; Moussa, Ahmed; Abumrad, Nada A; Ibrahimi, Azeddine

    2013-01-01

    CD36 is an integral membrane protein which is thought to have a hairpin-like structure with alpha-helices at the C and N terminals projecting through the membrane as well as a larger extracellular loop. This receptor interacts with a number of ligands including oxidized low density lipoprotein and long chain fatty acids (LCFAs). It is also implicated in lipid metabolism and heart diseases. It is therefore important to determine the 3D structure of the CD36 site involved in lipid binding. In this study, we predict the 3D structure of the fatty acid (FA) binding site [127-279 aa] of the CD36 receptor based on homology modeling with X-ray structure of Human Muscle Fatty Acid Binding Protein (PDB code: 1HMT). Qualitative and quantitative analysis of the resulting model suggests that this model was reliable and stable, taking in consideration over 97.8% of the residues in the most favored regions as well as the significant overall quality factor. Protein analysis, which relied on the secondary structure prediction of the target sequence and the comparison of 1HMT and CD36 [127-279 aa] secondary structures, led to the determination of the amino acid sequence consensus. These results also led to the identification of the functional sites on CD36 and revealed the presence of residues which may play a major role during ligand-protein interactions.

  1. Three Dimensional Structure Prediction of Fatty Acid Binding Site on Human Transmembrane Receptor CD36

    PubMed Central

    Tarhda, Zineb; Semlali, Oussama; Kettani, Anas; Moussa, Ahmed; Abumrad, Nada A.; Ibrahimi, Azeddine

    2013-01-01

    CD36 is an integral membrane protein which is thought to have a hairpin-like structure with alpha-helices at the C and N terminals projecting through the membrane as well as a larger extracellular loop. This receptor interacts with a number of ligands including oxidized low density lipoprotein and long chain fatty acids (LCFAs). It is also implicated in lipid metabolism and heart diseases. It is therefore important to determine the 3D structure of the CD36 site involved in lipid binding. In this study, we predict the 3D structure of the fatty acid (FA) binding site [127–279 aa] of the CD36 receptor based on homology modeling with X-ray structure of Human Muscle Fatty Acid Binding Protein (PDB code: 1HMT). Qualitative and quantitative analysis of the resulting model suggests that this model was reliable and stable, taking in consideration over 97.8% of the residues in the most favored regions as well as the significant overall quality factor. Protein analysis, which relied on the secondary structure prediction of the target sequence and the comparison of 1HMT and CD36 [127–279 aa] secondary structures, led to the determination of the amino acid sequence consensus. These results also led to the identification of the functional sites on CD36 and revealed the presence of residues which may play a major role during ligand-protein interactions. PMID:24348024

  2. Identification of amino acids involved in histamine potentiation of GABAA receptors

    PubMed Central

    Thiel, Ulrike; Platt, Sarah J.; Wolf, Steffen; Hatt, Hanns; Gisselmann, Günter

    2015-01-01

    Histamine is a neurotransmitter involved in a number of physiological and neuronal functions. In mammals, such as humans, and rodents, the histaminergic neurons found in the tuberomamillary nucleus project widely throughout the central nervous system. Histamine acts as positive modulator of GABAA receptors (GABAARs) and, in high concentrations (10 mM), as negative modulator of the strychnine-sensitive glycine receptor. However, the exact molecular mechanisms by which histamine acts on GABAARs are unknown. In our study, we aimed to identify amino acids potentially involved in the modulatory effect of histamine on GABAARs. We expressed GABAARs with 12 different point mutations in Xenopus laevis oocytes and characterized the effect of histamine on GABA-induced currents using the two-electrode voltage clamp technique. Our data demonstrate that the amino acid residues β2(N265) and β2(M286), which are important for modulation by propofol, are not involved in the action of histamine. However, we found that histamine modulation is dependent on the amino acid residues α1(R120), β2(Y157), β2(D163), β3(V175), and β3(Q185). We showed that the amino acid residues β2(Y157) and β3(Q185) mediate the positive modulatory effect of histamine on GABA-induced currents, whereas α1(R120) and β2(D163) form a potential histamine interaction site in GABAARs. PMID:26074818

  3. Cyclin D1 represses peroxisome proliferator-activated receptor alpha and inhibits fatty acid oxidation

    PubMed Central

    Hanse, Eric A.; Mashek, Douglas G.; Mashek, Mara T.; Hendrickson, Anna M.; Mullany, Lisa K.; Albrecht, Jeffrey H.

    2016-01-01

    Cyclin D1 is a cell cycle protein that promotes proliferation by mediating progression through key checkpoints in G1 phase. It is also a proto-oncogene that is commonly overexpressed in human cancers. In addition to its canonical role in controlling cell cycle progression, cyclin D1 affects other aspects of cell physiology, in part through transcriptional regulation. In this study, we find that cyclin D1 inhibits the activity of a key metabolic transcription factor, peroxisome proliferator-activated receptor α (PPARα), a member of nuclear receptor family that induces fatty acid oxidation and may play an anti-neoplastic role. In primary hepatocytes, cyclin D1 inhibits PPARα transcriptional activity and target gene expression in a cdk4-independent manner. In liver and breast cancer cells, knockdown of cyclin D1 leads to increased PPARα transcriptional activity, expression of PPARα target genes, and fatty acid oxidation. Similarly, cyclin D1 depletion enhances binding of PPARα to target sequences by chromatin immunoprecipitation. In proliferating hepatocytes and regenerating liver in vivo, induction of endogenous cyclin D1 is associated with diminished PPARα activity. Cyclin D1 expression is both necessary and sufficient for growth factor-mediated repression of fatty acid oxidation in proliferating hepatocytes. These studies indicate that in addition to playing a pivotal role in cell cycle progression, cyclin D1 represses PPARα activity and inhibits fatty acid oxidation. Our findings establish a new link between cyclin D1 and metabolism in both tumor cells and physiologic hepatocyte proliferation. PMID:27351284

  4. Cyclin D1 represses peroxisome proliferator-activated receptor alpha and inhibits fatty acid oxidation.

    PubMed

    Kamarajugadda, Sushama; Becker, Jennifer R; Hanse, Eric A; Mashek, Douglas G; Mashek, Mara T; Hendrickson, Anna M; Mullany, Lisa K; Albrecht, Jeffrey H

    2016-07-26

    Cyclin D1 is a cell cycle protein that promotes proliferation by mediating progression through key checkpoints in G1 phase. It is also a proto-oncogene that is commonly overexpressed in human cancers. In addition to its canonical role in controlling cell cycle progression, cyclin D1 affects other aspects of cell physiology, in part through transcriptional regulation. In this study, we find that cyclin D1 inhibits the activity of a key metabolic transcription factor, peroxisome proliferator-activated receptor α (PPARα), a member of nuclear receptor family that induces fatty acid oxidation and may play an anti-neoplastic role. In primary hepatocytes, cyclin D1 inhibits PPARα transcriptional activity and target gene expression in a cdk4-independent manner. In liver and breast cancer cells, knockdown of cyclin D1 leads to increased PPARα transcriptional activity, expression of PPARα target genes, and fatty acid oxidation. Similarly, cyclin D1 depletion enhances binding of PPARα to target sequences by chromatin immunoprecipitation. In proliferating hepatocytes and regenerating liver in vivo, induction of endogenous cyclin D1 is associated with diminished PPARα activity. Cyclin D1 expression is both necessary and sufficient for growth factor-mediated repression of fatty acid oxidation in proliferating hepatocytes. These studies indicate that in addition to playing a pivotal role in cell cycle progression, cyclin D1 represses PPARα activity and inhibits fatty acid oxidation. Our findings establish a new link between cyclin D1 and metabolism in both tumor cells and physiologic hepatocyte proliferation.

  5. Reversible receptor methylation is essential for normal chemotaxis of Escherichia coli in gradients of aspartic acid.

    PubMed Central

    Weis, R M; Koshland, D E

    1988-01-01

    The chemotaxis of wild-type cells of Escherichia coli and double mutants lacking the methyltransferase and the methylesterase activities of the receptor modification system has been compared in spatial gradients of aspartic acid. Previous studies showing that a chemotactic response can be observed for the mutant raised questions about the role of methylation in the bacterial memory. To clarify the role of methylation, the redistribution of bacteria in stabilized defined gradients of aspartic acid was monitored by light scattering. There was no redistribution of the mutant cells in nonsaturating gradients of aspartic acid, but over the same range these mutant bacteria were observed to respond and to adapt during tethering experiments. In large saturating gradients of aspartate, slight movement of the mutant up the gradient was observed. These results show that dynamic receptor methylation is required for the chemotactic response to gentle gradients of aspartic acid and that methylation resets to zero and is part of the normal wild-type memory. There are certain gradients, however, in which the methylation-deficient mutants show chemotactic ability, thus explaining the apparent anomaly. Images PMID:2829179

  6. Molecular basis for designing selective modulators of retinoic acid receptor transcriptional activities.

    PubMed

    Lefebvre, P

    2001-08-01

    Retinoic acid receptors are ligand-regulated transcription factors belonging to the nuclear receptor superfamily, which comprises 49 members in the human genome. all-trans retinoic acid and 9-cis retinoic acid receptors (RARs and RXRs) are each encoded by three distinct genes and several isoforms arise from alternative splicing and the use of different promoters. While RXRs are promiscuous dimerization partners of several other nuclear receptors, RARs are active, in-vivo, when associated to RXRs. Retinoids are therefore regulators of multiple physiological processes, from embryogenesis to metabolism. Different combinations of RXR:RAR heterodimers occur as a function of their tissue-specific expression and their activity is mostly conditioned by the activation status of RAR. These heterodimers are defined as non permissive heterodimers, in opposition to permissive dimers whose transcriptional activity may be modulated through RXR and its dimerization partner. The transcriptional activity of these dimers also relies on their ability to recruit nuclear coactivators and corepressors, which function as multi proteic complexes harboring several enzymatic activities (acetylases, kinases). The structure of the ligand bound to the RAR moiety of the dimer, as well as the nature of the DNA sequence to which dimers are bound, dictate the relative affinity of dimers for coactivators and thus its overall transcriptional activity. RARs are also able to repress the activity of unrelated transcription factors such as AP1 and NF-kappa-B, and therefore have potent anti proliferative and anti inflammatory properties. This review summarizes our current view of molecular mechanisms governing these various activities and emphasizes the need for a detailed understanding of how retinoids may dictate transactivating and transrepressive properties of RARs and RXRs, which may be considered as highly valuable therapeutic targets in many diseases such as cancer, skin hyperproliferation and

  7. PLZF is a negative regulator of retinoic acid receptor transcriptional activity

    PubMed Central

    Martin, Perrine J; Delmotte, Marie-Hélène; Formstecher, Pierre; Lefebvre, Philippe

    2003-01-01

    Background Retinoic acid receptors (RARs) are ligand-regulated transcription factors controlling cellular proliferation and differentiation. Receptor-interacting proteins such as corepressors and coactivators play a crucial role in specifying the overall transcriptional activity of the receptor in response to ligand treatment. Little is known however on how receptor activity is controlled by intermediary factors which interact with RARs in a ligand-independent manner. Results We have identified the promyelocytic leukemia zinc finger protein (PLZF), a transcriptional corepressor, to be a RAR-interacting protein using the yeast two-hybrid assay. We confirmed this interaction by GST-pull down assays and show that the PLZF N-terminal zinc finger domain is necessary and sufficient for PLZF to bind RAR. The RAR ligand binding domain displayed the highest affinity for PLZF, but corepressor and coactivator binding interfaces did not contribute to PLZF recruitment. The interaction was ligand-independent and correlated to a decreased transcriptional activity of the RXR-RAR heterodimer upon overexpression of PLZF. A similar transcriptional interference could be observed with the estrogen receptor alpha and the glucocorticoid receptor. We further show that PLZF is likely to act by preventing RXR-RAR heterodimerization, both in-vitro and in intact cells. Conclusion Thus RAR and PLZF interact physically and functionally. Intriguingly, these two transcription factors play a determining role in hematopoiesis and regionalization of the hindbrain and may, upon chromosomal translocation, form fusion proteins. Our observations therefore define a novel mechanism by which RARs activity may be controlled. PMID:14521715

  8. Ginseng pharmacology: a new paradigm based on gintonin-lysophosphatidic acid receptor interactions

    PubMed Central

    Choi, Sun-Hye; Jung, Seok-Won; Lee, Byung-Hwan; Kim, Hyeon-Joong; Hwang, Sung-Hee; Kim, Ho-Kyoung; Nah, Seung-Yeol

    2015-01-01

    Ginseng, the root of Panax ginseng, is used as a traditional medicine. Despite the long history of the use of ginseng, there is no specific scientific or clinical rationale for ginseng pharmacology besides its application as a general tonic. The ambiguous description of ginseng pharmacology might be due to the absence of a predominant active ingredient that represents ginseng pharmacology. Recent studies show that ginseng abundantly contains lysophosphatidic acids (LPAs), which are phospholipid-derived growth factor with diverse biological functions including those claimed to be exhibited by ginseng. LPAs in ginseng form a complex with ginseng proteins, which can bind and deliver LPA to its cognate receptors with a high affinity. As a first messenger, gintonin produces second messenger Ca2+ via G protein-coupled LPA receptors. Ca2+ is an intracellular mediator of gintonin and initiates a cascade of amplifications for further intercellular communications by activation of Ca2+-dependent kinases, receptors, gliotransmitter, and neurotransmitter release. Ginsenosides, which have been regarded as primary ingredients of ginseng, cannot elicit intracellular [Ca2+]i transients, since they lack specific cell surface receptor. However, ginsenosides exhibit non-specific ion channel and receptor regulations. This is the key characteristic that distinguishes gintonin from ginsenosides. Although the current discourse on ginseng pharmacology is focused on ginsenosides, gintonin can definitely provide a mode of action for ginseng pharmacology that ginsenosides cannot. This review article introduces a novel concept of ginseng ligand-LPA receptor interaction and proposes to establish a paradigm that shifts the focus from ginsenosides to gintonin as a major ingredient representing ginseng pharmacology. PMID:26578955

  9. Ligand Induction of Retinoic Acid Receptors Alters an Acute Infection by Murine Cytomegalovirus†

    PubMed Central

    Angulo, Ana; Chandraratna, Roshantha A. S.; LeBlanc, James F.; Ghazal, Peter

    1998-01-01

    Here we report that administration of retinoids can alter the outcome of an acute murine cytomegalovirus (MCMV) infection. We show that a crucial viral control element, the major immediate-early enhancer, can be activated by retinoic acid (RA) via multiple RA-responsive elements (DR2) that bind retinoid X receptor-retinoic acid receptor (RAR) heterodimers with apparent dissociation constants ranging from 15 to 33 nM. Viral growth is dramatically increased upon RA treatment of infected tissue culture cells. Using synthetic retinoid receptor-specific agonists and antagonists, we provide evidence that RAR activation in cells is required for mediating the response of MCMV to RA. Oral administration of RA to infected immunocompetent mice selectively exacerbates an infection by MCMV, while cotreatment with an RAR antagonist protects against the adverse effects of RA on MCMV infection. In conclusion, these chemical genetic experiments provide evidence that an RAR-mediated pathway can modulate in vitro and in vivo infections by MCMV. PMID:9573222

  10. Ursodeoxycholic Acid Induces Death Receptor-mediated Apoptosis in Prostate Cancer Cells

    PubMed Central

    Lee, Won Sup; Jung, Ji Hyun; Panchanathan, Radha; Yun, Jeong Won; Kim, Dong Hoon; Kim, Hye Jung; Kim, Gon Sup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2017-01-01

    Background Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. Methods Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. Results UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. Conclusions UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer. PMID:28382282

  11. Functional analysis of free fatty acid receptor GPR120 in human eosinophils: implications in metabolic homeostasis.

    PubMed

    Konno, Yasunori; Ueki, Shigeharu; Takeda, Masahide; Kobayashi, Yoshiki; Tamaki, Mami; Moritoki, Yuki; Oyamada, Hajime; Itoga, Masamichi; Kayaba, Hiroyuki; Omokawa, Ayumi; Hirokawa, Makoto

    2015-01-01

    Recent evidence has shown that eosinophils play an important role in metabolic homeostasis through Th2 cytokine production. GPR120 (FFA4) is a G protein-coupled receptor (GPCR) for long-chain fatty acids that functions as a regulator of physiological energy metabolism. In the present study, we aimed to investigate whether human eosinophils express GPR120 and, if present, whether it possesses a functional capacity on eosinophils. Eosinophils isolated from peripheral venous blood expressed GPR120 at both the mRNA and protein levels. Stimulation with a synthetic GPR120 agonist, GW9508, induced rapid down-regulation of cell surface expression of GPR120, suggesting ligand-dependent receptor internalization. Although GPR120 activation did not induce eosinophil chemotactic response and degranulation, we found that GW9508 inhibited eosinophil spontaneous apoptosis and Fas receptor expression. The anti-apoptotic effect was attenuated by phosphoinositide 3-kinase (PI3K) inhibitors and was associated with inhibition of caspase-3 activity. Eosinophil response investigated using ELISpot assay indicated that stimulation with a GPR120 agonist induced IL-4 secretion. These findings demonstrate the novel functional properties of fatty acid sensor GPR120 on human eosinophils and indicate the previously unrecognized link between nutrient metabolism and the immune system.

  12. Expression and distribution of sialic acid influenza virus receptors in wild birds.

    PubMed

    França, M; Stallknecht, D E; Howerth, E W

    2013-02-01

    Avian influenza (AI) viruses have been detected in more than 105 wild bird species from 12 different orders but species-related differences in susceptibility to AI viruses exist. Expression of α2,3-linked (avian-type) and α2,6-linked (human-type) sialic acid (SA) influenza virus receptors in tissues is considered one of the determinants of the host range and tissue tropism of influenza viruses. We investigated the expression of these SA receptors in 37 wild bird species from 11 different orders by lectin histochemistry. Two isoforms of Maackia amurensis (MAA) lectin, MAA1 and MAA2, were used to detect α2,3-linked SA, and Sambucus nigra lectin was used to detect α2,6-linked SA. All species evaluated expressed α2,3-linked and α2,6-linked SA receptors in endothelial cells and renal tubular epithelial cells. Both α2,3-linked and α-2,6-linked SA receptors were expressed in respiratory and intestinal tract tissues of aquatic and terrestrial wild bird species from different taxa, but differences in SA expression and in the predominant isoform of MAA lectin bound were observed. With a few possible exceptions, these observed differences were not generally predictive of reported species susceptibility to AI viruses based on published experimental and field data.

  13. Waking action of ursodeoxycholic acid (UDCA) involves histamine and GABAA receptor block.

    PubMed

    Yanovsky, Yevgenij; Schubring, Stephan R; Yao, Quiaoling; Zhao, Yan; Li, Sha; May, Andrea; Haas, Helmut L; Lin, Jian-Sheng; Sergeeva, Olga A

    2012-01-01

    Since ancient times ursodeoxycholic acid (UDCA), a constituent of bile, is used against gallstone formation and cholestasis. A neuroprotective action of UDCA was demonstrated recently in models of Alzheimer's disease and retinal degeneration. The mechanisms of UDCA action in the nervous system are poorly understood. We show now that UDCA promotes wakefulness during the active period of the day, lacking this activity in histamine-deficient mice. In cultured hypothalamic neurons UDCA did not affect firing rate but synchronized the firing, an effect abolished by the GABA(A)R antagonist gabazine. In histaminergic neurons recorded in slices UDCA reduced amplitude and duration of spontaneous and evoked IPSCs. In acutely isolated histaminergic neurons UDCA inhibited GABA-evoked currents and sIPSCs starting at 10 µM (IC(50) = 70 µM) and did not affect NMDA- and AMPA-receptor mediated currents at 100 µM. Recombinant GABA(A) receptors composed of α1, β1-3 and γ2L subunits expressed in HEK293 cells displayed a sensitivity to UDCA similar to that of native GABA(A) receptors. The mutation α1V256S, known to reduce the inhibitory action of pregnenolone sulphate, reduced the potency of UDCA. The mutation α1Q241L, which abolishes GABA(A)R potentiation by several neurosteroids, had no effect on GABA(A)R inhibition by UDCA. In conclusion, UDCA enhances alertness through disinhibition, at least partially of the histaminergic system via GABA(A) receptors.

  14. Programmable Multivalent Display of Receptor Ligands using Peptide Nucleic Acid Nanoscaffolds

    PubMed Central

    Englund, Ethan A.; Wang, Deyun; Fujigaki, Hidetsugu; Sakai, Hiroyasu; Micklitsch, Christopher M.; Ghirlando, Rodolfo; Martin-Manso, Gema; Pendrak, Michael L.; Roberts, David D.; Durell, Stewart R.; Appella, Daniel H.

    2012-01-01

    Multivalent effects dictate the binding affinity of multiple ligands on one molecular entity to receptors. Integrins are receptors that mediate cell attachment through multivalent binding to peptide sequences within the extracellular matrix, and overexpression promotes the metastasis of some cancers. Multivalent display of integrin antagonists enhances their efficacy, but current scaffolds have limited ranges and precision for the display of ligands. Here we present an approach to study multivalent effects across wide ranges of ligand number, density, and three-dimensional arrangement. Using L-lysine γ-substituted peptide nucleic acids, the multivalent effects of an integrin antagonist were examined over a range of 1 to 45 ligands. The optimal construct improves the inhibitory activity of the antagonist by two orders of magnitude against the binding of melanoma cells to the extracellular matrix in both in vitro and in vivo models. PMID:22233624

  15. G-protein coupling and nuclear translocation of the human abscisic acid receptor LANCL2

    PubMed Central

    Fresia, Chiara; Vigliarolo, Tiziana; Guida, Lucrezia; Booz, Valeria; Bruzzone, Santina; Sturla, Laura; Di Bona, Melody; Pesce, Mattia; Usai, Cesare; De Flora, Antonio; Zocchi, Elena

    2016-01-01

    Abscisic acid (ABA), a long known phytohormone, has been recently demonstrated to be present also in humans, where it targets cells of the innate immune response, mesenchymal and hemopoietic stem cells and cells involved in the regulation of systemic glucose homeostasis. LANCL2, a peripheral membrane protein, is the mammalian ABA receptor. We show that N-terminal glycine myristoylation causes LANCL2 localization to the plasmamembrane and to cytoplasmic membrane vesicles, where it interacts with the α subunit of a Gi protein and starts the ABA signaling pathway via activation of adenylate cyclase. Demyristoylation of LANCL2 by chemical or genetic means triggers its nuclear translocation. Nuclear enrichment of native LANCL2 is also induced by ABA treatment. Therefore human LANCL2 is a non-transmembrane G protein-coupled receptor susceptible to hormone-induced nuclear translocation. PMID:27222287

  16. Effects of retinoic acid on growth hormone-releasing hormone receptor, growth hormone secretagogue receptor gene expression and growth hormone secretion in rat anterior pituitary cells.

    PubMed

    Maliza, Rita; Fujiwara, Ken; Tsukada, Takehiro; Azuma, Morio; Kikuchi, Motoshi; Yashiro, Takashi

    2016-06-30

    Retinoic acid (RA) is an important signaling molecule in embryonic development and adult tissue. The actions of RA are mediated by the nuclear receptors retinoic acid receptor (RAR) and retinoid X receptor (RXR), which regulate gene expression. RAR and RXR are widely expressed in the anterior pituitary gland. RA was reported to stimulate growth hormone (GH) gene expression in the anterior pituitary cells. However, current evidence is unclear on the role of RA in gene expression of growth hormone-releasing hormone receptor (Ghrh-r), growth hormone secretagogue receptor (Ghs-r) and somatostatin receptors (Sst-rs). Using isolated anterior pituitary cells of rats, we examined the effects of RA on gene expression of these receptors and GH release. Quantitative real-time PCR revealed that treatment with all-trans retinoic acid (ATRA; 10(-6) M) for 24 h increased gene expression levels of Ghrh-r and Ghs-r; however, expressions of Sst-r2 and Sst-r5 were unchanged. Combination treatment with the RAR-agonist Am80 and RXR-agonist PA024 mimicked the effects of ATRA on Ghrh-r and Ghs-r gene expressions. Exposure of isolated pituitary cells to ATRA had no effect on basal GH release. In contrast, ATRA increased growth hormone-releasing hormone (GHRH)- and ghrelin-stimulated GH release from cultured anterior pituitary cells. Our results suggest that expressions of Ghrh-r and Ghs-r are regulated by RA through the RAR-RXR receptor complex and that RA enhances the effects of GHRH and ghrelin on GH release from the anterior pituitary gland.

  17. Nipecotic acid ethyl ester: a cholinergic agonist that may differentiate muscarinic receptor subtypes

    SciTech Connect

    Zorn, S.H.; Duman, R.S.; Enna, S.J.; Krogsgaard-Larsen, P.; Micheletti, R.; Giraldo, E.; Giachetti, A.

    1986-03-05

    Reports indicate that nipecotic acid ethyl ester (NAEE) displays cholinomimetic properties in vivo. In the present study a series of physiological and biochemical tests were conducted to characterize this action. NAEE had a negative inotropic effect on the guinea pig atrium, and stimulated contraction of the guinea pig ileum and isolated mouse stomach strip at concentrations similar to bethanechol (BCH). The atrial and ilial effects were reversed by atropine. Unlike BCH, NAEE had no effect on basal acid secretion in the isolated mouse stomach at concentrations < 100 ..mu..M. NAEE was more potent than carbachol (CCH) in displacing /sup 3/H-ONB binding from rat brain membranes. The potency of NAEE to inhibit antagonist binding in rat heart membranes was enhanced by Mg/sup + +/ (Hill coefficient < 1.0) and reduced by Gpp(NH)p. Like CCH, NAEE inhibited GTP-stimulated adenylate cyclase in rat brain striatal membranes. As compared to CCH, NAEE had little effect (< 5%) as a stimulator of inositol phosphate (IP) production in rat brain slices. The results indicate that NAEE is a direct-acting muscarinic receptor agonist. Moreover, its differential effects on acid secretion, IP accumulation, and adenylate cyclase suggest that it may be useful for defining cholinergic receptor subclasses.

  18. Activation of the aryl hydrocarbon receptor induces hepatic steatosis via the upregulation of fatty acid transport.

    PubMed

    Kawano, Yuki; Nishiumi, Shin; Tanaka, Shinwa; Nobutani, Kentaro; Miki, Akira; Yano, Yoshihiko; Seo, Yasushi; Kutsumi, Hiromu; Ashida, Hitoshi; Azuma, Takeshi; Yoshida, Masaru

    2010-12-15

    The aryl hydrocarbon receptor (AHR) is a basic helix-loop-helix/Per-ARNT-Sim domain transcription factor, which is activated by various xenobiotic ligands. AHR is known to be abundant in liver tissue and to be associated with hepatic steatosis. However, it has not yet been elucidated how the activation of AHR promotes hepatic steatosis. The aim of this study is to clarify the role of AHR in hepatic steatosis. The intraperitoneal injection of 3-methylcholanthrene (3MC), a potent AHR ligand, into C57BL/6J mice significantly increased the levels of triglycerides and six long-chain monounsaturated fatty acids in the livers of mice, resulting in hepatic microvesicular steatosis. 3MC significantly enhanced the expression level of fatty acid translocase (FAT), a factor regulating the uptake of long-chain fatty acids into hepatocytes, in the liver. In an in vitro experiment using human hepatoma HepG2 cells, 3MC increased the expression level of FAT, and the downregulation of AHR by AHR siRNA led to the suppression of 3MC-induced FAT expression. In addition, the mRNA level of peroxisome proliferator-activated receptor (PPAR) α, an upstream factor of FAT, was increased in the livers of 3MC-treated mice. Taking together, AHR activation induces hepatic microvesicular steatosis by increasing the expression level of FAT.

  19. Identification of Darmstoff analogs as selective agonists and antagonists of lysophosphatidic acid receptors.

    PubMed

    Gududuru, Veeresa; Zeng, Kui; Tsukahara, Ryoko; Makarova, Natalia; Fujiwara, Yuko; Pigg, Kathryn R; Baker, Daniel L; Tigyi, Gabor; Miller, Duane D

    2006-01-15

    Darmstoff describes a family of gut smooth muscle-stimulating acetal phosphatidic acids initially isolated and characterized from the bath fluid of stimulated gut over 50 years ago. Despite similar structural and biological profiles, Darmstoff analogs have not previously been examined as potential LPA mimetics. Here, we report a facile method for the synthesis of potassium salts of Darmstoff analogs. To understand the effect of stereochemistry on lysophosphatidic acid mimetic activity, synthesis of optically pure stereoisomers of selected Darmstoff analogs was achieved starting with chiral methyl glycerates. Each Darmstoff analog was evaluated for subtype-specific LPA receptor agonist/antagonist activity, PPARgamma activation, and autotaxin inhibition. From this study we identified compound 12 as a pan-antagonist and several pan-agonists for the LPA(1-3) receptors. Introduction of an aromatic ring in the lipid chain such as analog 22 produced a subtype-specific LPA(3) agonist with an EC(50) of 692 nM. Interestingly, regardless of their LPA(1/2/3) ligand properties all of the Darmstoff analogs tested activated PPARgamma. However, these compounds are weak inhibitors of autotaxin. The results indicate that Darmstoff analogs constitute a novel class of lysophosphatidic acid mimetics.

  20. Loss of Free Fatty Acid Receptor 2 leads to impaired islet mass and beta cell survival

    PubMed Central

    Villa, Stephanie R.; Priyadarshini, Medha; Fuller, Miles H.; Bhardwaj, Tanya; Brodsky, Michael R.; Angueira, Anthony R.; Mosser, Rockann E.; Carboneau, Bethany A.; Tersey, Sarah A.; Mancebo, Helena; Gilchrist, Annette; Mirmira, Raghavendra G.; Gannon, Maureen; Layden, Brian T.

    2016-01-01

    The regulation of pancreatic β cell mass is a critical factor to help maintain normoglycemia during insulin resistance. Nutrient-sensing G protein-coupled receptors (GPCR) contribute to aspects of β cell function, including regulation of β cell mass. Nutrients such as free fatty acids (FFAs) contribute to precise regulation of β cell mass by signaling through cognate GPCRs, and considerable evidence suggests that circulating FFAs promote β cell expansion by direct and indirect mechanisms. Free Fatty Acid Receptor 2 (FFA2) is a β cell-expressed GPCR that is activated by short chain fatty acids, particularly acetate. Recent studies of FFA2 suggest that it may act as a regulator of β cell function. Here, we set out to explore what role FFA2 may play in regulation of β cell mass. Interestingly, Ffar2−/− mice exhibit diminished β cell mass at birth and throughout adulthood, and increased β cell death at adolescent time points, suggesting a role for FFA2 in establishment and maintenance of β cell mass. Additionally, activation of FFA2 with Gαq/11-biased agonists substantially increased β cell proliferation in in vitro and ex vivo proliferation assays. Collectively, these data suggest that FFA2 may be a novel therapeutic target to stimulate β cell growth and proliferation. PMID:27324831

  1. Membrane bile acid receptor TGR5 predicts good prognosis in ampullary adenocarcinoma patients with hyperbilirubinemia

    PubMed Central

    Chen, Min-Chan; Chen, Yi-Ling; Wang, Tzu-Wen; Hsu, Hui-Ping; Lai, Ming-Derg

    2016-01-01

    Bile acids are potential carcinogens in gastrointestinal cancer, and interact with nuclear and membrane receptors to initiate downstream signaling. The effect of TGR5 [also known as G protein-coupled bile acid receptor 1 (GPBAR1)] on cancer progression is dependent on the tissue where it is activated. In this report, the function of TGR5 expression in cancer was studied using a bioinformatic approach. TGR5 expression in ampullary adenocarcinoma and normal duodenum was compared by western blotting, reverse transcription polymerase chain reaction, and immunohistochemistry (IHC). High GPBAR1 gene expression was found to be an indicator of worse prognosis in gastric and breast cancer patients, and an indication of better prognosis in ovarian cancer patients. The level of GPBAR1 gene expression was higher in bile-acid exposed cancer than in other types of cancer, and was increased in well-differentiated ampullary adenocarcinoma. Negative, weak or mild expression of TGR5 was correlated with younger age, higher plasma level of total/direct bilirubin, higher plasma concentration of CA-125, advanced tumor stage and advanced AJCC TNM stage. The disease-specific survival rate was highest in ampullary adenocarcinoma patients with high TGR5 expression and high total bilirubin level. In summary, TGR5 functions as a tumor-suppressor in patients with ampullary adenocarcinoma and preoperative hyperbilirubinemia. Further study of the suppressive mechanism may provide a new therapeutic option for patients with ampullary adenocarcinoma. PMID:27510297

  2. Characterization of influenza virus sialic acid receptors in minor poultry species.

    PubMed

    Kimble, Brian; Nieto, Gloria Ramirez; Perez, Daniel R

    2010-12-09

    It is commonly accepted that avian influenza viruses (AIVs) bind to terminal α2,3 sialic acid (SA) residues whereas human influenza viruses bind to α2,6 SA residues. By a series of amino acid changes on the HA surface protein, AIVs can switch receptor specificity and recognize α2,6 SA positive cells, including human respiratory epithelial cells. Animal species, like pigs and Japanese quail, that contain both α2,3 and α2,6 SA become ideal environments for receptor switching. Here, we describe the SA patterns and distributions in 6 common minor domestic poultry species: Peking duck, Toulouse geese, Chinese ring-neck pheasant, white midget turkey, bobwhite quail, and pearl guinea fowl. Lectins specific to α2,3 and α2,6 SA (Maakia amurensis agglutinin and Sambuca nigra agglutinin, respectively) were used to detect SA by an alkaline phosphotase-based method and a fluorescent-based method. Differences in SA moieties and their ability to bind influenza viruses were visualized by fluorescent labeling of 4 different H3N2 influenza viruses known to be specific for one receptor or the other. The geese and ducks showed α2,3 SA throughout the respiratory tract and marginal α2,6 SA only in the colon. The four other avian species showed both α2,3 and α2,6 SA in the respiratory tract and the intestines. Furthermore, the turkey respiratory tract showed a positive correlation between age and α2,6 SA levels. The fact that these birds have both avian and human flu receptors, combined with their common presence in backyard farms and live bird markets worldwide, mark them as potential mixing bowl species and necessitates improved surveillance and additional research about the role of these birds in influenza host switching.

  3. Free fatty acids-sensing G protein-coupled receptors in drug targeting and therapeutics.

    PubMed

    Yonezawa, Tomo; Kurata, Riho; Yoshida, Kaori; Murayama, Masanori A; Cui, Xiaofeng; Hasegawa, Akihiko

    2013-01-01

    G protein-coupled receptor (GPCR) (also known as seven-transmembrane domain receptor) superfamily represents the largest protein family in the human genome. These receptors respond to various physiological ligands such as photons, odors, pheromones, hormones, ions, and small molecules including amines, amino acids to large peptides and steroids. Thus, GPCRs are involved in many diseases and the target of around half of all conventional drugs. The physiological roles of free fatty acids (FFAs), in particular, long-chain FFAs, are important for the development of many metabolic disease including obesity, diabetes, and atherosclerosis. In the past half decade, deorphanization of several GPCRs has revealed that GPR40, GPR41, GPR43, GPR84 and GPR120 sense concentration of extracellular FFAs with various carbon chain lengths. GPR40 and GPR120 are activated by medium- and long-chain FFAs. GPR84 is activated by medium- chain, but not long-chain, FFAs. GPR41 and GPR43 are activated by short-chain FFAs. GPR40 is highly expressed in pancreatic beta cells and plays a crucial role in FFAs-induced insulin secretion. GPR120 is mainly expressed in enteroendocrine cells and plays an important role for FFAs-induced glucagon-like peptide-1. GPR43 is abundant in leukocytes and adipose tissue, whilst GPR41 is highly expressed in adipose tissue, the pancreas and leukocytes. GPR84 is expressed in leukocytes and monocyte/macrophage. This review aims to shed light on the physiological roles and development of drugs targeting these receptors.

  4. Oleanolic acid acrylate elicits antidepressant-like effect mediated by 5-HT1A receptor

    PubMed Central

    Fajemiroye, James O.; Polepally, Prabhakar R.; Chaurasiya, Narayan D.; Tekwani, Babu L.; Zjawiony, Jordan K.; Costa, Elson A.

    2015-01-01

    The development of new drugs for the treatment of depression is strategic to achieving clinical needs of patients. This study evaluates antidepressant-like effect and neural mechanisms of four oleanolic acid derivatives i.e. acrylate (D1), methacrylate (D2), methyl fumarate (D3) and ethyl fumarate (D4). All derivatives were obtained by simple one-step esterification of oleanolic acid prior to pharmacological screening in the forced swimming (FS) and open field (OF) tests. Pharmacological tools like α-methyl-p-tyrosine (AMPT, catecholamine depletor), p-chlorophenylalanine (serotonin depletor), prazosin (PRAZ, selective α1-receptor antagonist), WAY-100635 (selective serotonin 5-HT1A receptor antagonist) as well as monoamine oxidase (MAO) and functional binding assays were conducted to investigate possible neural mechanisms. In the FS test, D1 showed the most promising antidepressant-like effect without eliciting locomotor incoordination. Unlike group of mice pretreated with AMPT 100 mg/kg, PCPA 100 mg/kg or PRAZ 1 mg/kg, the effect of D1 was attenuated by WAY-100635 0.3 mg/kg pretreatment. D1 demonstrated moderate inhibition of MAO-A (IC50 = 48.848 ± 1.935 μM), potency (pEC50 = 6.1 ± 0.1) and intrinsic activity (Emax = 26 ± 2.0%) on 5-HT1A receptor. In conclusion, our findings showed antidepressant-like effect of D1 and possible involvement of 5-HT1A receptor. PMID:26199018

  5. Leptin receptor polymorphisms interact with polyunsaturated fatty acids to augment risk of insulin resistance and metabolic syndrome in adults

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The leptin receptor (LEPR) is associated with insulin resistance, a key feature of metabolic syndrome (MetS). Gene-fatty acid interactions may affect MetS risk. The objective was to investigate the relationship among LEPR polymorphisms, insulin resistance, andMetSrisk and whether plasma fatty acids,...

  6. The short-chain fatty acid receptor, FFA2, contributes to gestational glucose homeostasis.

    PubMed

    Fuller, Miles; Priyadarshini, Medha; Gibbons, Sean M; Angueira, Anthony R; Brodsky, Michael; Hayes, M Geoffrey; Kovatcheva-Datchary, Petia; Bäckhed, Fredrik; Gilbert, Jack A; Lowe, William L; Layden, Brian T

    2015-11-15

    The structure of the human gastrointestinal microbiota can change during pregnancy, which may influence gestational metabolism; however, a mechanism of action remains unclear. Here we observed that in wild-type (WT) mice the relative abundance of Actinobacteria and Bacteroidetes increased during pregnancy. Along with these changes, short-chain fatty acids (SCFAs), which are mainly produced through gut microbiota fermentation, significantly changed in both the cecum and peripheral blood throughout gestation in these mice. SCFAs are recognized by G protein-coupled receptors (GPCRs) such as free fatty acid receptor-2 (FFA2), and we have previously demonstrated that the fatty acid receptor-2 gene (Ffar2) expression is higher in pancreatic islets during pregnancy. Using female Ffar2-/- mice, we explored the physiological relevance of signaling through this GPCR and found that Ffar2-deficient female mice developed fasting hyperglycemia and impaired glucose tolerance in the setting of impaired insulin secretion compared with WT mice during, but not before, pregnancy. Insulin tolerance tests were similar in Ffar2-/- and WT mice before and during pregnancy. Next, we examined the role of FFA2 in gestational β-cell mass, observing that Ffar2-/- mice had diminished gestational expansion of β-cells during pregnancy. Interestingly, mouse genotype had no significant impact on the composition of the gut microbiome, but did affect the observed SCFA profiles, suggesting a functional difference in the microbiota. Together, these results suggest a potential link between increased Ffar2 expression in islets and the alteration of circulating SCFA levels, possibly explaining how changes in the gut microbiome contribute to gestational glucose homeostasis.

  7. Human gestation-associated tissues express functional cytosolic nucleic acid sensing pattern recognition receptors.

    PubMed

    Bryant, Aled H; Menzies, Georgina E; Scott, Louis M; Spencer-Harty, Samantha; Davies, Lleucu B; Smith, Rachel A; Jones, Ruth H; Thornton, Catherine A

    2017-03-13

    The role of viral infections in adverse pregnancy outcomes has gained interest in recent years. Innate immune pattern recognition receptors (PRRs) and their signalling pathways that yield a cytokine output in response to pathogenic stimuli have been postulated to link infection at the maternal-fetal interface and adverse pregnancy outcomes. The objective of this study was to investigate the expression and functional response of nucleic acid ligand responsive Toll-like receptors (TLR3, 7, 8 and 9), and retinoic acid-inducible gene 1 (RIG-I)-like receptors (RIG-I, MDA5 and LGP2) in human term gestation-associated tissues (placenta, choriodecidua and amnion) using an explant model. Immunohistochemistry revealed that these PRRs were expressed by the term placenta, choriodecidua and amnion. A statistically significant increase in interleukin (IL)-6 and/or IL-8 production in response to specific agonists for TLR3 (Poly(I:C); low and high molecular weight), TLR7 (Imiquimod), TLR8 (ssRNA40) and RIG-I/MDA5 (Poly(I:C)LyoVec) was observed; there was no response to a TLR9 (ODN21798) agonist. A hierarchical clustering approach was used to compare the response of each tissue type to the ligands studied and revealed that the placenta and choriodecidua generate a more similar IL-8 response, while the choriodecidua and amnion generate a more similar IL-6 response to nucleic acid ligands. These findings demonstrate that responsiveness via TLR3, TLR7, TLR8 and RIG-1/MDA5 is a broad feature of human term gestation-associated tissues with differential responses by tissue that might underpin adverse obstetric outcomes. This article is protected by copyright. All rights reserved.

  8. Up-regulation of the alpha-secretase ADAM10 by retinoic acid receptors and acitretin.

    PubMed

    Tippmann, Frank; Hundt, Jana; Schneider, Anja; Endres, Kristina; Fahrenholz, Falk

    2009-06-01

    Late-onset Alzheimer's disease is often connected with nutritional misbalance, such as enhanced cholesterol intake, deficiency in polyunsaturated fatty acids, or hypovitaminosis. The alpha-secretase ADAM10 has been found to be regulated by retinoic acid, the bioreactive metabolite of vitamin A. Here we show that retinoids induce gene expression of ADAM10 and alpha-secretase activity by nonpermissive retinoid acid receptor/retinoid X receptor (RAR/RXR) heterodimers, whereby alpha- and beta-isotypes of RAR play a major role. However, ligands of other RXR binding partners, such as the vitamin D receptor, do not stimulate alpha-secretase activity. On the basis of these findings, we examined the effect of synthetic retinoids and found a strong enhancement of nonamyloidogenic processing of the amyloid precursor protein by the vitamin A analog acitretin: it stimulated ADAM10 promoter activity with an EC(50) of 1.5 microM and led to an increase of mature ADAM10 protein that resulted in a two- to three-fold increase of the ratio between alpha- and beta-secretase activity in neuroblastoma cells. The alpha-secretase stimulation by acitretin was completely inhibited by the ADAM10-specific inhibitor GI254023X. Intracerebral injection of acitretin in APP/PS1-21 transgenic mice led to a reduction of Abeta(40) and Abeta(42). The results of this study may have clinical relevance because acitretin has been approved for the treatment of psoriasis since 1997 and found generally safe for long-term use in humans.

  9. Felbamate displays in vitro antiepileptic effects as a broad spectrum excitatory amino acid receptor antagonist.

    PubMed

    Domenici, M R; Sagratella, S; Ongini, E; Longo, R; Scotti de Carolis, A

    1994-12-27

    The in vitro antiepileptic activity of the novel anticonvulsant drug felbamate was tested in rat hippocampal slices on the CA1 epileptiform bursting induced by different chemical epileptogenic agents. The effects of felbamate were compared with those of the anticonvulsant drugs diphenylhydantoin and pentobarbitone and with the effects of excitatory amino acid antagonists acting at both N-methyl-D-aspartate (NMDA) and non-NMDA receptors. Like the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), felbamate at a minimum effective concentration of 1 mM induced a significant (P < 0.01) reduction of the duration of the CA1 epileptiform bursting due to the K+ channel blocker, 4-aminopyridine, and the excitatory amino acids, kainate and quisqualate. Like the NMDA receptor antagonist ketamine, felbamate (1.6 mM) significantly (P < 0.01) decreased the duration of the CA1 epileptiform bursting caused by 'Mg(2+)-free' solutions. Conversely, felbamate (1.6 mM), CNQX (100 microM) and ketamine (100 microM) failed to affect the epileptiform bursting induced by the GABA antagonist penicillin. Pentobarbitone (100 microM) significantly (P < 0.01) decreased the duration of the CA1 epileptiform bursting caused by 'Mg(2+)-free' solutions, 4-aminopyridine or penicillin, while diphenylhydantoin (up to concentrations of 100 microM) failed to have an effect. The results indicate that felbamate displays a unique profile of in vitro antiepileptic effects as a broad spectrum antagonist of excitatory amino acid transmission.

  10. Recessive and Dominant Mutations in Retinoic Acid Receptor Beta in Cases with Microphthalmia and Diaphragmatic Hernia

    PubMed Central

    Srour, Myriam; Chitayat, David; Caron, Véronique; Chassaing, Nicolas; Bitoun, Pierre; Patry, Lysanne; Cordier, Marie-Pierre; Capo-Chichi, José-Mario; Francannet, Christine; Calvas, Patrick; Ragge, Nicola; Dobrzeniecka, Sylvia; Hamdan, Fadi F.; Rouleau, Guy A.; Tremblay, André; Michaud, Jacques L.

    2013-01-01

    Anophthalmia and/or microphthalmia, pulmonary hypoplasia, diaphragmatic hernia, and cardiac defects are the main features of PDAC syndrome. Recessive mutations in STRA6, encoding a membrane receptor for the retinol-binding protein, have been identified in some cases with PDAC syndrome, although many cases have remained unexplained. Using whole-exome sequencing, we found that two PDAC-syndrome-affected siblings, but not their unaffected sibling, were compound heterozygous for nonsense (c.355C>T [p.Arg119∗]) and frameshift (c.1201_1202insCT [p.Ile403Serfs∗15]) mutations in retinoic acid receptor beta (RARB). Transfection studies showed that p.Arg119∗ and p.Ile403Serfs∗15 altered RARB had no transcriptional activity in response to ligands, confirming that the mutations induced a loss of function. We then sequenced RARB in 15 subjects with anophthalmia and/or microphthalmia and at least one other feature of PDAC syndrome. Surprisingly, three unrelated subjects with microphthalmia and diaphragmatic hernia showed de novo missense mutations affecting the same codon; two of the subjects had the c.1159C>T (Arg387Cys) mutation, whereas the other one carried the c.1159C>A (p.Arg387Ser) mutation. We found that compared to the wild-type receptor, p.Arg387Ser and p.Arg387Cys altered RARB induced a 2- to 3-fold increase in transcriptional activity in response to retinoic acid ligands, suggesting a gain-of-function mechanism. Our study thus suggests that both recessive and dominant mutations in RARB cause anophthalmia and/or microphthalmia and diaphragmatic hernia, providing further evidence of the crucial role of the retinoic acid pathway during eye development and organogenesis. PMID:24075189

  11. The short chain fatty acid receptor, FFA2, contributes to gestational glucose homeostasis

    DOE PAGES

    Fuller, Miles; Priyadarshini, Medha; Gibbons, Sean M.; ...

    2015-09-22

    The structure of the human gastrointestinal microbiota can change during pregnancy, which may influence gestational metabolism; however, a mechanism of action remains unclear. Here we observed that in wild-type (WT) mice the relative abundance of Actinobacteria and Bacteroidetes increased during pregnancy. Along with these changes, short-chain fatty acids (SCFAs), which are mainly produced through gut microbiota fermentation, significantly changed in both the cecum and peripheral blood throughout gestation in these mice. SCFAs are recognized by G protein-coupled receptors (GPCRs) such as free fatty acid receptor-2 (FFA2), and we have previously demonstrated that the fatty acid receptor-2 gene (Ffar2) expression ismore » higher in pancreatic islets during pregnancy. Using female Ffar2-/- mice, we explored the physiological relevance of signaling through this GPCR and found that Ffar2-deficient female mice developed fasting hyperglycemia and impaired glucose tolerance in the setting of impaired insulin secretion compared with WT mice during, but not before, pregnancy. Insulin tolerance tests were similar in Ffar2-/- and WT mice before and during pregnancy. Next, we examined the role of FFA2 in gestational β-cell mass, observing that Ffar2-/- mice had diminished gestational expansion of β-cells during pregnancy. Interestingly, mouse genotype had no significant impact on the composition of the gut microbiome, but did affect the observed SCFA profiles, suggesting a functional difference in the microbiota. Altogether, these results suggest a potential link between increased Ffar2 expression in islets and the alteration of circulating SCFA levels, possibly explaining how changes in the gut microbiome contribute to gestational glucose homeostasis.« less

  12. The short chain fatty acid receptor, FFA2, contributes to gestational glucose homeostasis

    SciTech Connect

    Fuller, Miles; Priyadarshini, Medha; Gibbons, Sean M.; Angueira, Anthony R.; Brodsky, Michael; Hayes, M. Geoffrey; Kovatcheva-Datchary, Petia; Backhed, Fredrik; Gilbert, Jack A.; Lowe, Jr., William L.; Layden, Brian T.

    2015-09-22

    The structure of the human gastrointestinal microbiota can change during pregnancy, which may influence gestational metabolism; however, a mechanism of action remains unclear. Here we observed that in wild-type (WT) mice the relative abundance of Actinobacteria and Bacteroidetes increased during pregnancy. Along with these changes, short-chain fatty acids (SCFAs), which are mainly produced through gut microbiota fermentation, significantly changed in both the cecum and peripheral blood throughout gestation in these mice. SCFAs are recognized by G protein-coupled receptors (GPCRs) such as free fatty acid receptor-2 (FFA2), and we have previously demonstrated that the fatty acid receptor-2 gene (Ffar2) expression is higher in pancreatic islets during pregnancy. Using female Ffar2-/- mice, we explored the physiological relevance of signaling through this GPCR and found that Ffar2-deficient female mice developed fasting hyperglycemia and impaired glucose tolerance in the setting of impaired insulin secretion compared with WT mice during, but not before, pregnancy. Insulin tolerance tests were similar in Ffar2-/- and WT mice before and during pregnancy. Next, we examined the role of FFA2 in gestational β-cell mass, observing that Ffar2-/- mice had diminished gestational expansion of β-cells during pregnancy. Interestingly, mouse genotype had no significant impact on the composition of the gut microbiome, but did affect the observed SCFA profiles, suggesting a functional difference in the microbiota. Altogether, these results suggest a potential link between increased Ffar2 expression in islets and the alteration of circulating SCFA levels, possibly explaining how changes in the gut microbiome contribute to gestational glucose homeostasis.

  13. Overexpression of Poplar Pyrabactin Resistance-Like Abscisic Acid Receptors Promotes Abscisic Acid Sensitivity and Drought Resistance in Transgenic Arabidopsis.

    PubMed

    Yu, Jingling; Yang, Lei; Liu, Xiaobing; Tang, Renjie; Wang, Yuan; Ge, Haiman; Wu, Mengting; Zhang, Jiang; Zhao, Fugeng; Luan, Sheng; Lan, Wenzhi

    2016-01-01

    Drought stress is an important environmental factor limiting productivity of plants, especially fast growing species with high water consumption like poplar. Abscisic acid (ABA) is a phytohormone that positively regulates seed dormancy and drought resistance. The PYR1 (Pyrabactin Resistance 1)/ PYRL (PYR-Like)/ RCAR (Regulatory Component of ABA Receptor) (PYR/PYL/RCAR) ABA receptor family has been identified and widely characterized in Arabidopsis thaliana. However, their functions in poplars remain unknown. Here, we report that 2 of 14 PYR/PYL/RCAR orthologues in poplar (Populus trichocarpa) (PtPYRLs) function as a positive regulator of the ABA signal transduction pathway. The Arabidopsis transient expression and yeast two-hybrid assays showed the interaction among PtPYRL1 and PtPYRL5, a clade A protein phosphatase 2C, and a SnRK2, suggesting that a core signalling complex for ABA signaling pathway exists in poplars. Phenotypic analysis of PtPYRL1 and PtPYRL5 transgenic Arabidopsis showed that these two genes positively regulated the ABA responses during the seed germination. More importantly, the overexpression of PtPYRL1 and PtPYRL5 substantially improved ABA sensitivity and drought stress tolerance in transgenic plants. In summary, we comprehensively uncovered the properties of PtPYRL1 and PtPYRL5, which might be good target genes to genetically engineer drought-Resistant plants.

  14. Overexpression of Poplar Pyrabactin Resistance-Like Abscisic Acid Receptors Promotes Abscisic Acid Sensitivity and Drought Resistance in Transgenic Arabidopsis

    PubMed Central

    Liu, Xiaobing; Tang, Renjie; Wang, Yuan; Ge, Haiman; Wu, Mengting; Zhang, Jiang; Zhao, Fugeng; Luan, Sheng; Lan, Wenzhi

    2016-01-01

    Drought stress is an important environmental factor limiting productivity of plants, especially fast growing species with high water consumption like poplar. Abscisic acid (ABA) is a phytohormone that positively regulates seed dormancy and drought resistance. The PYR1 (Pyrabactin Resistance 1)/ PYRL (PYR-Like)/ RCAR (Regulatory Component of ABA Receptor) (PYR/PYL/RCAR) ABA receptor family has been identified and widely characterized in Arabidopsis thaliana. However, their functions in poplars remain unknown. Here, we report that 2 of 14 PYR/PYL/RCAR orthologues in poplar (Populus trichocarpa) (PtPYRLs) function as a positive regulator of the ABA signal transduction pathway. The Arabidopsis transient expression and yeast two-hybrid assays showed the interaction among PtPYRL1 and PtPYRL5, a clade A protein phosphatase 2C, and a SnRK2, suggesting that a core signalling complex for ABA signaling pathway exists in poplars. Phenotypic analysis of PtPYRL1 and PtPYRL5 transgenic Arabidopsis showed that these two genes positively regulated the ABA responses during the seed germination. More importantly, the overexpression of PtPYRL1 and PtPYRL5 substantially improved ABA sensitivity and drought stress tolerance in transgenic plants. In summary, we comprehensively uncovered the properties of PtPYRL1 and PtPYRL5, which might be good target genes to genetically engineer drought-Resistant plants. PMID:27992471

  15. Bioorthogonal click chemistry to assay mu-opioid receptor palmitoylation using 15-hexadecynoic acid and immunoprecipitation

    PubMed Central

    Ebersole, Brittany; Petko, Jessica; Levenson, Robert

    2014-01-01

    We have developed a modification of bioorthogonal click chemistry to assay the palmitoylation of cellular proteins. This assay utilizes 15-hexadecynoic acid (15-HDYA) as a chemical probe in combination with protein immunoprecipitation using magnetic beads in order to detect S-palmitoylation of proteins of interest. Here we demonstrate the utility of this approach for the mu-opioid receptor (MOR), a GPCR responsible for mediating the analgesic and addictive properties of most clinically relevant opioid agonist drugs. This technique provides a rapid, non-isotopic, and efficient method to assay the palmitoylation status of a variety of cellular proteins including most GPCRs. PMID:24463015

  16. Retinoic Acid-Related Orphan Receptors (RORs): Regulatory Functions in Immunity, Development, Circadian Rhythm, and Metabolism

    PubMed Central

    Cook, Donald N.; Kang, Hong Soon; Jetten, Anton M.

    2015-01-01

    In this overview, we provide an update on recent progress made in understanding the mechanisms of action, physiological functions, and roles in disease of retinoic acid related orphan receptors (RORs). We are particularly focusing on their roles in the regulation of adaptive and innate immunity, brain function, retinal development, cancer, glucose and lipid metabolism, circadian rhythm, metabolic and inflammatory diseases and neuropsychiatric disorders. We also summarize the current status of ROR agonists and inverse agonists, including their regulation of ROR activity and their therapeutic potential for management of various diseases in which RORs have been implicated. PMID:26878025

  17. Aminosulfhydryl and Aminodisulfide Compounds Enhance Binding of the Glucocorticoid Receptor Complex to Deoxyribonucleic Acid-Coated Cellulose and to Chromatin

    DTIC Science & Technology

    1993-01-01

    glucocorticoid receptor [21]. Diaminosulfhydryl chloroacetic acid was obtained from the Fisher compounds are more active at enhancing GRC Scientific...phase consisting of 0. I M BASE containing 25mM KCI and 3 mM chloroacetic acid and 5mM d/-10-camphorsul- MgCI2, pH 7.6 at 0 0C) was added to each tube...Enhance Binding of the Glucocorticoid Receptor Complex to Deoxy- ribonucleic Acid -Coated Cellulose and to Chromatin 4. AUThOR(S)’ J.M. Karle, R. Olmeda and

  18. A Conserved Aspartic Acid Is Important for Agonist (VUAA1) and Odorant/Tuning Receptor-Dependent Activation of the Insect Odorant Co-Receptor (Orco)

    PubMed Central

    Kumar, Brijesh N.; Taylor, Robert W.; Pask, Gregory M.; Zwiebel, Laurence J.; Newcomb, Richard D.; Christie, David L.

    2013-01-01

    Insect odorant receptors function as heteromeric odorant-gated cation channels comprising a conventional odorant-sensitive tuning receptor, and a conserved co-receptor (Orco). An Orco agonist, VUAA1, is able to activate both heteromeric and homomeric Orco-containing channels. Very little is known about specific residues in Orco that contribute to cation permeability and gating. We investigated the importance of two conserved Asp residues, one in each of transmembrane domains 5 and 7, for channel function by mutagenesis. Drosophila melanogaster Orco and its substitution mutants were expressed in HEK cells and VUAA1-stimulated channel activity was determined by Ca2+ influx and whole-cell patch clamp electrophysiology. Substitution of D466 in transmembrane 7 with amino acids other than glutamic acid resulted in a substantial reduction in channel activity. The D466E Orco substitution mutant was ∼2 times more sensitive to VUAA1. The permeability of the D466E Orco mutant to cations was unchanged relative to wild-type Orco. When D466E Orco is co-expressed with a conventional tuning odorant receptor, the heteromeric complex also shows increased sensitivity to an odorant. Thus, the effect of the D466E mutation is not specific to VUAA1 agonism or dependent on homomeric Orco assembly. We suggest the gain-of-activation characteristic of the D466E mutant identifies an amino acid that is likely to be important for activation of both heteromeric and homomeric insect odorant receptor channels. PMID:23894621

  19. Loss of Nuclear Receptor SHP Impairs but Does Not Eliminate Negative Feedback Regulation of Bile Acid Synthesis

    PubMed Central

    Kerr, Thomas A.; Saeki, Shigeru; Schneider, Manfred; Schaefer, Karen; Berdy, Sara; Redder, Thadd; Shan, Bei; Russell, David W.; Schwarz, Margrit

    2014-01-01

    Summary The in vivo role of the nuclear receptor SHP in feedback regulation of bile acid synthesis was examined. Loss of SHP in mice caused abnormal accumulation and increased synthesis of bile acids due to derepression of rate-limiting CYP7A1 and CYP8B1 hydroxylase enzymes in the biosynthetic pathway. Dietary bile acids induced liver damage and restored feedback regulation. A synthetic agonist of the nuclear receptor FXR was not hepatotoxic and had no regulatory effects. Reduction of the bile acid pool with cholestyramine enhanced CYP7A1 and CYP8B1 expression. We conclude that input from three negative regulatory pathways controls bile acid synthesis. One is mediated by SHP, and two are SHP independent and invoked by liver damage and changes in bile acid pool size. PMID:12062084

  20. Hydroxy monounsaturated fatty acids as agonists for peroxisome proliferator-activated receptors.

    PubMed

    Yokoi, Hiroshi; Mizukami, Hajime; Nagatsu, Akito; Tanabe, Hiroki; Inoue, Makoto

    2010-01-01

    The physiological and pathological role of oxidized polyunsaturated fatty acids (PUFAs) has been extensively studied, whereas those of hydroxy monounsaturated fatty acids (MUFAs) are not well understood. This study demonstrated that 11-hydroxy-(9Z)-octadecenoic acid ((9Z)-11-HOE), which was isolated from adlay seeds (Coix lacryma-jobi L. var. ma-yuen STAF.), can activate peroxisome proliferator-activated receptor (PPAR)alpha, delta and gamma in luciferase reporter assays more efficiently than (9Z)-octadecenoic acid (oleic acid), and to the same degree as linoleic acid. (9Z)-11-HOE increased the mRNA levels of UCP2 and CD36 in C2C12 myotubes and THP- 1 cells, respectively, and these effects were blocked by the PPARdelta- and gamma-specific antagonists GSK0660 and T0070907, respectively. Evaluation of the structure.activity relationship between hydroxy MUFAs and PPAR activation revealed that (9E)-11-HOE, the geometrical isomer of (9Z)-11-HOE, activated PPARs more potently than (9Z)-11-HOE, and that PPAR activation by hydroxyl MUFAs was not markedly influenced by the position of the hydroxy group or the double bond, although PPARdelta seemed to possess ligand specificity different to that of PPARalpha or gamma . Additionally, the finding that 11-hydroxy octadecanoic acid, the hydrogenated product of (9E)-11- HOE, was also capable of activating PPARs to a similar extent as (9E)-11-HOE indicates that the double bond in hydroxy MUFAs is not essential for PPAR activation. In conclusion, (9Z)-11-HOE derived from alday seeds and hydroxy MUFAs with a chain length of 16 or 18 acted as PPAR agonists. Hydroxylation of MUFAs may change these compounds from silent PPAR ligands to active PPAR agonists.

  1. Synthesis and Activity of Dafachronic Acid Ligands for the C. elegans DAF-12 Nuclear Hormone Receptor

    PubMed Central

    Sharma, Kamalesh K.; Wang, Zhu; Motola, Daniel L.; Cummins, Carolyn L.; Mangelsdorf, David J.; Auchus, Richard J.

    2009-01-01

    The nuclear hormone receptor DAF-12 from Caenorhabditis elegans is activated by dafachronic acids, which derive from sterols upon oxidation by DAF-9, a cytochrome P450. DAF-12 activation is a critical checkpoint in C. elegans for acquisition of reproductive competence and for entry into adulthood rather than dauer diapause. Previous studies implicated the (25S)-Δ7-dafachronic acid isomer as the most potent compound, but the (25S)-Δ4-isomer was also identified as an activator of DAF-12. To explore the tolerance of DAF-12 for structural variations in the ligand and to enable further studies requiring large amounts of ligands for DAF-12 and homologs in other nematodes, we synthesized (25R)- and (25S)-isomers of five dafachronic acids differing in A/B-ring configurations. Both the (25S)- and (25R)-Δ7-dafachronic acids are potent transcriptional activators in a Gal4-transactivation assay using HEK-293 cells, with EC50 values of 23 and 33 nm, respectively, as are (25S)- and (25R)-Δ4-dafachronic acids, with EC50 values of 23 and 66 nm, respectively. The (25S)- and (25R)-Δ5-isomers were much less potent, with EC50 values approaching 1000 nm, and saturated 5α- and 5β-dafachronic acids showed mostly intermediate potencies. Rescue assays using daf- 9-null mutants confirmed the results from transactivation experiments, but this in vivo assay accentuated the greater potencies of the (25S)-epimers, particularly for the (25S)-Δ7-isomer. We conclude that DAF-12 accommodates a large range of structural variation in ligand geometry, but (25S)-Δ7-dafachronic acid is the most potent and probably biologically relevant isomer. Potency derives more from the A/B-ring configuration than from the stereochemistry at C-25. PMID:19196833

  2. Phosphatase inhibitors remove the run-down of γ-aminobutyric acid type A receptors in the human epileptic brain

    PubMed Central

    Palma, E.; Ragozzino, D. A.; Di Angelantonio, S.; Spinelli, G.; Trettel, F.; Martinez-Torres, A.; Torchia, G.; Arcella, A.; Di Gennaro, G.; Quarato, P. P.; Esposito, V.; Cantore, G.; Miledi, R.; Eusebi, F.

    2004-01-01

    The properties of γ-aminobutyric acid (GABA) type A receptors (GABAA receptors) microtransplanted from the human epileptic brain to the plasma membrane of Xenopus oocytes were compared with those recorded directly from neurons, or glial cells, in human brains slices. Cell membranes isolated from brain specimens, surgically obtained from six patients afflicted with drug-resistant temporal lobe epilepsy (TLE) were injected into frog oocytes. Within a few hours, these oocytes acquired GABAA receptors that generated GABA currents with an unusual run-down, which was inhibited by orthovanadate and okadaic acid. In contrast, receptors derived from membranes of a nonepileptic hippocampal uncus, membranes from mouse brain, or recombinant rat α1β2γ2-GABA receptors exhibited a much less pronounced GABA-current run-down. Moreover, the GABAA receptors of pyramidal neurons in temporal neocortex slices from the same six epileptic patients exhibited a stronger run-down than the receptors of rat pyramidal neurons. Interestingly, the GABAA receptors of neighboring glial cells remained substantially stable after repetitive activation. Therefore, the excessive GABA-current run-down observed in the membrane-injected oocytes recapitulates essentially what occurs in neurons, rather than in glial cells. Quantitative RT-PCR analyses from the same TLE neocortex specimens revealed that GABAA-receptor β1, β2, β3, and γ2 subunit mRNAs were significantly overexpressed (8- to 33-fold) compared with control autopsy tissues. Our results suggest that an abnormal GABA-receptor subunit transcription in the TLE brain leads to the expression of run-down-enhanced GABAA receptors. Blockage of phosphatases stabilizes the TLE GABAA receptors and strengthens GABAergic inhibition. It may be that this process can be targeted to develop new treatments for intractable epilepsy. PMID:15218107

  3. Abscisic acid (ABA) receptors: light at the end of the tunnel

    PubMed Central

    McCormick, Sheila

    2010-01-01

    The plant hormone abscisic acid (ABA) plays a role in several aspects of plant growth and development. Understanding how this hormonal stimulus is sensed and transduced turned out to be one of the major tasks in the field of plant signaling. A series of recent papers proposed several different proteins that could receive the ABA signal and initiate the signaling cascade. The winner appears to be PYR/PYL/RCAR (PYrabactin Resistance/PYrabactin Resistance-Like/Regulatory Component of Abscisic acid Receptor) proteins, as crystal structures were recently published. The crystal structures support the idea that upon ABA binding to a PYR/PYL/RCAR protein, the activity of a phosphatase 2C, with known repressive activity on ABA signaling, is inhibited. PMID:20948817

  4. Eicosapentaenoic acid (EPA) induces peroxisome proliferator-activated receptors and ameliorates experimental autoimmune encephalomyelitis.

    PubMed

    Unoda, Kiichi; Doi, Yoshimitsu; Nakajima, Hideto; Yamane, Kazushi; Hosokawa, Takafumi; Ishida, Shimon; Kimura, Fumiharu; Hanafusa, Toshiaki

    2013-03-15

    Eicosapentaenoic acid (EPA), one of the n-3 polyunsaturated fatty acids, is a neuroprotective lipid with anti-inflammatory properties. We investigated the possible therapeutic effect of EPA on experimental autoimmune encephalomyelitis (EAE). EAE mice were fed a diet with or without EPA. The clinical EAE scores of the EPA-fed mice were significantly lower than those of the non-EPA mice. In the EPA-treated mice, IFN-γ and IL-17 productions were remarkably inhibited and the expression levels of peroxisome proliferator-activated receptors were significantly enhanced in the CNS-infiltrating CD4T cells. Thus EPA shows promise as a potential new therapeutic agent against multiple sclerosis.

  5. A novel role for the dioxin receptor in fatty acid metabolism and hepatic steatosis

    PubMed Central

    Lee, Jung Hoon; Wada, Taira; Febbraio, Maria; He, Jinhan; Matsubara, Tsutomu; Lee, Min Jae; Gonzalez, Frank J.; Xie, Wen

    2010-01-01

    Background & Aims The aryl hydrocarbon receptor (AhR) is a PAS domain transcription factor previously known as the “dioxin receptor” or “xenobiotic receptor.” The goal of this study is to determine the endobiotic role of AhR in hepatic steatosis. Methods Wild type, constitutively activated AhR (CA-AhR) transgenic, AhR null (AhR-/-), and fatty acid translocase CD36/FAT null (CD36-/-) mice were used to investigate the role of AhR in steatosis and the involvement of CD36 in the steatotic effect of AhR. The promoters of the mouse and human CD36 genes were cloned and their regulation by AhR was analyzed. Results Activation of AhR induced spontaneous hepatic steatosis characterized by the accumulation of triglycerides. The steatotic effect of AhR is likely due to the combined upregulation of CD36 and fatty acid transport proteins (FATPs), suppression of fatty acid oxidation, inhibition of hepatic export of triglycerides, increase in peripheral fat mobilization, and increased hepatic oxidative stress. Promoter analysis established CD36 as a novel transcriptional target of AhR. Activation of AhR in liver cells induced CD36 gene expression and enhanced fatty acid uptake. The steatotic effect of an AhR agonist was inhibited in CD36-/- mice. Conclusions Our study reveals a novel link between AhR-induced steatosis and the expression of CD36. Industrial or military exposures to dioxin and related compounds have been linked to increased prevalence of fatty liver in humans. Results from this study may help to establish AhR and its target CD36 as novel therapeutic and preventive targets for fatty liver disease. PMID:20303349

  6. Human sweet taste receptor mediates acid-induced sweetness of miraculin.

    PubMed

    Koizumi, Ayako; Tsuchiya, Asami; Nakajima, Ken-ichiro; Ito, Keisuke; Terada, Tohru; Shimizu-Ibuka, Akiko; Briand, Loïc; Asakura, Tomiko; Misaka, Takumi; Abe, Keiko

    2011-10-04

    Miraculin (MCL) is a homodimeric protein isolated from the red berries of Richadella dulcifica. MCL, although flat in taste at neutral pH, has taste-modifying activity to convert sour stimuli to sweetness. Once MCL is held on the tongue, strong sweetness is sensed over 1 h each time we taste a sour solution. Nevertheless, no molecular mechanism underlying the taste-modifying activity has been clarified. In this study, we succeeded in quantitatively evaluating the acid-induced sweetness of MCL using a cell-based assay system and found that MCL activated hT1R2-hT1R3 pH-dependently as the pH decreased from 6.5 to 4.8, and that the receptor activation occurred every time an acid solution was applied. Although MCL per se is sensory-inactive at pH 6.7 or higher, it suppressed the response of hT1R2-hT1R3 to other sweeteners at neutral pH and enhanced the response at weakly acidic pH. Using human/mouse chimeric receptors and molecular modeling, we revealed that the amino-terminal domain of hT1R2 is required for the response to MCL. Our data suggest that MCL binds hT1R2-hT1R3 as an antagonist at neutral pH and functionally changes into an agonist at acidic pH, and we conclude this may cause its taste-modifying activity.

  7. Human sweet taste receptor mediates acid-induced sweetness of miraculin

    PubMed Central

    Koizumi, Ayako; Tsuchiya, Asami; Nakajima, Ken-ichiro; Ito, Keisuke; Terada, Tohru; Shimizu-Ibuka, Akiko; Briand, Loïc; Asakura, Tomiko; Misaka, Takumi; Abe, Keiko

    2011-01-01

    Miraculin (MCL) is a homodimeric protein isolated from the red berries of Richadella dulcifica. MCL, although flat in taste at neutral pH, has taste-modifying activity to convert sour stimuli to sweetness. Once MCL is held on the tongue, strong sweetness is sensed over 1 h each time we taste a sour solution. Nevertheless, no molecular mechanism underlying the taste-modifying activity has been clarified. In this study, we succeeded in quantitatively evaluating the acid-induced sweetness of MCL using a cell-based assay system and found that MCL activated hT1R2-hT1R3 pH-dependently as the pH decreased from 6.5 to 4.8, and that the receptor activation occurred every time an acid solution was applied. Although MCL per se is sensory-inactive at pH 6.7 or higher, it suppressed the response of hT1R2-hT1R3 to other sweeteners at neutral pH and enhanced the response at weakly acidic pH. Using human/mouse chimeric receptors and molecular modeling, we revealed that the amino-terminal domain of hT1R2 is required for the response to MCL. Our data suggest that MCL binds hT1R2-hT1R3 as an antagonist at neutral pH and functionally changes into an agonist at acidic pH, and we conclude this may cause its taste-modifying activity. PMID:21949380

  8. Activation of a tunicate (Ciona intestinalis) xenobiotic receptor orthologue by both natural toxins and synthetic toxicants.

    PubMed

    Fidler, Andrew E; Holland, Patrick T; Reschly, Erica J; Ekins, Sean; Krasowski, Matthew D

    2012-02-01

    Vertebrate xenobiotic receptors are ligand-activated nuclear receptors (NRs) that bind exogenous biologically active chemicals before activating the transcription of genes involved in xenobiotic metabolism and excretion. Typically, xenobiotic receptors have ligand binding domains (LBDs) that can accommodate a structurally diverse array of molecules and in addition display high levels of inter-taxa sequence diversity suggestive of positive selection. Pursuing the idea that xenobiotic receptors may adaptively evolve to bind toxic chemicals commonly present in an organism's environment/diet, we examined ligand binding by a xenobiotic receptor orthologue of a marine filter-feeding organism. The solitary tunicate Ciona intestinalis (Phylum Chordata) genome encodes an orthologue of the vertebrate pregnane X receptor (PXR) and vitamin D receptor (VDR), here denoted CiVDR/PXRα. In a luciferase reporter assay the CiVDR/PXRα was activated, at nanomolar concentrations, by two of four natural marine microalgal biotoxins tested (okadaic acid, EC50 = 18.2 ± 0.9 nM and pectenotoxin-2, EC50 = 37.0 ± 3.5 nM) along with 1 of 11 synthetic toxicants (esfenvalerate: EC50 = 0.59 ± 0.7 μM). Two related C. intestinalis NRs, orthologous to vertebrate farnesoid X receptor and liver X receptors, respectively, along with the PXR of a freshwater fish (zebrafish, Danio rerio), were not activated by any of the 15 chemicals tested. In contrast, human PXR was activated by okadaic acid at similar concentrations to CiVDR/PXRα (EC50 = 7.2 ± 1.1 nM) but not by pectenotoxin-2. A common features pharmacophore developed for the CiVDR/PXRα ligand consisted of an off-center hydrogen bond acceptor flanked by two hydrophobic regions. The results of this study are consistent with the original hypothesis that natural toxins, present in the diet of filter-feeding marine invertebrates, may have acted as selective agents in the molecular evolution of tunicate xenobiotic receptors. Bioassays based on

  9. The retinoid X receptor in a marine invertebrate chordate: evolutionary insights from urochordates.

    PubMed

    Maeng, Sejung; Lee, Jung Hwan; Choi, Sung-Chang; Kim, Mi Ae; Shin, Yun Kyung; Sohn, Young Chang

    2012-09-01

    Retinoid X receptors (RXRs) are highly conserved members of the nuclear hormone receptor family that mediate various physiological processes in vertebrates and invertebrates. We examined the expression patterns of RXR in the ascidian Halocynthia roretzi across a wide range of tissues and stages of embryo development, as well as the regulation of gene transcription by the ascidian RXR. H. roretzi RXR cDNA (HrRXR) was cloned from 64-cell stage embryos. The overall amino acid sequence of HrRXR showed high sequence identity with a urochordate Ciona intestinalis RXR (58%), but the ligand-binding domain of HrRXR was more similar to vertebrate orthologs than to those of invertebrate RXRs. Based on a phylogenetic analysis, HrRXR belongs to a group of urochordates that are separate from vertebrate RXRs, showing a clear evolutionary history. Real-time quantitative polymerase chain reaction and whole-mount in situ hybridization analyses revealed that the HrRXR mRNA is of maternal origin during embryogenesis, and in the examined adult tissues it is expressed in the muscles, gills, gonads, and the hepatopancreas. Immunofluorescence and immunohistochemical staining demonstrated that HrRXR is localized to the nucleus and highly expressed in the gills and hepatopancreas. Unlike human RXRα, HrRXR did not show 9-cis retinoic acid- and bexarotene (LGD1069)-dependent transactivation. While a synthetic ligand for farnesoid X receptor (FXR), GW4064, did not increase the transcriptional activation in HrRXR- or HrRXR/HrFXR-transfected HEK-293 cells, the ligand showed weak but significant activity for a single amino acid mutant of HrRXR ((Phe)231(Cys)) and HrFXR cotransfected cells. The present study suggests that the marine invertebrate chordate RXR may possess endogenous ligands that are different than vertebrate RXR ligands and which function during early embryonic stages.

  10. Structural analysis of binding functionality of folic acid-PEG dendrimers against folate receptor.

    PubMed

    Sampogna-Mireles, Diana; Araya-Durán, Ingrid D; Márquez-Miranda, Valeria; Valencia-Gallegos, Jesús A; González-Nilo, Fernando D

    2017-03-01

    Dendrimers functionalized with folic acid (FA) are drug delivery systems that can selectively target cancer cells with folate receptors (FR-α) overexpression. Incorporation of polyethylene glycol (PEG) can enhance dendrimers solubility and pharmacokinetics, but ligand-receptor binding must not be affected. In this work we characterized, at atomic level, the binding functionality of conventional site-specific dendrimers conjugated with FA with PEG 750 or PEG 3350 as a linker. After Molecular Dynamics simulation, we observed that both PEG's did not interfere over ligand-receptor binding functionality. Although binding kinetics could be notably affected, the folate fragment from both dendrimers remained exposed to the solvent before approaching selectively to FR-α. PEG 3350 provided better solubility and protection from enzymatic degradation to the dendrimer than PEG 750. Also, FA-PEG3350 dendrimer showed a slightly better interaction with FR-α than FA-PEG750 dendrimer. Therefore, theoretical evidence supports that both dendrimers are suitable as drug delivery systems for cancer therapies.

  11. Variant Ionotropic Receptors in the Malaria Vector Mosquito Anopheles gambiae Tuned to Amines and Carboxylic Acids

    PubMed Central

    Pitts, R. Jason; Derryberry, Stephen L.; Zhang, Zhiwei; Zwiebel, Laurence J.

    2017-01-01

    The principal Afrotropical human malaria vector mosquito, Anopheles gambiae, remains a significant threat to global health. A critical component in the transmission of malaria is the ability of An. gambiae females to detect and respond to human-derived chemical kairomones in their search for blood meal hosts. The basis for host odor responses resides in olfactory receptor neurons (ORNs) that express chemoreceptors encoded by large gene families, including the odorant receptors (ORs) and the variant ionotropic receptors (IRs). While ORs have been the focus of extensive investigation, functional IR complexes and the chemical compounds that activate them have not been identified in An. gambiae. Here we report the transcriptional profiles and functional characterization of three An. gambiae IR (AgIr) complexes that specifically respond to amines or carboxylic acids - two classes of semiochemicals that have been implicated in mediating host-seeking by adult females but are not known to activate An. gambiae ORs (AgOrs). Our results suggest that AgIrs play critical roles in the detection and behavioral responses to important classes of host odors that are underrepresented in the AgOr chemical space. PMID:28067294

  12. Pharmacodynamics of long-acting folic acid-receptor targeted ritonavir boosted atazanavir nanoformulations

    PubMed Central

    Puligujja, Pavan; Balkundi, Shantanu; Kendrick, Lindsey; Baldridge, Hannah; Hilaire, James; Bade, Aditya N.; Dash, Prasanta K.; Zhang, Gang; Poluektova, Larisa; Gorantla, Santhi; Liu, Xin-Ming; Ying, Tianlei; Feng, Yang; Wang, Yanping; Dimitrov, Dimiter S.; McMillan, JoEllyn M.; Gendelman, Howard E.

    2014-01-01

    Long-acting nanoformulated antiretroviral therapy (nanoART) that target monocyte-macrophage could improve the drug’s half-life and protein binding capacities while facilitating cell and tissue depots. To this end, ART nanoparticles that target the folic acid (FA) receptor and permit cell-based drug depots were examined using pharmacokinetic and pharmacodynamic (PD) tests. FA receptor-targeted poloxamer 407 nanocrystals, containing ritonavir-boosted atazanavir (ATV/r), significantly affected several therapeutic factors: drug bioavailability increased as much as 5 times and PD activity improved as much as 100 times. Drug particles administered to human peripheral blood lymphocyte reconstituted NOD.Cg-PrkdcscidIl2rgtm1Wjl/SzJ mice and infected with HIV-1ADA at a tissue culture infective dose50 of 104 infectious viral particles/ml led to ATV/r drug concentrations that paralleled FA receptor beta staining in both the macrophage-rich parafollicular areas of spleen and lymph nodes. Drug levels were higher in these tissues than what could be achieved by either native drug or untargeted nanoART particles. The data also mirrored potent reductions in viral loads, tissue viral RNA and numbers of HIV-1p24+ cells in infected and treated animals. We conclude that FA-P407 coating of ART nanoparticles readily facilitate drug carriage and facilitate antiretroviral responses. PMID:25522973

  13. Pharmacodynamics of long-acting folic acid-receptor targeted ritonavir-boosted atazanavir nanoformulations.

    PubMed

    Puligujja, Pavan; Balkundi, Shantanu S; Kendrick, Lindsey M; Baldridge, Hannah M; Hilaire, James R; Bade, Aditya N; Dash, Prasanta K; Zhang, Gang; Poluektova, Larisa Y; Gorantla, Santhi; Liu, Xin-Ming; Ying, Tianlei; Feng, Yang; Wang, Yanping; Dimitrov, Dimiter S; McMillan, JoEllyn M; Gendelman, Howard E

    2015-02-01

    Long-acting nanoformulated antiretroviral therapy (nanoART) that targets monocyte-macrophages could improve the drug's half-life and protein-binding capacities while facilitating cell and tissue depots. To this end, ART nanoparticles that target the folic acid (FA) receptor and permit cell-based drug depots were examined using pharmacokinetic and pharmacodynamic (PD) tests. FA receptor-targeted poloxamer 407 nanocrystals, containing ritonavir-boosted atazanavir (ATV/r), significantly increased drug bioavailability and PD by five and 100 times, respectively. Drug particles administered to human peripheral blood lymphocyte reconstituted NOD.Cg-Prkdc(scid)Il2rg(tm1Wjl)/SzJ mice and infected with HIV-1ADA led to ATV/r drug concentrations that paralleled FA receptor beta staining in both the macrophage-rich parafollicular areas of spleen and lymph nodes. Drug levels were higher in these tissues than what could be achieved by either native drug or untargeted nanoART particles. The data also mirrored potent reductions in viral loads, tissue viral RNA and numbers of HIV-1p24+ cells in infected and treated animals. We conclude that FA-P407 coating of ART nanoparticles readily facilitates drug carriage and antiretroviral responses.

  14. Salvianolic Acid A, as a Novel ETA Receptor Antagonist, Shows Inhibitory Effects on Tumor in Vitro.

    PubMed

    Zhang, Qiao; Wang, Shifeng; Yu, Yangyang; Sun, Shengnan; Zhang, Yuxin; Zhang, Yanling; Yang, Wei; Li, Shiyou; Qiao, Yanjiang

    2016-08-02

    Endothelin-1 (ET-1) autocrine and paracrine signaling modulate cell proliferation of tumor cells by activating its receptors, endothelin A receptor (ETAR) and endothelin B receptor (ETBR). Dysregulation of ETAR activation promotes tumor development and progression. The potential of ETAR antagonists and the dual-ETAR and ETBR antagonists as therapeutic approaches are under preclinical and clinical studies. Salvianolic acid A (Sal A) is a hydrophilic polyphenolic derivative isolated from Salvia miltiorrhiza Bunge (Danshen), which has been reported as an anti-cancer and cardio-protective herbal medicine. In this study, we demonstrate that Sal A inhibits ETAR activation induced by ET-1 in both recombinant and endogenous ETAR expression cell lines. The IC50 values were determined as 5.7 µM in the HEK293/ETAR cell line and 3.14 µM in HeLa cells, respectively. Furthermore, our results showed that Sal A suppressed cell proliferation and extended the doubling times of multiple cancer cells, including HeLa, DU145, H1975, and A549 cell lines. In addition, Sal A inhibited proliferation of DU145 cell lines stimulated by exogenous ET-1 treatment. Moreover, the cytotoxicity and cardio-toxicity of Sal A were assessed in human umbilical vein endothelial cells (HUVEC) and Human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs), which proved that Sal A demonstrates no cytotoxicity or cardiotoxicity. Collectively, our findings indicate that Sal A is a novel anti-cancer candidate through targeting ETAR.

  15. Activation of Lysophosphatidic Acid Receptor Type 1 Contributes to Pathophysiology of Spinal Cord Injury

    PubMed Central

    Santos-Nogueira, Eva; López-Serrano, Clara; Hernández, Joaquim; Lago, Natalia; Astudillo, Alma M.; Balsinde, Jesús; Estivill-Torrús, Guillermo; de Fonseca, Fernando Rodriguez; Chun, Jerold

    2015-01-01

    Lysophosphatidic acid (LPA) is an extracellular lipid mediator involved in many physiological functions that signals through six known G-protein-coupled receptors (LPA1–LPA6). A wide range of LPA effects have been identified in the CNS, including neural progenitor cell physiology, astrocyte and microglia activation, neuronal cell death, axonal retraction, and development of neuropathic pain. However, little is known about the involvement of LPA in CNS pathologies. Herein, we demonstrate for the first time that LPA signaling via LPA1 contributes to secondary damage after spinal cord injury. LPA levels increase in the contused spinal cord parenchyma during the first 14 d. To model this potential contribution of LPA in the spinal cord, we injected LPA into the normal spinal cord, revealing that LPA induces microglia/macrophage activation and demyelination. Use of a selective LPA1 antagonist or mice lacking LPA1 linked receptor-mediated signaling to demyelination, which was in part mediated by microglia. Finally, we demonstrate that selective blockade of LPA1 after spinal cord injury results in reduced demyelination and improvement in locomotor recovery. Overall, these results support LPA–LPA1 signaling as a novel pathway that contributes to secondary damage after spinal cord contusion in mice and suggest that LPA1 antagonism might be useful for the treatment of acute spinal cord injury. SIGNIFICANCE STATEMENT This study reveals that LPA signaling via LPA receptor type 1 activation causes demyelination and functional deficits after spinal cord injury. PMID:26180199

  16. Alisol B 23-acetate protects against non-alcoholic steatohepatitis in mice via farnesoid X receptor activation

    PubMed Central

    Meng, Qiang; Duan, Xing-ping; Wang, Chang-yuan; Liu, Zhi-hao; Sun, Peng-yuan; Huo, Xiao-kui; Sun, Hui-jun; Peng, Jin-yong; Liu, Ke-xin

    2017-01-01

    Alisol B 23-acetate (AB23A) is a natural triterpenoid isolated from the traditional Chinese medicine rhizoma alismatis, which exhibits a number of pharmacological activities, including anti-hepatitis virus, anti-cancer and antibacterial effects. In this study we examined whether AB23A protected against non-alcoholic steatohepatitis (NASH) in mice, and the mechanisms underlying the protective effects. NASH was induced in mice fed a methionine and choline-deficient (MCD) diet for 4 weeks. The mice were simultaneously treated with AB23A (15, 30, and 60 mg·kg−1·d−1, ig) for 4 weeks. On the last day, blood samples and livers were collected. Serum liver functional enzymes, inflammatoru markers were assessed. The livers were histologically examined using H&E, Oil Red O, Masson's trichrome and Sirius Red staining. Mouse primary hepatocytes were used for in vitro experiments. The mechanisms underlying AB23A protection were analyzed using siRNA, qRT-PCR, and Western blot assays. AB23A treatment significantly and dose-dependently decreased the elevated levels of serum ALT and AST in MCD diet-fed mice. Furthermore, AB23A treatment significantly reduced hepatic triglyceride accumulation, inflammatory cell infiltration and hepatic fibrosis in the mice. AB23A-induced decreases in serum and hepatic lipids were related to decreased hepatic lipogenesis through decreasing hepatic levels of SREBP-1c, FAS, ACC1 and SCD1 and increased lipid metabolism via inducing PPARα, CPT1α, ACADS and LPL. The reduction in inflammatory cell infiltration corresponded to deceased serum levels of mKC and MCP-1 and decreased hepatic gene expression of MCP-1 and VCAM-1. The reduction in hepatic fibrosis was correlated with decreased hepatic gene expression of fibrosis markers. The protective effects of AB23A were FXR-dependent, because treatment with the FXR agonist CDCA mimicked AB23A-induced hepato-protection in the mice, whereas co-administration of FXR antagonist guggulsterone abrogated AB23A-induced hepato-protection. In mouse primary hepatocytes, FXR gene silencing abrogated AB23A-induced changes in gene expression of Apo C-II, CPT1α, ACADS and LPL. AB23A produces protective effects against NASH in mice via FXR activation. PMID:27773935

  17. Neuroendocrine factors regulate retinoic acid receptors in normal and hypoplastic lung development

    PubMed Central

    Pereira-Terra, Patrícia; Moura, Rute S; Nogueira-Silva, Cristina; Correia-Pinto, Jorge

    2015-01-01

    Congenital diaphragmatic hernia (CDH) is characterised by a spectrum of lung hypoplasia and consequent pulmonary hypertension, leading to high morbidity and mortality rates. Moreover, CDH has been associated with an increase in the levels of pulmonary neuroendocrine factors, such as bombesin and ghrelin, and a decrease in the action of retinoic acid (RA). The present study aimed to elucidate the interaction between neuroendocrine factors and RA. In vitro analyses were performed on Sprague–Dawley rat embryos. Normal lung explants were treated with bombesin, ghrelin, a bombesin antagonist, a ghrelin antagonist, dimethylsulfoxide (DMSO), RA dissolved in DMSO, bombesin plus RA and ghrelin plus RA. Hypoplastic lung explants (nitrofen model) were cultured with bombesin, ghrelin, bombesin antagonist or ghrelin antagonist. The lung explants were analysed morphometrically, and retinoic acid receptor (RAR) α, β and γ expression levels were assessed via Western blotting. Immunohistochemistry analysis of RAR was performed in normal and hypoplastic lungs 17.5 days post-conception (dpc). Compared with the controls, hypoplastic lungs exhibited significantly higher RARα/γ expression levels. Furthermore considering hypoplastic lungs, bombesin and ghrelin antagonists decreased RARα/γ expression. Normal lung explants (13.5 dpc) treated with RA, bombesin plus RA, ghrelin plus RA, bombesin or ghrelin exhibited increased lung growth. Moreover, bombesin and ghrelin increased RARα/γ expression levels, whereas the bombesin and ghrelin antagonists decreased RARα/γ expression. This study demonstrates for the first time that neuroendocrine factors function as lung growth regulators, sensitising the lung to the action of RA through up-regulation of RARα and RARγ. Key points Retinoic acid (RA) and ghrelin levels are altered in human hypoplastic lungs when compared to healthy lungs. Although considerable data have been obtained about RA, ghrelin and bombesin in the congenital

  18. Mutations of lysophosphatidic acid receptor-1 gene during progression of lung tumors in rats

    SciTech Connect

    Yamada, Takanori; Obo, Yumi; Furukawa, Mami; Hotta, Mayuko; Yamasaki, Ayako; Honoki, Kanya; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

    2009-01-16

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. In this study, mutations of lysophosphatidic acid receptor-1 (LPA1) gene were investigated to clarify the possible molecular mechanisms underlying the development of lung tumors induced by N-nitrosobis(2-hydroxypropyl)amine (BHP) in rats. Male Wistar rats, 6 weeks of age, were given 2000 ppm BHP in their drinking water for 12 weeks and then maintained without further treatment until sacrifice at 25 weeks. Genomic DNAs were extracted from paraffin-embedded tissues and exons 2-4 were examined for mutations, using polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) analysis. No LPA1 mutations were detected in 15 hyperplasias, but 2 out of 12 adenomas (16.7%) and 7 out of 17 adenocarcinomas (41.2%). These results suggest that mutations of LPA1 gene may be involved in the acquisition of growth advantage from adenomas to adenocarcinomas in lung carcinogenesis induced in rats by BHP.

  19. Kynurenic acid, an aryl hydrocarbon receptor ligand, is elevated in serum of Zucker fatty rats

    PubMed Central

    Oxenkrug, G; Cornicelli, J; van der Hart, M; Roeser, J; Summergrad, P

    2016-01-01

    Obesity is an increasingly urgent global problem and the molecular mechanisms of obesity are not fully understood. Dysregulation of the tryptophan (Trp) – kynurenine (Kyn) metabolic pathway (TKP) have been suggested as a mechanism of obesity and described in obese humans and in animal models of obesity. However, to the best of our knowledge, TKP metabolism has not been studied in leptin-receptor-deficient Zucker fatty rats (ZFR) (fa/fa), the best-known and most widely used rat model of obesity. We were interested to determine if there are any deviations of TKP in ZFR. Concentrations of major TKP metabolites were evaluated (HPLC- MS method) in serum of ZFR (fa/fa) and age-matched lean rats (FA/-). Concentrations of kynurenic acid (KYNA) were 50% higher in ZFR than in lean rats (p<0.004, Mann-Whitney two-tailed test). Anthranilic acid (AA) concentrations, while elevated by 33%, did not reach statistical significance (p<0.04, one-tailed test). Elevated KYNA serum concentrations might contribute to development of obesity via KYNA-induced activation of aryl hydrocarbon receptor. Present results warrant further studies of KYNA and AA in ZFR and other animal models of obesity. PMID:27738521

  20. Regulation of GABA-modulin phosphorylation and GABA receptor binding by excitatory amino acids

    SciTech Connect

    Vaccarino, F.; Guidotti, A.

    1987-05-01

    Primary cultures of cerebellar granule cells phosphorylate numerous proteins including GABA-modulin (GM), which is a putative allosteric modulator of GABA receptors. Cell depolarization and treatment with dicarboxylic excitatory amino acids, which activate PI turnover, Ca/sup 2 +/ influx and guanylate cyclase in granule cells increase the phosphorylation of specific proteins. To determine GM phosphorylation by endogenous protein kinases in living granule cell cultures, GM was isolated by immunoprecipitation and reverse-phase HPLC. High K/sup +/, veratridine, glutamate and NMDA treatment stimulated GM phosphorylation over 2-fold. This increase was abolished by the absence of extracellular Ca/sup 2 +/ and was antagonized by Mg/sup 2 +/ ions and by AVP. The excitatory amino acid action was mimicked by phorbol esters but not by forskolin or by cGMP, and thus may be mediated by an activation of protein kinase C (PKC). Moreover, excitatory amino acids increase /sup 3/H-labelled phorbol ester binding sites in granule cell membrane. The same cultures, treated with glutamate or kainate, showed a 50-fold greater efficacy of muscimol for the stimulation of benzodiazepine (BZ) binding. These data-suggest that excitatory amino acid stimulation of neurons triggers PKC translocation and the activated enzyme phosphorylates GM. The extent of GM phosphorylation may regulate the coupling between GABA and BZ binding sites.

  1. Acid-responsive PEGylated doxorubicin prodrug nanoparticles for neuropilin-1 receptor-mediated targeted drug delivery.

    PubMed

    Song, Huijuan; Zhang, Ju; Wang, Weiwei; Huang, Pingsheng; Zhang, Yumin; Liu, Jianfeng; Li, Chen; Kong, Deling

    2015-12-01

    Self-assembled prodrug nanoparticles have demonstrated great promise in cancer chemotherapy. In the present study, we developed a new kind of prodrug nanoparticles for targeted drug delivery. PEGylated doxorubicin conjugate with an acid-cleavable cis-aconityl spacer was prepared. Then it was functionalized with a tumor-penetrating peptide, Cys-Arg-Gly-Asp-Lys (CRGDK), providing the prodrug nanoparticles with the specific binding ability to neurophilin-1 receptor. In acid mediums, doxorubicin could be released from the prodrug nanoparticles with an accumulative release around 60% through the acid-triggered hydrolysis of cis-aconityl bond and nanoparticle disassembly. Whereas, drug release was slow under a neutral pH and the accumulative drug release was less than 16%. In the cell culture tests, our prodrug nanoparticles showed enhanced endocytosis and cytotoxicity in cancer cells including HepG2, MCF-7 and MDA-MB-231 cells, but lower cytotoxicity in human cardiomyocyte H2C9. In the animal experiments, the prodrug nanoparticles were intravenously injected into Balb/c nude mice bearing MDA-MB-231 tumors. Enhanced drug penetration and accumulation in tumors, accompanying with a rapid early tumor-binding behavior, was observed after intravenous injection of the peptide modified prodrug nanoparticles. These data suggests that the acid-sensitive and tumor-targeting PEGylated doxorubicin prodrug nanoparticle may be an efficient drug delivery system for cancer chemotherapy.

  2. Acidic tumor microenvironment and pH-sensing G protein-coupled receptors.

    PubMed

    Justus, Calvin R; Dong, Lixue; Yang, Li V

    2013-12-05

    The tumor microenvironment is acidic due to glycolytic cancer cell metabolism, hypoxia, and deficient blood perfusion. It is proposed that acidosis in the tumor microenvironment is an important stress factor and selection force for cancer cell somatic evolution. Acidic pH has pleiotropic effects on the proliferation, migration, invasion, metastasis, and therapeutic response of cancer cells and the function of immune cells, vascular cells, and other stromal cells. However, the molecular mechanisms by which cancer cells and stromal cells sense and respond to acidic pH in the tumor microenvironment are poorly understood. In this article the role of a family of pH-sensing G protein-coupled receptors (GPCRs) in tumor biology is reviewed. Recent studies show that the pH-sensing GPCRs, including GPR4, GPR65 (TDAG8), GPR68 (OGR1), and GPR132 (G2A), regulate cancer cell metastasis and proliferation, immune cell function, inflammation, and blood vessel formation. Activation of the proton-sensing GPCRs by acidosis transduces multiple downstream G protein signaling pathways. Since GPCRs are major drug targets, small molecule modulators of the pH-sensing GPCRs are being actively developed and evaluated. Research on the pH-sensing GPCRs will continue to provide important insights into the molecular interaction between tumor and its acidic microenvironment and may identify new targets for cancer therapy and chemoprevention.

  3. A Novel Allosteric Activator of Free Fatty Acid 2 Receptor Displays Unique Gi-functional Bias*

    PubMed Central

    Bolognini, Daniele; Moss, Catherine E.; Nilsson, Karolina; Petersson, Annika U.; Donnelly, Iona; Sergeev, Eugenia; König, Gabriele M.; Kostenis, Evi; Kurowska-Stolarska, Mariola; Miller, Ashley; Dekker, Niek; Tobin, Andrew B.

    2016-01-01

    The short chain fatty acid receptor FFA2 is able to stimulate signaling via both Gi- and Gq/G11-promoted pathways. These pathways are believed to control distinct physiological end points but FFA2 receptor ligands appropriate to test this hypothesis have been lacking. Herein, we characterize AZ1729, a novel FFA2 regulator that acts as a direct allosteric agonist and as a positive allosteric modulator, increasing the activity of the endogenously produced short chain fatty acid propionate in Gi-mediated pathways, but not at those transduced by Gq/G11. Using AZ1729 in combination with direct inhibitors of Gi and Gq/G11 family G proteins demonstrated that although both arms contribute to propionate-mediated regulation of phospho-ERK1/2 MAP kinase signaling in FFA2-expressing 293 cells, the Gq/G11-mediated pathway is predominant. We extend these studies by employing AZ1729 to dissect physiological FFA2 signaling pathways. The capacity of AZ1729 to act at FFA2 receptors to inhibit β-adrenoreceptor agonist-promoted lipolysis in primary mouse adipocytes and to promote chemotaxis of isolated human neutrophils confirmed these as FFA2 processes mediated by Gi signaling, whereas, in concert with blockade by the Gq/G11 inhibitor FR900359, the inability of AZ1729 to mimic or regulate propionate-mediated release of GLP-1 from mouse colonic preparations defined this physiological response as an end point transduced via activation of Gq/G11. PMID:27385588

  4. Triphenyl phosphate-induced developmental toxicity in zebrafish: Potential role of the retinoic acid receptor

    PubMed Central

    Isales, Gregory M.; Hipszer, Rachel A.; Raftery, Tara D.; Chen, Albert; Stapleton, Heather M.; Volz, David C.

    2015-01-01

    Using zebrafish as a model, we previously reported that developmental exposure to triphenyl phosphate (TPP) – a high-production volume organophosphate-based flame retardant – results in dioxin-like cardiac looping impairments that are independent of the aryl hydrocarbon receptor. Using a pharmacologic approach, the objective of this study was to investigate the potential role of retinoic acid receptor (RAR) – a nuclear receptor that regulates vertebrate heart morphogenesis – in mediating TPP-induced developmental toxicity in zebrafish. We first revealed that static exposure of zebrafish from 5-72 hours post-fertilization (hpf) to TPP in the presence of non-toxic concentrations of an RAR antagonist (BMS493) significantly enhanced TPP-induced toxicity (relative to TPP alone), even though identical non-toxic BMS493 concentrations mitigated retinoic acid (RA)-induced toxicity. BMS493-mediated enhancement of TPP toxicity was not a result of differential TPP uptake or metabolism, as internal embryonic doses of TPP and diphenyl phosphate (DPP) – a primary TPP metabolite - were not different in the presence or absence of BMS493. Using real-time PCR, we then quantified the relative change in expression of cytochrome P450 26a1 (cyp26a1) – a major target gene for RA-induced RAR activation in zebrafish – and found that RA and TPP exposure resulted in a ∼5-fold increase and decrease in cyp26a1 expression, respectively, relative to vehicle-exposed embryos. To address whether TPP may interact with human RARs, we then exposed Chinese hamster ovary cells stably transfected with chimeric human RARα-, RARβ-, or RARγ to TPP in the presence of RA, and found that TPP significantly inhibited RA-induced luciferase activity in a concentration-dependent manner. Overall, our findings suggest that zebrafish RARs may be involved in mediating TPP-induced developmental toxicity, a mechanism of action that may have relevance to humans. PMID:25725299

  5. Efficient Modulation of γ-Aminobutyric Acid Type A Receptors by Piperine Derivatives

    PubMed Central

    2014-01-01

    Piperine activates TRPV1 (transient receptor potential vanilloid type 1 receptor) receptors and modulates γ-aminobutyric acid type A receptors (GABAAR). We have synthesized a library of 76 piperine analogues and analyzed their effects on GABAAR by means of a two-microelectrode voltage-clamp technique. GABAAR were expressed in Xenopus laevis oocytes. Structure–activity relationships (SARs) were established to identify structural elements essential for efficiency and potency. Efficiency of piperine derivatives was significantly increased by exchanging the piperidine moiety with either N,N-dipropyl, N,N-diisopropyl, N,N-dibutyl, p-methylpiperidine, or N,N-bis(trifluoroethyl) groups. Potency was enhanced by replacing the piperidine moiety by N,N-dibutyl, N,N-diisobutyl, or N,N-bistrifluoroethyl groups. Linker modifications did not substantially enhance the effect on GABAAR. Compound 23 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dipropyl-2,4-pentadienamide] induced the strongest modulation of GABAA (maximal GABA-induced chloride current modulation (IGABA-max = 1673% ± 146%, EC50 = 51.7 ± 9.5 μM), while 25 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dibutyl-2,4-pentadienamide] displayed the highest potency (EC50 = 13.8 ± 1.8 μM, IGABA-max = 760% ± 47%). Compound 23 induced significantly stronger anxiolysis in mice than piperine and thus may serve as a starting point for developing novel GABAAR modulators. PMID:24905252

  6. Bone marrow stromal cell transplantation preserves gammaaminobutyric acid receptor function in the injured spinal cord.

    PubMed

    Yano, Shunsuke; Kuroda, Satoshi; Shichinohe, Hideo; Seki, Toshitaka; Ohnishi, Takako; Tamagami, Hiroshi; Hida, Kazutoshi; Iwasaki, Yoshinobu

    2006-11-01

    A surprising shortage of information surrounds the mechanisms by which bone marrow stromal cells (BMSC) restore lost neurologic functions when transplanted into the damaged central nervous system. In the present study, we sought to elucidate whether BMSCs express the neuron-specific gamma-aminobutyric acid (GABA) receptor when transplanted into injured spinal cord. To examine this, we harvested and cultured rat femoral BMSCs. We then subjected Sprague-Dawley rats to thoracic spinal cord injury (SCI) with a pneumatic impact device. Fluorescence-labeled BMSCs (n = 7) were transplanted stereotactically or the vehicle in which these cells were cultured (n = 4) was introduced stereotactically into the rostral site of SCI at 7 days after injury. We evaluated GABA receptor function by measuring the binding potential for 125I-iomazenil (125I-IMZ) through in vitro autoradiography at 4 weeks after BMSC transplantation and simultaneously examined the fate of the transplanted BMSCs by immunocytochemistry. We found that the transplanted BMSC migrated toward the core of the injury and were densely distributed in the marginal region at 4 weeks after transplantation. BMSC transplantation significantly increased the binding potential for 125I-IMZ (p = 0.0376) and increased the number of GABA receptor-positive cells (p = 0.0077) in the marginal region of the injury site. Some of the transplanted BMSCs were positive for microtubule-associated protein-2 and the alpha1 subunit of GABA(A) receptor in the region of injury. These findings suggest that BMSCs have the potential to support the survival of neurons in the marginal region of SCI and can partly differentiate into neurons, regenerating spinal cord tissue at the site of injury.

  7. Nicotinic acid-adenine dinucleotide phosphate activates the skeletal muscle ryanodine receptor.

    PubMed Central

    Hohenegger, Martin; Suko, Josef; Gscheidlinger, Regina; Drobny, Helmut; Zidar, Andreas

    2002-01-01

    Calcium is a universal second messenger. The temporal and spatial information that is encoded in Ca(2+)-transients drives processes as diverse as neurotransmitter secretion, axonal outgrowth, immune responses and muscle contraction. Ca(2+)-release from intracellular Ca(2+) stores can be triggered by diffusible second messengers like Ins P (3), cyclic ADP-ribose or nicotinic acid-adenine dinucleotide phosphate (NAADP). A target has not yet been identified for the latter messenger. In the present study we show that nanomolar concentrations of NAADP trigger Ca(2+)-release from skeletal muscle sarcoplasmic reticulum. This was due to a direct action on the Ca(2+)-release channel/ryanodine receptor type-1, since in single channel recordings, NAADP increased the open probability of the purified channel protein. The effects of NAADP on Ca(2+)-release and open probability of the ryanodine receptor occurred over a similar concentration range (EC(50) approximately 30 nM) and were specific because (i) they were blocked by Ruthenium Red and ryanodine, (ii) the precursor of NAADP, NADP, was ineffective at equimolar concentrations, (iii) NAADP did not affect the conductance and reversal potential of the ryanodine receptor. Finally, we also detected an ADP-ribosyl cyclase activity in the sarcoplasmic reticulum fraction of skeletal muscle. This enzyme was not only capable of synthesizing cyclic GDP-ribose but also NAADP, with an activity of 0.25 nmol/mg/min. Thus, we conclude that NAADP is generated in the vicinity of type 1 ryanodine receptor and leads to activation of this ion channel. PMID:12102654

  8. Modulation of NMDA receptor function by inhibition of D-amino acid oxidase in rodent brain.

    PubMed

    Strick, Christine A; Li, Cheryl; Scott, Liam; Harvey, Brian; Hajós, Mihály; Steyn, Stefanus J; Piotrowski, Mary A; James, Larry C; Downs, James T; Rago, Brian; Becker, Stacey L; El-Kattan, Ayman; Xu, Youfen; Ganong, Alan H; Tingley, F David; Ramirez, Andres D; Seymour, Patricia A; Guanowsky, Victor; Majchrzak, Mark J; Fox, Carol B; Schmidt, Christopher J; Duplantier, Allen J

    2011-01-01

    Observations that N-Methyl-D-Aspartate (NMDA) antagonists produce symptoms in humans that are similar to those seen in schizophrenia have led to the current hypothesis that schizophrenia might result from NMDA receptor hypofunction. Inhibition of D-amino acid oxidase (DAAO), the enzyme responsible for degradation of D-serine, should lead to increased levels of this co-agonist at the NMDA receptor, and thereby provide a therapeutic approach to schizophrenia. We have profiled some of the preclinical biochemical, electrophysiological, and behavioral consequences of administering potent and selective inhibitors of DAAO to rodents to begin to test this hypothesis. Inhibition of DAAO activity resulted in a significant dose and time dependent increase in D-serine only in the cerebellum, although a time delay was observed between peak plasma or brain drug concentration and cerebellum D-serine response. Pharmacokinetic/pharmacodynamic (PK/PD) modeling employing a mechanism-based indirect response model was used to characterize the correlation between free brain drug concentration and D-serine accumulation. DAAO inhibitors had little or no activity in rodent models considered predictive for antipsychotic activity. The inhibitors did, however, affect cortical activity in the Mescaline-Induced Scratching model, produced a modest but significant increase in NMDA receptor-mediated synaptic currents in primary neuronal cultures from rat hippocampus, and resulted in a significant increase in evoked hippocampal theta rhythm, an in vivo electrophysiological model of hippocampal activity. These findings demonstrate that although DAAO inhibition did not cause a measurable increase in D-serine in forebrain, it did affect hippocampal and cortical activity, possibly through augmentation of NMDA receptor-mediated currents.

  9. Megalin and cubilin expression in gallbladder epithelium and regulation by bile acids.

    PubMed

    Erranz, Benjamín; Miquel, Juan Francisco; Argraves, W Scott; Barth, Jeremy L; Pimentel, Fernando; Marzolo, María-Paz

    2004-12-01

    Cholesterol crystal formation in the gallbladder is a key step in gallstone pathogenesis. Gallbladder epithelial cells might prevent luminal gallstone formation through a poorly understood cholesterol absorption process. Genetic studies in mice have highlighted potential gallstone susceptibility alleles, Lith genes, which include the gene for megalin. Megalin, in conjunction with the large peripheral membrane protein cubilin, mediates the endocytosis of numerous ligands, including HDL/apolipoprotein A-I (apoA-I). Although the bile contains apoA-I and several cholesterol-binding megalin ligands, the expression of megalin and cubilin in the gallbladder has not been investigated. Here, we show that both proteins are expressed by human and mouse gallbladder epithelia. In vitro studies using a megalin-expressing cell line showed that lithocholic acid strongly inhibits and cholic and chenodeoxycholic acids increase megalin expression. The effects of bile acids (BAs) were also demonstrated in vivo, analyzing gallbladder levels of megalin and cubilin from mice fed with different BAs. The BA effects could be mediated by the farnesoid X receptor, expressed in the gallbladder. Megalin protein was also strongly increased after feeding a lithogenic diet. These results indicate a physiological role for megalin and cubilin in the gallbladder and provide support for a role for megalin in gallstone pathogenesis.

  10. γ-Aminobutyric Acid B Receptor Mediated Inhibition of Gonadotropin-Releasing Hormone Neurons Is Suppressed by Kisspeptin-G Protein-Coupled Receptor 54 Signaling

    PubMed Central

    Zhang, Chunguang; Bosch, Martha A.; Rønnekleiv, Oline K.; Kelly, Martin J.

    2009-01-01

    γ-Aminobutyric acid (GABA) is one of the most important neurotransmitters that regulate the excitability of GnRH neurons. Numerous studies have shown that GABA activates Cl− currents in GnRH neurons, and these effects are antagonized by GABAA receptor antagonists. The GABAB receptor is a heterodimer composed of GABAB R1 and R2, and although both subunits have been localized in GnRH neurons, nothing is known about the cellular signaling of this Gαi,o-coupled receptor in GnRH neurons. Using whole-cell recordings from mouse enhanced green fluorescent protein-GnRH neurons, we found that the GABAB receptor agonist baclofen hyperpolarized GnRH neurons through activation of an inwardly rectifying K+ current in a concentration-dependent manner. The effects of baclofen were antagonized by the selective GABAB receptor antagonist CGP 52432 with a Ki (inhibitory constant) of 85 nm. Furthermore, in the presence of the GABAA receptor antagonist picrotoxin, GABA hyperpolarized GnRH neurons in a similar manner. Treatment with 17β-estradiol as compared with oil vehicle did not significantly alter either the EC50 for the baclofen-induced response (0.8 ± 0.1 vs. 1.0 ± 0.1 μm, respectively) or the maximal outward current (10.8 ± 1.7 pA vs. 11.4 ± 0.6 pA, respectively) in GnRH neurons. However, the outward current (and membrane hyperpolarization) was abrogated by submaximal concentrations of the G protein-coupled receptor 54 (GPR54) agonist kisspeptin-10 in both groups, indicating that Gαq-coupled (GPR54) can desensitize the GABAB receptor-mediated response. Therefore, the activation of GABAB receptors in GnRH neurons may provide increased inhibitory tone during estrogen-negative feedback states that is attenuated by kisspeptin during positive feedback. PMID:19164470

  11. Activities of natural methyl farnesoids on pupariation and metamorphosis of Drosophila melanogaster

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Methyl farnesoate (MF) and juvenile hormone (JH III), which respectively bind to the receptors USP and MET, and bisepoxy JH III (bisJHIII) were assessed for several activities during Drosophila larval development, and during prepupal development to eclosed adults. Dietary MF and JH III were similar...

  12. Evidence for a bladder cell glycolipid receptor for Escherichia coli and the effect of neuraminic acid and colominic acid on adherence.

    PubMed Central

    Davis, C P; Avots-Avotins, A E; Fader, R C

    1981-01-01

    The rat bladder epithelial cell receptors involved in mannose-sensitive adherence of Escherichia coli strains were studied. Sodium metaperiodate and lipase pretreatment of epithelial cells significantly reduced bacterial adherence to cells whereas trypsin and phospholipase C had a marginal or insignificant effect on adherence. Neuraminidase and colominic acid significantly increased adherence, whereas N-acetylneuraminic acid significantly reduced adherence. These data suggest that the rat bladder epithelial cell receptors involved in mannose-sensitive adherence are glycolipids. In addition, the data suggested that sialic acid on bladder epithelial cells acts as a nonspecific inhibitor of adherence, whereas colominic acid, a component of some E. coli K1 capsules, may act as a promoter of adherence. PMID:6277793

  13. Bile acid signaling in lipid metabolism: metabolomic and lipidomic analysis of lipid and bile acid markers linked to anti-obesity and anti-diabetes in mice.

    PubMed

    Qi, Yunpeng; Jiang, Changtao; Cheng, Jie; Krausz, Kristopher W; Li, Tiangang; Ferrell, Jessica M; Gonzalez, Frank J; Chiang, John Y L

    2015-01-01

    Bile acid synthesis is the major pathway for catabolism of cholesterol. Cholesterol 7α-hydroxylase (CYP7A1) is the rate-limiting enzyme in the bile acid biosynthetic pathway in the liver and plays an important role in regulating lipid, glucose and energy metabolism. Transgenic mice overexpressing CYP7A1 (CYP7A1-tg mice) were resistant to high-fat diet (HFD)-induced obesity, fatty liver, and diabetes. However the mechanism of resistance to HFD-induced obesity of CYP7A1-tg mice has not been determined. In this study, metabolomic and lipidomic profiles of CYP7A1-tg mice were analyzed to explore the metabolic alterations in CYP7A1-tg mice that govern the protection against obesity and insulin resistance by using ultra-performance liquid chromatography-coupled with electrospray ionization quadrupole time-of-flight mass spectrometry combined with multivariate analyses. Lipidomics analysis identified seven lipid markers including lysophosphatidylcholines, phosphatidylcholines, sphingomyelins and ceramides that were significantly decreased in serum of HFD-fed CYP7A1-tg mice. Metabolomics analysis identified 13 metabolites in bile acid synthesis including taurochenodeoxycholic acid, taurodeoxycholic acid, tauroursodeoxycholic acid, taurocholic acid, and tauro-β-muricholic acid (T-β-MCA) that differed between CYP7A1-tg and wild-type mice. Notably, T-β-MCA, an antagonist of the farnesoid X receptor (FXR) was significantly increased in intestine of CYP7A1-tg mice. This study suggests that reducing 12α-hydroxylated bile acids and increasing intestinal T-β-MCA may reduce high fat diet-induced increase of phospholipids, sphingomyelins and ceramides, and ameliorate diabetes and obesity. This article is part of a Special Issue entitled Linking transcription to physiology in lipodomics.

  14. Discovery of novel dihydrobenzofuran cyclopropane carboxylic acid based calcium sensing receptor antagonists for the treatment of osteoporosis.

    PubMed

    Liang, Gui-Bai; Zhou, Changyou; Huo, Xianghong; Wang, Hank; Yang, Xuelin; Huang, Shaoqiang; Wang, Haisheng; Wilkinson, Hilary; Luo, Lusong; Tang, Wei; Sutton, David; Li, Hong; Zaller, Dennis; Meinke, Peter T

    2016-08-15

    In a search for novel small molecule calcium-sensing receptor (CaSR) antagonists as oral bone anabolic agents, we discovered dihydrobenzofuran cyclopropane carboxylic acid derivatives, such as 12f (IC50=27.6nM), are highly potent calcium-sensing receptor antagonists. Studies in rats established that compound 12f stimulates parathyroid hormone (PTH) release in a fast-acting, pulsatile manner.

  15. Postsynaptic clustering of γ-aminobutyric acid type A receptors by the γ3 subunit in vivo

    PubMed Central

    Baer, Kristin; Essrich, Christian; Benson, Jack A.; Benke, Dietmar; Bluethmann, Horst; Fritschy, Jean-Marc; Lüscher, Bernhard

    1999-01-01

    Synaptic localization of γ-aminobutyric acid type A (GABAA) receptors is a prerequisite for synaptic inhibitory function, but the mechanism by which different receptor subtypes are localized to postsynaptic sites is poorly understood. The γ2 subunit and the postsynaptic clustering protein gephyrin are required for synaptic localization and function of major GABAA receptor subtypes. We now show that transgenic overexpression of the γ3 subunit in γ2 subunit-deficient mice restores benzodiazepine binding sites, benzodiazepine-modulated whole cell currents, and postsynaptic miniature currents, suggesting the formation of functional, postsynaptic receptors. Moreover, the γ3 subunit can substitute for γ2 in the formation of GABAA receptors that are synaptically clustered and colocalized with gephyrin in vivo. These clusters were formed even in brain regions devoid of endogenous γ3 subunit, indicating that the factors present for clustering of γ2 subunit-containing receptors are sufficient to cluster γ3 subunit-containing receptors. The GABAA receptor and gephyrin-clustering properties of the ectopic γ3 subunit were also observed for the endogenous γ3 subunit, but only in the absence of the γ2 subunit, suggesting that the γ3 subunit is at a competitive disadvantage with the γ2 subunit for clustering of postsynaptic GABAA receptors in wild-type mice. PMID:10536013

  16. Allosteric Regulation in the Ligand Binding Domain of Retinoic Acid Receptorγ

    PubMed Central

    Amal, Ismail; Lutzing, Régis; Stote, Roland H.; Rochette-Egly, Cécile; Rochel, Natacha; Dejaegere, Annick

    2017-01-01

    Retinoic acid (RA) plays key roles in cell differentiation and growth arrest through nuclear retinoic acid receptors (RARs), which are ligand-dependent transcription factors. While the main trigger of RAR activation is the binding of RA, phosphorylation of the receptors has also emerged as an important regulatory signal. Phosphorylation of the RARγ N-terminal domain (NTD) is known to play a functional role in neuronal differentiation. In this work, we investigated the phosphorylation of RARγ ligand binding domain (LBD), and present evidence that the phosphorylation status of the LBD affects the phosphorylation of the NTD region. We solved the X-ray structure of a phospho-mimetic mutant of the LBD (RARγ S371E), which we used in molecular dynamics simulations to characterize the consequences of the S371E mutation on the RARγ structural dynamics. Combined with simulations of the wild-type LBD, we show that the conformational equilibria of LBD salt bridges (notably R387-D340) are affected by the S371E mutation, which likely affects the recruitment of the kinase complex that phosphorylates the NTD. The molecular dynamics simulations also showed that a conservative mutation in this salt bridge (R387K) affects the dynamics of the LBD without inducing large conformational changes. Finally, cellular assays showed that the phosphorylation of the NTD of RARγ is differentially regulated by retinoic acid in RARγWT and in the S371N, S371E and R387K mutants. This multidisciplinary work highlights an allosteric coupling between phosphorylations of the LBD and the NTD of RARγ and supports the importance of structural dynamics involving electrostatic interactions in the regulation of RARs activity. PMID:28125680

  17. Expanding Duplication of Free Fatty Acid Receptor-2 (GPR43) Genes in the Chicken Genome.

    PubMed

    Meslin, Camille; Desert, Colette; Callebaut, Isabelle; Djari, Anis; Klopp, Christophe; Pitel, Frédérique; Leroux, Sophie; Martin, Pascal; Froment, Pascal; Guilbert, Edith; Gondret, Florence; Lagarrigue, Sandrine; Monget, Philippe

    2015-04-24

    Free fatty acid receptors (FFAR) belong to a family of five G-protein coupled receptors that are involved in the regulation of lipid metabolism, so that their loss of function increases the risk of obesity. The aim of this study was to determine the expansion of genes encoding paralogs of FFAR2 in the chicken, considered as a model organism for developmental biology and biomedical research. By estimating the gene copy number using quantitative polymerase chain reaction, genomic DNA resequencing, and RNA sequencing data, we showed the existence of 23 ± 1.5 genes encoding FFAR2 paralogs in the chicken genome. The FFAR2 paralogs shared an identity from 87.2% up to 99%. Extensive gene conversion was responsible for this high degree of sequence similarities between these genes, and this concerned especially the four amino acids known to be critical for ligand binding. Moreover, elevated nonsynonymous/synonymous substitution ratios on some amino acids within or in close-vicinity of the ligand-binding groove suggest that positive selection may have reduced the effective rate of gene conversion in this region, thus contributing to diversify the function of some FFAR2 paralogs. All the FFAR2 paralogs were located on a microchromosome in a same linkage group. FFAR2 genes were expressed in different tissues and cells such as spleen, peripheral blood mononuclear cells, abdominal adipose tissue, intestine, and lung, with the highest rate of expression in testis. Further investigations are needed to determine whether these chicken-specific events along evolution are the consequence of domestication and may play a role in regulating lipid metabolism in this species.

  18. Metabotropic glutamate receptors are involved in the detection of IMP and L-amino acids by mouse taste sensory cells.

    PubMed

    Pal Choudhuri, S; Delay, R J; Delay, E R

    2016-03-01

    G-protein-coupled receptors are thought to be involved in the detection of umami and L-amino acid taste. These include the heterodimer taste receptor type 1 member 1 (T1r1)+taste receptor type 1 member 3 (T1r3), taste and brain variants of mGluR4 and mGluR1, and calcium sensors. While several studies suggest T1r1+T1r3 is a broadly tuned lLamino acid receptor, little is known about the function of metabotropic glutamate receptors (mGluRs) in L-amino acid taste transduction. Calcium imaging of isolated taste sensory cells (TSCs) of T1r3-GFP and T1r3 knock-out (T1r3 KO) mice was performed using the ratiometric dye Fura 2 AM to investigate the role of different mGluRs in detecting various L-amino acids and inosine 5' monophosphate (IMP). Using agonists selective for various mGluRs such as (RS)-3,5-dihydroxyphenylglycine (DHPG) (an mGluR1 agonist) and L-(+)-2-amino-4-phosphonobutyric acid (l-AP4) (an mGluR4 agonist), we evaluated TSCs to determine if they might respond to these agonists, IMP, and three L-amino acids (monopotassium L-glutamate, L-serine and L-arginine). Additionally, we used selective antagonists against different mGluRs such as (RS)-L-aminoindan-1,5-dicarboxylic acid (AIDA) (an mGluR1 antagonist), and (RS)-α-methylserine-O-phosphate (MSOP) (an mGluR4 antagonist) to determine if they can block responses elicited by these L-amino acids and IMP. We found that L-amino acid- and IMP-responsive cells also responded to each agonist. Antagonists for mGluR4 and mGluR1 significantly blocked the responses elicited by IMP and each of the L-amino acids. Collectively, these data provide evidence for the involvement of taste and brain variants of mGluR1 and mGluR4 in L-amino acid and IMP taste responses in mice, and support the concept that multiple receptors contribute to IMP and L-amino acid taste.

  19. [Relationship between the crystal lattice structure and the biological action of some agonists of amino acid receptors].

    PubMed

    Kertser, L S; Baev, K V

    1992-01-01

    The crystal structures of glycine, taurine, GABA, beta-alanine were compared. The quantity and the accuracy of distances coincidence between nitrogen and oxygen atoms were used as a criterion of similarity of the crystalline structures. The conclusion is made about a correlation between crystalline structure of agonists and their effect on amino acid receptors. It is assumed that in case of a cooperative effect of agonist on the receptor a mutual arrangement of molecules on the receptor surface is similar to their arrangement in the agonist crystal.

  20. Agonist-directed signaling of serotonin 5-HT2C receptors: differences between serotonin and lysergic acid diethylamide (LSD).

    PubMed

    Backstrom, J R; Chang, M S; Chu, H; Niswender, C M; Sanders-Bush, E

    1999-08-01

    For more than 40 years the hallucinogen lysergic acid diethylamide (LSD) has been known to modify serotonin neurotransmission. With the advent of molecular and cellular techniques, we are beginning to understand the complexity of LSD's actions at the serotonin 5-HT2 family of receptors. Here, we discuss evidence that signaling of LSD at 5-HT2C receptors differs from the endogenous agonist serotonin. In addition, RNA editing of the 5-HT2C receptor dramatically alters the ability of LSD to stimulate phosphatidylinositol signaling. These findings provide a unique opportunity to understand the mechanism(s) of partial agonism.

  1. Concentration-dependent mode of interaction of angiotensin II receptor blockers with uric acid transporter.

    PubMed

    Iwanaga, Takashi; Sato, Masanobu; Maeda, Tomoji; Ogihara, Toshio; Tamai, Ikumi

    2007-01-01

    Serum uric acid (SUA) is currently recognized as a risk factor for cardiovascular disease. It has been reported that an angiotensin II receptor blocker (ARB), losartan, decreases SUA level, whereas other ARBs, such as candesartan, have no lowering effect. Because the renal uric acid transporter (URAT1) is an important factor controlling the SUA level, we examined the involvement of URAT1 in those differential effects of various ARBs on SUA level at clinically relevant concentrations. This study was done by using URAT1-expressing Xenopus oocytes. Losartan, pratosartan, and telmisartan exhibited cis-inhibitory effects on the uptake of uric acid by URAT1, whereas at higher concentrations, only telmisartan did, and these ARBs reduced the uptake in competitive inhibition kinetics. On the other hand, candesartan, EXP3174 [2-n-butyl-4-chloro-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yI)methyl]imidazole-5-carboxylic acid] (a major metabolite of losartan), olmesartan, and valsartan were not inhibitory. Preloading of those ARBs in the oocytes enhanced the URAT1-mediated uric acid uptake, showing a trans-stimulatory effect. The present study is a first demonstration of the differential effects of ARBs on URAT1 that some ARBs are both cis-inhibitory and trans-stimulatory, depending on concentration, whereas others exhibit either a trans-stimulatory or cis-inhibitory effect alone, which could explain the clinically observed differential effects of ARBs on SUA level. Furthermore, it was found that such differential effects of ARBs on URAT1 could be predicted from the partial chemical structures of ARBs, which will be useful information for the appropriate use and development of ARBs without an increase of SUA.

  2. (+)Lysergic acid diethylamide, but not its nonhallucinogenic congeners, is a potent serotonin 5HT1C receptor agonist

    SciTech Connect

    Burris, K.D.; Breeding, M.; Sanders-Bush, E. )

    1991-09-01

    Activation of central serotonin 5HT2 receptors is believed to be the primary mechanism whereby lysergic acid diethylamide (LSD) and other hallucinogens induce psychoactive effects. This hypothesis is based on extensive radioligand binding and electrophysiological and behavioral studies in laboratory animals. However, the pharmacological profiles of 5HT2 and 5HT1C receptors are similar, making it difficult to distinguish between effects due to activation of one or the other receptor. For this reason, it was of interest to investigate the interaction of LSD with 5HT1C receptors. Agonist-stimulated phosphoinositide hydrolysis in rat choroid plexus was used as a direct measure of 5HT1C receptor activation. (+)LSD potently stimulated phosphoinositide hydrolysis in intact choroid plexus and in cultures of choroid plexus epithelial cells, with EC50 values of 9 and 26 nM, respectively. The effect of (+)LSD in both systems was blocked by 5HT receptor antagonists with an order of activity consistent with interaction at 5HT1C receptors. Neither (+)-2-bromo-LSD nor lisuride, two nonhallucinogenic congeners of LSD, were able to stimulate 5HT1C receptors in cultured cells or intact choroid plexus. In contrast, lisuride, like (+)LSD, is a partial agonist at 5HT2 receptors in cerebral cortex slices and in NIH 3T3 cells transfected with 5HT2 receptor cDNA. The present finding that (+)LSD, but not its nonhallucinogenic congeners, is a 5HT1C receptor agonist suggests a possible role for these receptors in mediating the psychoactive effects of LSD.

  3. Critical Role for an acidic amino acid region in platelet signaling by the HemITAM (hemi-immunoreceptor tyrosine-based activation motif) containing receptor CLEC-2 (C-type lectin receptor-2).

    PubMed

    Hughes, Craig E; Sinha, Uma; Pandey, Anjali; Eble, Johannes A; O'Callaghan, Christopher A; Watson, Steve P

    2013-02-15

    CLEC-2 is a member of new family of C-type lectin receptors characterized by a cytosolic YXXL downstream of three acidic amino acids in a sequence known as a hemITAM (hemi-immunoreceptor tyrosine-based activation motif). Dimerization of two phosphorylated CLEC-2 molecules leads to recruitment of the tyrosine kinase Syk via its tandem SH2 domains and initiation of a downstream signaling cascade. Using Syk-deficient and Zap-70-deficient cell lines we show that hemITAM signaling is restricted to Syk and that the upstream triacidic amino acid sequence is required for signaling. Using surface plasmon resonance and phosphorylation studies, we demonstrate that the triacidic amino acids are required for phosphorylation of the YXXL. These results further emphasize the distinct nature of the proximal events in signaling by hemITAM relative to ITAM receptors.

  4. The bile acid sensor FXR regulates insulin transcription and secretion.

    PubMed

    Renga, Barbara; Mencarelli, Andrea; Vavassori, Piero; Brancaleone, Vincenzo; Fiorucci, Stefano

    2010-03-01

    Farnesoid X Receptor plays an important role in maintaining bile acid, cholesterol homeostasis and glucose metabolism. Here we investigated whether FXR is expressed by pancreatic beta-cells and regulates insulin signaling in pancreatic beta-cell line and human islets. We found that FXR activation induces positive regulatory effects on glucose-induced insulin transcription and secretion by genomic and non-genomic activities. Genomic effects of FXR activation relay on the induction of the glucose regulated transcription factor KLF11. Indeed, results from silencing experiments of KLF11 demonstrate that this transcription factor is essential for FXR activity on glucose-induced insulin gene transcription. In addition FXR regulates insulin secretion by non-genomic effects. Thus, activation of FXR in betaTC6 cells increases Akt phosphorylation and translocation of the glucose transporter GLUT2 at plasma membrane, increasing the glucose uptake by these cells. In vivo experiments on Non Obese Diabetic (NOD) mice demonstrated that FXR activation delays development of signs of diabetes, hyperglycemia and glycosuria, by enhancing insulin secretion and by stimulating glucose uptake by the liver. These data established that an FXR-KLF11 regulated pathway has an essential role in the regulation of insulin transcription and secretion induced by glucose.

  5. Potentiation of acid-sensing ion channel activity by peripheral group I metabotropic glutamate receptor signaling.

    PubMed

    Gan, Xiong; Wu, Jing; Ren, Cuixia; Qiu, Chun-Yu; Li, Yan-Kun; Hu, Wang-Ping

    2016-05-01

    Glutamate activates peripheral group I metabotropic glutamate receptors (mGluRs) and contributes to inflammatory pain. However, it is still not clear the mechanisms are involved in group I mGluR-mediated peripheral sensitization. Herein, we report that group I mGluRs signaling sensitizes acid-sensing ion channels (ASICs) in dorsal root ganglion (DRG) neurons and contributes to acidosis-evoked pain. DHPG, a selective group I mGluR agonist, can potentiate the functional activity of ASICs, which mediated the proton-induced events. DHPG concentration-dependently increased proton-gated currents in DRG neurons. It shifted the proton concentration-response curve upwards, with a 47.3±7.0% increase of the maximal current response to proton. Group I mGluRs, especially mGluR5, mediated the potentiation of DHPG via an intracellular cascade. DHPG potentiation of proton-gated currents disappeared after inhibition of intracellular Gq/11 proteins, PLCβ, PKC or PICK1 signaling. Moreover, DHPG enhanced proton-evoked membrane excitability of rat DRG neurons and increased the amplitude of the depolarization and the number of spikes induced by acid stimuli. Finally, peripherally administration of DHPG dose-dependently exacerbated nociceptive responses to intraplantar injection of acetic acid in rats. Potentiation of ASIC activity by group I mGluR signaling in rat DRG neurons revealed a novel peripheral mechanism underlying group I mGluRs involvement in hyperalgesia.

  6. Pore architecture and ion sites in acid-sensing ion channels and P2X receptors.

    PubMed

    Gonzales, Eric B; Kawate, Toshimitsu; Gouaux, Eric

    2009-07-30

    Acid-sensing ion channels are proton-activated, sodium-selective channels composed of three subunits, and are members of the superfamily of epithelial sodium channels, mechanosensitive and FMRF-amide peptide-gated ion channels. These ubiquitous eukaryotic ion channels have essential roles in biological activities as diverse as sodium homeostasis, taste and pain. Despite their crucial roles in biology and their unusual trimeric subunit stoichiometry, there is little knowledge of the structural and chemical principles underlying their ion channel architecture and ion-binding sites. Here we present the structure of a functional acid-sensing ion channel in a desensitized state at 3 A resolution, the location and composition of the approximately 8 A 'thick' desensitization gate, and the trigonal antiprism coordination of caesium ions bound in the extracellular vestibule. Comparison of the acid-sensing ion channel structure with the ATP-gated P2X(4) receptor reveals similarity in pore architecture and aqueous vestibules, suggesting that there are unanticipated yet common structural and mechanistic principles.

  7. Poly (ADP-ribose) glycohydrolase regulates retinoic acid receptor-mediated gene expression.

    PubMed

    Le May, Nicolas; Iltis, Izarn; Amé, Jean-Christophe; Zhovmer, Alexander; Biard, Denis; Egly, Jean-Marc; Schreiber, Valérie; Coin, Frédéric

    2012-12-14

    Poly-(ADP-ribose) glycohydrolase (PARG) is a catabolic enzyme that cleaves ADP-ribose polymers synthesized by poly-(ADP-ribose) polymerases. Here, transcriptome profiling and differentiation assay revealed a requirement of PARG for retinoic acid receptor (RAR)-mediated transcription. Mechanistically, PARG accumulates early at promoters of RAR-responsive genes upon retinoic acid treatment to promote the formation of an appropriate chromatin environment suitable for transcription. Silencing of PARG or knockout of its enzymatic activity maintains the H3K9me2 mark at the promoter of the RAR-dependent genes, leading to the absence of preinitiation complex formation. In the absence of PARG, we found that the H3K9 demethylase KDM4D/JMJD2D became PARsylated. Mutation of two glutamic acids located in the Jumonji N domain of KDM4D inhibited PARsylation. PARG becomes dispensable for ligand-dependent transcription when either a PARP inhibitor or a non-PARsylable KDM4D/JMJD2D mutant is used. Our results define PARG as a coactivator regulating chromatin remodeling during RA-dependent gene expression.

  8. International Union of Basic and Clinical Pharmacology. LXXXII: Nomenclature and Classification of Hydroxy-carboxylic Acid Receptors (GPR81, GPR109A, and GPR109B).

    PubMed

    Offermanns, Stefan; Colletti, Steven L; Lovenberg, Timothy W; Semple, Graeme; Wise, Alan; IJzerman, Adriaan P

    2011-06-01

    The G-protein-coupled receptors GPR81, GPR109A, and GPR109B share significant sequence homology and form a small group of receptors, each of which is encoded by clustered genes. In recent years, endogenous ligands for all three receptors have been described. These endogenous ligands have in common that they are hydroxy-carboxylic acid metabolites, and we therefore have proposed that this receptor family be named hydroxy-carboxylic acid (HCA) receptors. The HCA(1) receptor (GPR81) is activated by 2-hydroxy-propanoic acid (lactate), the HCA(2) receptor (GPR109A) is a receptor for the ketone body 3-hydroxy-butyric acid, and the HCA(3) receptor (GPR109B) is activated by the β-oxidation intermediate 3-hydroxy-octanoic acid. HCA(1) and HCA(2) receptors are found in most mammalian species, whereas the HCA(3) receptor is present only in higher primates. The three receptors have in common that they are expressed in adipocytes and are coupled to G(i)-type G-proteins mediating antilipolytic effects in fat cells. HCA(2) and HCA(3) receptors are also expressed in a variety of immune cells. HCA(2) is a receptor for the antidyslipidemic drug nicotinic acid (niacin) and related compounds, and there is an increasing number of synthetic ligands mainly targeted at HCA(2) and HCA(3) receptors. The aim of this article is to give an overview on the discovery and pharmacological characterization of HCAs, and to introduce an International Union of Basic and Clinical Pharmacology (IUPHAR)-recommended nomenclature. We will also discuss open questions regarding this receptor family as well as their physiological role and therapeutic potential.

  9. FXR agonist obeticholic acid reduces hepatic inflammation and fibrosis in a rat model of toxic cirrhosis

    PubMed Central

    Verbeke, Len; Mannaerts, Inge; Schierwagen, Robert; Govaere, Olivier; Klein, Sabine; Vander Elst, Ingrid; Windmolders, Petra; Farre, Ricard; Wenes, Mathias; Mazzone, Massimiliano; Nevens, Frederik; van Grunsven, Leo A.; Trebicka, Jonel; Laleman, Wim

    2016-01-01

    Hepatic inflammation drives hepatic stellate cells (HSC), resulting in liver fibrosis. The Farnesoid-X receptor (FXR) antagonizes inflammation through NF-κB inhibition. We investigated preventive and therapeutic effects of FXR agonist obeticholic acid (OCA) on hepatic inflammation and fibrosis in toxic cirrhotic rats. Cirrhosis was induced by thioacetamide (TAA) intoxication. OCA was given during or after intoxication with vehicle-treated rats as controls. At sacrifice, fibrosis, hemodynamic and biochemical parameters were assessed. HSC activation, cell turn-over, hepatic NF-κB activation, pro-inflammatory and pro-fibrotic cytokines were determined. The effect of OCA was further evaluated in isolated HSC, Kupffer cells, hepatocytes and liver sinusoidal endothelial cells (LSEC). OCA decreased hepatic inflammation and fibrogenesis during TAA-administration and reversed fibrosis in established cirrhosis. Portal pressure decreased through reduced intrahepatic vascular resistance. This was paralleled by decreased expression of pro-fibrotic cytokines (transforming growth-factor β, connective tissue growth factor, platelet-derived growth factor β-receptor) as well as markers of hepatic cell turn-over, by blunting effects of pro-inflammatory cytokines (e.g. monocyte chemo-attractant protein-1). In vitro, OCA inhibited both LSEC and Kupffer cell activation; while HSC remained unaffected. This related to NF-κB inhibition via up-regulated IκBα. In conclusion, OCA inhibits hepatic inflammation in toxic cirrhotic rats resulting in decreased HSC activation and fibrosis. PMID:27634375

  10. A New Pain Regulatory System via the Brain Long Chain Fatty Acid Receptor GPR40/FFA1 Signal.

    PubMed

    Nakamoto, Kazuo

    2017-01-01

     An increasingly large number of pharmacological and physiological works on fatty acids have shown that the functional properties of fatty acids are regulated by the amount of individual fatty acid intake and the distribution of fatty acids among organs. Recently, it has been determined that G-protein-coupled receptor 40/free fatty acid receptor 1 (GPR40/FFA1) is activated by long-chain fatty acids, such as docosahexaenoic acid (DHA). GPR40/FFA1 is mainly expressed in the β cell of the pancreas, spinal cord and brain. It is reported that this receptor has a functional role in controlling blood glucose levels via the modulation of insulin secretion. However, its physiological function in the brain remains unknown. Our previous studies have shown that GPR40/FFA1 is expressed in pro-opiomelanocortin (POMC)-positive neurons of the arcuate nucleus, serotonergic neurons in the nucleus raphe magnus, and in noradrenergic neurons in the locus coeruleus. Furthermore, the intracerebroventricular injection of DHA or GW9508, which is a selective GPR40/FFA1 agonist, attenuates formalin-induced inflammatory pain behavior through increasing β-endorphin release in the hypothalamus. It also suppresses complete Freund's adjuvant-induced mechanical allodynia and thermal hyperalgesia. Our findings suggest that brain free long-chain fatty acids-GPR40/FFA1 signaling might have an important role in the modulation of endogenous pain control systems. In this review, I discuss the current status and our recent study regarding a new pain regulatory system via the brain long chain fatty acid receptor GPR40/FFA1 signal.

  11. Chlorogenic Acid Improves Late Diabetes through Adiponectin Receptor Signaling Pathways in db/db Mice

    PubMed Central

    Jin, Shasha; Chang, Cuiqing; Zhang, Lantao; Liu, Yang; Huang, Xianren; Chen, Zhimin

    2015-01-01

    The aim of this study was to examine the effects of chlorogenic acid (CGA) on glucose and lipid metabolism in late diabetic db/db mice, as well as on adiponectin receptors and their signaling molecules, to provide evidence for CGA in the prevention of type 2 diabetes. We randomly divided 16 female db/db mice into db/db-CGA and db/db-control (CON) groups equally; db/m mice were used as control mice. The mice in both the db/db-CGA and db/m-CGA groups were administered 80 mg/kg/d CGA by lavage for 12 weeks, whereas the mice in both CON groups were given equal volumes of phosphate-buffered saline (PBS) by lavage. At the end of the intervention, we assessed body fat and the parameters of glucose and lipid metabolism in the plasma, liver and skeletal muscle tissues as well as the levels of aldose reductase (AR) and transforming growth factor-β1 (TGF-β1) in the kidneys and measured adiponectin receptors and the protein expression of their signaling molecules in liver and muscle tissues. After 12 weeks of intervention, compared with the db/db-CON group, the percentage of body fat, fasting plasma glucose (FPG) and glycosylated hemoglobin (HbA1c) in the db/db-CGA group were all significantly decreased; TGF-β1 protein expression and AR activity in the kidney were both decreased; and the adiponectin level in visceral adipose was increased. The protein expression of adiponectin receptors (ADPNRs), the phosphorylation of AMP-activated protein kinase (AMPK) in the liver and muscle, and the mRNA and protein levels of peroxisome proliferator-activated receptor alpha (PPAR-α) in the liver were all significantly greater. CGA could lower the levels of fasting plasma glucose and HbA1c during late diabetes and improve kidney fibrosis to some extent through the modulation of adiponectin receptor signaling pathways in db/db mice. PMID:25849026

  12. L-Amino Acids Elicit Diverse Response Patterns in Taste Sensory Cells: A Role for Multiple Receptors.

    PubMed

    Pal Choudhuri, Shreoshi; Delay, Rona J; Delay, Eugene R

    2015-01-01

    Umami, the fifth basic taste, is elicited by the L-amino acid, glutamate. A unique characteristic of umami taste is the response potentiation by 5' ribonucleotide monophosphates, which are also capable of eliciting an umami taste. Initial reports using human embryonic kidney (HEK) cells suggested that there is one broadly tuned receptor heterodimer, T1r1+T1r3, which detects L-glutamate and all other L-amino acids. However, there is growing evidence that multiple receptors detect glutamate in the oral cavity. While much is understood about glutamate transduction, the mechanisms for detecting the tastes of other L-amino acids are less well understood. We used calcium imaging of isolated taste sensory cells and taste cell clusters from the circumvallate and foliate papillae of C57BL/6J and T1r3 knockout mice to determine if other receptors might also be involved in detection of L-amino acids. Ratiometric imaging with Fura-2 was used to study calcium responses to monopotassium L-glutamate, L-serine, L-arginine, and L-glutamine, with and without inosine 5' monophosphate (IMP). The results of these experiments showed that the response patterns elicited by L-amino acids varied significantly across taste sensory cells. L-amino acids other than glutamate also elicited synergistic responses in a subset of taste sensory cells. Along with its role in synergism, IMP alone elicited a response in a large number of taste sensory cells. Our data indicate that synergistic and non-synergistic responses to L-amino acids and IMP are mediated by multiple receptors or possibly a receptor complex.

  13. L-Amino Acids Elicit Diverse Response Patterns in Taste Sensory Cells: A Role for Multiple Receptors

    PubMed Central

    Pal Choudhuri, Shreoshi; Delay, Rona J.; Delay, Eugene R.

    2015-01-01

    Umami, the fifth basic taste, is elicited by the L-amino acid, glutamate. A unique characteristic of umami taste is the response potentiation by 5’ ribonucleotide monophosphates, which are also capable of eliciting an umami taste. Initial reports using human embryonic kidney (HEK) cells suggested that there is one broadly tuned receptor heterodimer, T1r1+T1r3, which detects L-glutamate and all other L-amino acids. However, there is growing evidence that multiple receptors detect glutamate in the oral cavity. While much is understood about glutamate transduction, the mechanisms for detecting the tastes of other L-amino acids are less well understood. We used calcium imaging of isolated taste sensory cells and taste cell clusters from the circumvallate and foliate papillae of C57BL/6J and T1r3 knockout mice to determine if other receptors might also be involved in detection of L-amino acids. Ratiometric imaging with Fura-2 was used to study calcium responses to monopotassium L-glutamate, L-serine, L-arginine, and L-glutamine, with and without inosine 5’ monophosphate (IMP). The results of these experiments showed that the response patterns elicited by L-amino acids varied significantly across taste sensory cells. L-amino acids other than glutamate also elicited synergistic responses in a subset of taste sensory cells. Along with its role in synergism, IMP alone elicited a response in a large number of taste sensory cells. Our data indicate that synergistic and non-synergistic responses to L-amino acids and IMP are mediated by multiple receptors or possibly a receptor complex. PMID:26110622

  14. Activation function 2 (AF-2) of retinoic acid receptor and 9-cis retinoic acid receptor: presence of a conserved autonomous constitutive activating domain and influence of the nature of the response element on AF-2 activity.

    PubMed Central

    Durand, B; Saunders, M; Gaudon, C; Roy, B; Losson, R; Chambon, P

    1994-01-01

    A motif essential for the transcriptional activation function 2 (AF-2) present in the E region of retinoic acid receptor (RAR) alpha and 9-cis retinoic acid receptor (RXR) alpha has been characterized as an amphipathic alpha-helix whose main features are conserved between transcriptionally active members of the nuclear receptor superfamily. This conserved motif, which can activate autonomously in the absence of ligand in animal and yeast cells, can be swapped between nuclear receptors without affecting the ligand dependency for activation of transcription, thus indicating that a ligand-dependent conformational change is necessary to reveal the AF-2 activation potential within the E region of the nuclear receptor. Interestingly, we show that the precise nature of the direct repeat response element to which RAR/RXR heterodimers are bound can affect the activity of the AF-2s of the heterodimeric partners, as well as the relative efficiency with which all-trans and 9-cis retinoic acids activate the RAR partner. Images PMID:7957103

  15. Individual bile acids have differential effects on bile acid signaling in mice

    SciTech Connect

    Song, Peizhen Rockwell, Cheryl E. Cui, Julia Yue Klaassen, Curtis D.

    2015-02-15

    Bile acids (BAs) are known to regulate BA synthesis and transport by the farnesoid X receptor in the liver (FXR-SHP) and intestine (FXR-Fgf15). However, the relative importance of individual BAs in regulating these processes is not known. Therefore, mice were fed various doses of five individual BAs, including cholic acid (CA), chenodeoxycholic acid (CDCA), deoxoycholic acid (DCA), lithocholic acid (LCA), and ursodeoxycholic acid (UDCA) in their diets at various concentrations for one week to increase the concentration of one BA in the enterohepatic circulation. The mRNA of BA synthesis and transporting genes in liver and ileum were quantified. In the liver, the mRNA of SHP, which is the prototypical target gene of FXR, increased in mice fed all concentrations of BAs. In the ileum, the mRNA of the intestinal FXR target gene Fgf15 was increased at lower doses and to a higher extent by CA and DCA than by CDCA and LCA. Cyp7a1, the rate-limiting enzyme in BA synthesis, was decreased more by CA and DCA than CDCA and LCA. Cyp8b1, the enzyme that 12-hydroxylates BAs and is thus responsible for the synthesis of CA, was decreased much more by CA and DCA than CDCA and LCA. Surprisingly, neither a decrease in the conjugated BA uptake transporter (Ntcp) nor increase in BA efflux transporter (Bsep) was observed by FXR activation, but an increase in the cholesterol efflux transporter (Abcg5/Abcg8) was observed with FXR activation. Thus in conclusion, CA and DCA are more potent FXR activators than CDCA and LCA when fed to mice, and thus they are more effective in decreasing the expression of the rate limiting gene in BA synthesis Cyp7a1 and the 12-hydroxylation of BAs Cyp8b1, and are also more effective in increasing the expression of Abcg5/Abcg8, which is responsible for biliary cholesterol excretion. However, feeding BAs do not alter the mRNA or protein levels of Ntcp or Bsep, suggesting that the uptake or efflux of BAs is not regulated by FXR at physiological and

  16. Activation of peroxisome proliferator-activated receptor-{alpha} enhances fatty acid oxidation in human adipocytes

    SciTech Connect

    Lee, Joo-Young; Hashizaki, Hikari; Goto, Tsuyoshi; Sakamoto, Tomoya; Takahashi, Nobuyuki; Kawada, Teruo

    2011-04-22

    Highlights: {yields} PPAR{alpha} activation increased mRNA expression levels of adipocyte differentiation marker genes and GPDH activity in human adipocytes. {yields} PPAR{alpha} activation also increased insulin-dependent glucose uptake in human adipocytes. {yields} PPAR{alpha} activation did not affect lipid accumulation in human adipocytes. {yields} PPAR{alpha} activation increased fatty acid oxidation through induction of fatty acid oxidation-related genes in human adipocytes. -- Abstract: Peroxisome proliferator-activated receptor-{alpha} (PPAR{alpha}) is a key regulator for maintaining whole-body energy balance. However, the physiological functions of PPAR{alpha} in adipocytes have been unclarified. We examined the functions of PPAR{alpha} using human multipotent adipose tissue-derived stem cells as a human adipocyte model. Activation of PPAR{alpha} by GW7647, a potent PPAR{alpha} agonist, increased the mRNA expression levels of adipocyte differentiation marker genes such as PPAR{gamma}, adipocyte-specific fatty acid-binding protein, and lipoprotein lipase and increased both GPDH activity and insulin-dependent glucose uptake level. The findings indicate that PPAR{alpha} activation stimulates adipocyte differentiation. However, lipid accumulation was not changed, which is usually observed when PPAR{gamma} is activated. On the other hand, PPAR{alpha} activation by GW7647 treatment induced the mRNA expression of fatty acid oxidation-related genes such as CPT-1B and AOX in a PPAR{alpha}-dependent manner. Moreover, PPAR{alpha} activation increased the production of CO{sub 2} and acid soluble metabolites, which are products of fatty acid oxidation, and increased oxygen consumption rate in human adipocytes. The data indicate that activation of PPAR{alpha} stimulates both adipocyte differentiation and fatty acid oxidation in human adipocytes, suggesting that PPAR{alpha} agonists could improve insulin resistance without lipid accumulation in adipocytes. The expected

  17. Chemically engineering ligand selectivity at the free fatty acid receptor 2 based on pharmacological variation between species orthologs

    PubMed Central

    Hudson, Brian D.; Christiansen, Elisabeth; Tikhonova, Irina G.; Grundmann, Manuel; Kostenis, Evi; Adams, David R.; Ulven, Trond; Milligan, Graeme

    2012-01-01

    When it is difficult to develop selective ligands within a family of related G-protein-coupled receptors (GPCRs), chemically engineered receptors activated solely by synthetic ligands (RASSLs) are useful alternatives for probing receptor function. In the present work, we explored whether a RASSL of the free fatty acid receptor 2 (FFA2) could be developed on the basis of pharmacological variation between species orthologs. For this, bovine FFA2 was characterized, revealing distinct ligand selectivity compared with human FFA2. Homology modeling and mutational analysis demonstrated a single mutation in human FFA2 of C4.57G resulted in a human FFA2 receptor with ligand selectivity similar to the bovine receptor. This was exploited to generate human FFA2-RASSL by the addition of a second mutation at a known orthosteric ligand interaction site, H6.55Q. The resulting FFA2-RASSL displayed a >100-fold loss of activity to endogenous ligands, while responding to the distinct ligand sorbic acid with pEC50 values for inhibition of cAMP, 5.83 ± 0.11; Ca2+ mobilization, 4.63 ± 0.05; ERK phosphorylation, 5.61 ± 0.06; and dynamic mass redistribution, 5.35 ± 0.06. This FFA2-RASSL will be useful in future studies on this receptor and demonstrates that exploitation of pharmacological variation between species orthologs is a powerful method to generate novel chemically engineered GPCRs.—Hudson, B. D., Christiansen, E., Tikhonova, I. G., Grundmann, M., Kostenis, E., Adams, D. R., Ulven, T., Milligan, G. Chemically engineering ligand selectivity at the free fatty acid receptor 2 based on pharmacological variation between species orthologs. PMID:22919070

  18. Gamma-Aminobutyric acid and benzodiazepine receptors in the kindling model of epilepsy: a quantitative radiohistochemical study

    SciTech Connect

    Shin, C.; Pedersen, H.B.; McNamara, J.O.

    1985-10-01

    Quantitative radiohistochemistry was utilized to study alterations of gamma-aminobutyric acid (GABA) and benzodiazepine receptors in the kindling model of epilepsy. The radioligands used for GABA and benzodiazepine receptors were (TH) muscimol and (TH)flunitrazepam, respectively. GABA receptor binding was increased by 22% in fascia dentata of the hippocampal formation but not in neocortex or substantia nigra of kindled rats. Within fascia dentata, GABA receptor binding was increased to an equivalent extent in stratum granulosum and throughout stratum moleculare; no increase was found in dentate hilus or stratum lacunosummoleculare or stratum radiatum of CA1. The increased binding was present at 24 hr but not at 28 days after the last kindled seizure. The direction, anatomic distribution, and time course of the increased GABA receptor binding were paralleled by increased benzodiazepine receptor binding. The anatomic distribution of the increased GABA receptor binding is consistent with a localization to somata and dendritic trees of dentate granule cells. The authors suggest that increased GABA and benzodiazepine receptor binding may contribute to enhanced inhibition of dentate granule cells demonstrated electrophysiologically in kindled animals.

  19. Amino acid residues 4425-4621 localized on the three-dimensional structure of the skeletal muscle ryanodine receptor.

    PubMed Central

    Benacquista, B L; Sharma, M R; Samsó, M; Zorzato, F; Treves, S; Wagenknecht, T

    2000-01-01

    We have localized a region contained within the sequence of amino acid residues 4425-4621 on the three-dimensional structure of the skeletal muscle ryanodine receptor (RyR). Mouse monoclonal antibodies raised against a peptide comprising these residues have been complexed with ryanodine receptors and imaged in the frozen-hydrated state by cryoelectron microscopy. These images, along with images of antibody-free ryanodine receptor, were used to compute two-dimensional averaged images and three-dimensional reconstructions. Two-dimensional averages of immunocomplexes in which the ryanodine receptor was in the fourfold symmetrical orientation disclosed four symmetrical regions of density located on the edges of the receptor's cytoplasmic assembly that were absent from control averages of receptor without added antibody. Three-dimensional reconstructions revealed the antibody-binding sites to be on the so-called handle domains of the ryanodine receptor's cytoplasmic assembly, near their junction with the transmembrane assembly. This study is the first to demonstrate epitope mapping on the three-dimensional structure of the ryanodine receptor. PMID:10692321

  20. Salvianolic Acid A, as a Novel ETA Receptor Antagonist, Shows Inhibitory Effects on Tumor in Vitro

    PubMed Central

    Zhang, Qiao; Wang, Shifeng; Yu, Yangyang; Sun, Shengnan; Zhang, Yuxin; Zhang, Yanling; Yang, Wei; Li, Shiyou; Qiao, Yanjiang

    2016-01-01

    Endothelin-1 (ET-1) autocrine and paracrine signaling modulate cell proliferation of tumor cells by activating its receptors, endothelin A receptor (ETAR) and endothelin B receptor (ETBR). Dysregulation of ETAR activation promotes tumor development and progression. The potential of ETAR antagonists and the dual-ETAR and ETBR antagonists as therapeutic approaches are under preclinical and clinical studies. Salvianolic acid A (Sal A) is a hydrophilic polyphenolic derivative isolated from Salvia miltiorrhiza Bunge (Danshen), which has been reported as an anti-cancer and cardio-protective herbal medicine. In this study, we demonstrate that Sal A inhibits ETAR activation induced by ET-1 in both recombinant and endogenous ETAR expression cell lines. The IC50 values were determined as 5.7 µM in the HEK293/ETAR cell line and 3.14 µM in HeLa cells, respectively. Furthermore, our results showed that Sal A suppressed cell proliferation and extended the doubling times of multiple cancer cells, including HeLa, DU145, H1975, and A549 cell lines. In addition, Sal A inhibited proliferation of DU145 cell lines stimulated by exogenous ET-1 treatment. Moreover, the cytotoxicity and cardio-toxicity of Sal A were assessed in human umbilical vein endothelial cells (HUVEC) and Human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs), which proved that Sal A demonstrates no cytotoxicity or cardiotoxicity. Collectively, our findings indicate that Sal A is a novel anti-cancer candidate through targeting ETAR. PMID:27490540

  1. Loss of lysophosphatidic acid receptor-3 enhances cell migration in rat lung tumor cells

    SciTech Connect

    Hayashi, Mai; Okabe, Kyoko; Yamawaki, Yasuna; Teranishi, Miki; Honoki, Kanya; Mori, Toshio; Fukushima, Nobuyuki; Tsujiuchi, Toshifumi

    2011-02-18

    Research highlights: {yields} Loss of the Lpar3 expression due to aberrant DNA methylation occurred in rat lung tumor cells. {yields} The Lpar3 inhibited cell migration of rat lung tumor cells. {yields} The Lpar3 may act as a negative regulator of rat lung tumor cells. -- Abstract: Lysophosphatidic acid (LPA) indicates several biological effects, such as cell proliferation, differentiation and migration. LPA interacts with G protein-coupled transmembrane LPA receptors. In our previous report, we detected that loss of the LPA receptor-1 (Lpar1) expression is due to its aberrant DNA methylation in rat tumor cell lines. In this study, to assess an involvement of the other LPA receptor, Lpar3, in the pathogenesis of rat lung tumor cells, we measured the expression levels of the Lpar3 gene and its DNA methylation status by reverse transcription (RT)-polymerase chain reaction (PCR) and bisulfite sequencing analyses, respectively. RLCNR lung adenocarcinoma cells showed reduced expression of the Lpar3, compared with normal lung tissues. In the 5' upstream region of the Lpar3, normal lung tissues were unmethylated. By contrast, RLCNR cells were highly methylated, correlating with reduced expressions of the Lpar3. Based on these results, we generated the Lpar3-expressing RLCNR-a3 cells and measured the cell migration ability. Interestingly, the cell migration of RLCNR-a3 cells was significantly lower than that of RLCNR cells. This study suggests that loss of the Lpar3 due to aberrant DNA methylation may be involved in the progression of rat lung tumor cells.

  2. Identification of a chemoreceptor for tricarboxylic acid cycle intermediates: differential chemotactic response towards receptor ligands.

    PubMed

    Lacal, Jesús; Alfonso, Carlos; Liu, Xianxian; Parales, Rebecca E; Morel, Bertrand; Conejero-Lara, Francisco; Rivas, Germán; Duque, Estrella; Ramos, Juan L; Krell, Tino

    2010-07-23

    We report the identification of McpS as the specific chemoreceptor for 6 tricarboxylic acid (TCA) cycle intermediates and butyrate in Pseudomonas putida. The analysis of the bacterial mutant deficient in mcpS and complementation assays demonstrate that McpS is the only chemoreceptor of TCA cycle intermediates in the strain under study. TCA cycle intermediates are abundantly present in root exudates, and taxis toward these compounds is proposed to facilitate the access to carbon sources. McpS has an unusually large ligand-binding domain (LBD) that is un-annotated in InterPro and is predicted to contain 6 helices. The ligand profile of McpS was determined by isothermal titration calorimetry of purified recombinant LBD (McpS-LBD). McpS recognizes TCA cycle intermediates but does not bind very close structural homologues and derivatives like maleate, aspartate, or tricarballylate. This implies that functional similarity of ligands, such as being part of the same pathway, and not structural similarity is the primary element, which has driven the evolution of receptor specificity. The magnitude of chemotactic responses toward these 7 chemoattractants, as determined by qualitative and quantitative chemotaxis assays, differed largely. Ligands that cause a strong chemotactic response (malate, succinate, and fumarate) were found by differential scanning calorimetry to increase significantly the midpoint of protein unfolding (T(m)) and unfolding enthalpy (DeltaH) of McpS-LBD. Equilibrium sedimentation studies show that malate, the chemoattractant that causes the strongest chemotactic response, stabilizes the dimeric state of McpS-LBD. In this respect clear parallels exist to the Tar receptor and other eukaryotic receptors, which are discussed.

  3. Lysophosphatidic acid receptors (LPARs): Potential targets for the treatment of neuropathic pain.

    PubMed

    Velasco, María; O'Sullivan, Catherine; Sheridan, Graham K

    2017-02-01

    Neuropathic pain can arise from lesions to peripheral or central nerve fibres leading to spontaneous action potential generation and a lowering of the nociceptive threshold. Clinically, neuropathic pain can manifest in many chronic disease states such as cancer, diabetes or multiple sclerosis (MS). The bioactive lipid, lysophosphatidic acid (LPA), via activation of its receptors (LPARs), is thought to play a central role in both triggering and maintaining neuropathic pain. In particular, following an acute nerve injury, the excitatory neurotransmitters glutamate and substance P are released from primary afferent neurons leading to upregulated synthesis of lysophosphatidylcholine (LPC), the precursor for LPA production. LPC is converted to LPA by autotaxin (ATX), which can then activate macrophages/microglia and modulate neuronal functioning. A ubiquitous feature of animal models of neuropathic pain is demyelination of damaged nerves. It is thought that LPA contributes to demyelination through several different mechanisms. Firstly, high levels of LPA are produced following macrophage/microglial activation that triggers a self-sustaining feed-forward loop of de novo LPA synthesis. Secondly, macrophage/microglial activation contributes to inflammation-mediated demyelination of axons, thus initiating neuropathic pain. Therefore, targeting LPA production and/or the family of LPA-activated G protein-coupled receptors (GPCRs) may prove to be fruitful clinical approaches to treating demyelination and the accompanying neuropathic pain. This review discusses our current understanding of the role of LPA/LPAR signalling in the initiation of neuropathic pain and suggests potential targeted strategies for its treatment. This article is part of the Special Issue entitled 'Lipid Sensing G Protein-Coupled Receptors in the CNS'.

  4. Evolutionary diversification of retinoic acid receptor ligand-binding pocket structure by molecular tinkering

    PubMed Central

    Gutierrez-Mazariegos, Juliana; Nadendla, Eswar Kumar; Studer, Romain A.; Alvarez, Susana; de Lera, Angel R.; Kuraku, Shigehiro; Bourguet, William; Laudet, Vincent

    2016-01-01

    Whole genome duplications (WGDs) have been classically associated with the origin of evolutionary novelties and the so-called duplication–degeneration–complementation model describes the possible fates of genes after duplication. However, how sequence divergence effectively allows functional changes between gene duplicates is still unclear. In the vertebrate lineage, two rounds of WGDs took place, giving rise to paralogous gene copies observed for many gene families. For the retinoic acid receptors (RARs), for example, which are members of the nuclear hormone receptor (NR) superfamily, a unique ancestral gene has been duplicated resulting in three vertebrate paralogues: RARα, RARβ and RARγ. It has previously been shown that this single ancestral RAR was neofunctionalized to give rise to a larger substrate specificity range in the RARs of extant jawed vertebrates (also called gnathostomes). To understand RAR diversification, the members of the cyclostomes (lamprey and hagfish), jawless vertebrates representing the extant sister group of gnathostomes, provide an intermediate situation and thus allow the characterization of the evolutionary steps that shaped RAR ligand-binding properties following the WGDs. In this study, we assessed the ligand-binding specificity of cyclostome RARs and found that their ligand-binding pockets resemble those of gnathostome RARα and RARβ. In contrast, none of the cyclostome receptors studied showed any RARγ-like specificity. Together, our results suggest that cyclostome RARs cover only a portion of the specificity repertoire of the ancestral gnathostome RARs and indicate that the establishment of ligand-binding specificity was a stepwise event. This iterative process thus provides a rare example for the diversification of receptor–ligand interactions of NRs following WGDs. PMID:27069642

  5. A naturally occurring insertion of a single amino acid rewires transcriptional regulation by glucocorticoid receptor isoforms.

    PubMed

    Thomas-Chollier, Morgane; Watson, Lisa C; Cooper, Samantha B; Pufall, Miles A; Liu, Jennifer S; Borzym, Katja; Vingron, Martin; Yamamoto, Keith R; Meijsing, Sebastiaan H

    2013-10-29

    In addition to guiding proteins to defined genomic loci, DNA can act as an allosteric ligand that influences protein structure and activity. Here we compared genome-wide binding, transcriptional regulation, and, using NMR, the conformation of two glucocorticoid receptor (GR) isoforms that differ by a single amino acid insertion in the lever arm, a domain that adopts DNA sequence-specific conformations. We show that these isoforms differentially regulate gene expression levels through two mechanisms: differential DNA binding and altered communication between GR domains. Our studies suggest a versatile role for DNA in both modulating GR activity and also in directing the use of GR isoforms. We propose that the lever arm is a "fulcrum" for bidirectional allosteric signaling, conferring conformational changes in the DNA reading head that influence DNA sequence selectivity, as well as conferring changes in the dimerization domain that connect functionally with remote regulatory surfaces, thereby influencing which genes are regulated and the magnitude of their regulation.

  6. The N-terminal portion of domain E of retinoic acid receptors alpha and beta is essential for the recognition of retinoic acid and various analogs.

    PubMed

    Ostrowski, J; Hammer, L; Roalsvig, T; Pokornowski, K; Reczek, P R

    1995-03-14

    Utilizing a strategy involving domain exchange between retinoic acid receptors alpha and beta (RAR alpha and RAR beta) and monitoring the transcriptional activity of the resulting chimeric receptors with receptor-selective retinoids, we identified a 70-aa region within the N-terminal portion of the RAR alpha and -beta domain E which is important for an RAR alpha- or RAR beta-specific response. Two amino acid residues within this region, serine-232 (S232) and threonine-239 (T239) in RAR alpha and the corresponding alanine-225 (A225) and isoleucine-232 (I232) in RAR beta, were found to be essential for this effect. In addition, binding studies using the chimeric receptors expressed in Escherichia coli showed that the N-terminal portion of domain E was also important for the characteristic binding profile of t-RA and various retinoids with RAR alpha or RAR beta. Structural predictions of the primary amino acid sequence in this region indicate the presence of an amphipathic helix-turn-helix structure with five hydrophobic amino acids that resemble a leucine zipper motif. The amino acid residues identified by domain swapping, S232 and T239 in RAR alpha and A225 and I232 in RAR beta, were found within the hydrophobic face of an alpha-helix in close proximity to this zipper motif, suggesting that the ligand may interact with the receptor in the region adjacent to a surface involved in protein-protein interactions. This finding may link ligand binding to other processes important for transcriptional activation.

  7. The N-terminal portion of domain E of retinoic acid receptors alpha and beta is essential for the recognition of retinoic acid and various analogs.

    PubMed Central

    Ostrowski, J; Hammer, L; Roalsvig, T; Pokornowski, K; Reczek, P R

    1995-01-01

    Utilizing a strategy involving domain exchange between retinoic acid receptors alpha and beta (RAR alpha and RAR beta) and monitoring the transcriptional activity of the resulting chimeric receptors with receptor-selective retinoids, we identified a 70-aa region within the N-terminal portion of the RAR alpha and -beta domain E which is important for an RAR alpha- or RAR beta-specific response. Two amino acid residues within this region, serine-232 (S232) and threonine-239 (T239) in RAR alpha and the corresponding alanine-225 (A225) and isoleucine-232 (I232) in RAR beta, were found to be essential for this effect. In addition, binding studies using the chimeric receptors expressed in Escherichia coli showed that the N-terminal portion of domain E was also important for the characteristic binding profile of t-RA and various retinoids with RAR alpha or RAR beta. Structural predictions of the primary amino acid sequence in this region indicate the presence of an amphipathic helix-turn-helix structure with five hydrophobic amino acids that resemble a leucine zipper motif. The amino acid residues identified by domain swapping, S232 and T239 in RAR alpha and A225 and I232 in RAR beta, were found within the hydrophobic face of an alpha-helix in close proximity to this zipper motif, suggesting that the ligand may interact with the receptor in the region adjacent to a surface involved in protein-protein interactions. This finding may link ligand binding to other processes important for transcriptional activation. PMID:7892182

  8. Tributyltin and triphenyltin inhibit osteoclast differentiation through a retinoic acid receptor-dependent signaling pathway

    SciTech Connect

    Yonezawa, Takayuki; Hasegawa, Shin-ichi; Ahn, Jae-Yong; Cha, Byung-Yoon; Teruya, Toshiaki; Hagiwara, Hiromi; Nagai, Kazuo; Woo, Je-Tae; E-mail: jwoo@isc.chubu.ac.jp

    2007-03-30

    Organotin compounds, such as tributyltin (TBT) and triphenyltin (TPT), have been widely used in agriculture and industry. Although these compounds are known to have many toxic effects, including endocrine-disrupting effects, their effects on bone resorption are unknown. In this study, we investigated the effects of organotin compounds, such as monobutyltin (MBT), dibutyltin (DBT), TBT, and TPT, on osteoclast differentiation using mouse monocytic RAW264.7 cells. MBT and DBT had no effects, whereas TBT and TPT dose-dependently inhibited osteoclast differentiation at concentrations of 3-30 nM. Treatment with a retinoic acid receptor (RAR)-specific antagonist, Ro41-5253, restored the inhibition of osteoclastogenesis by TBT and TPT. TBT and TPT reduced receptor activator of nuclear factor-{kappa}B ligand (RANKL) induced nuclear factor of activated T cells (NFAT) c1 expression, and the reduction in NFATc1 expression was recovered by Ro41-5253. Our results suggest that TBT and TPT suppress osteoclastogenesis by inhibiting RANKL-induced NFATc1 expression via an RAR-dependent signaling pathway.

  9. Synaptic enhancement induced by gintonin via lysophosphatidic acid receptor activation in central synapses.

    PubMed

    Park, Hoyong; Kim, Sungmin; Rhee, Jeehae; Kim, Hyeon-Joong; Han, Jung-Soo; Nah, Seung-Yeol; Chung, ChiHye

    2015-03-01

    Lysophosphatidic acid (LPA) is one of the well-characterized, ubiquitous phospholipid molecules. LPA exerts its effect by activating G protein-coupled receptors known as LPA receptors (LPARs). So far, LPAR signaling has been critically implicated during early development stages, including the regulation of synapse formation and the morphology of cortical and hippocampal neurons. In adult brains, LPARs seem to participate in cognitive as well as emotional learning and memory. Recent studies using LPAR1-deficient mice reported impaired performances in a number of behavioral tasks, including the hippocampus-dependent spatial memory and fear conditioning tests. Nevertheless, the effect of LPAR activation in the synaptic transmission of central synapses after the completion of embryonic development has not been investigated. In this study, we took advantage of a novel extracellular agonist for LPARs called gintonin to activate LPARs in adult brain systems. Gintonin, a recently identified active ingredient in ginseng, has been shown to activate LPARs and mobilize Ca(2+) in an artificial cell system. We found that the activation of LPARs by application of gintonin acutely enhanced both excitatory and inhibitory transmission in central synapses, albeit through tentatively distinct mechanisms. Gintonin-mediated LPAR activation primarily resulted in synaptic enhancement and an increase in neuronal excitability in a phospholipase C-dependent manner. Our findings suggest that LPARs are able to directly potentiate synaptic transmission in central synapses when stimulated exogenously. Therefore, LPARs could serve as a useful target to modulate synaptic activity under pathological conditions, including neurodegenerative diseases.

  10. Designed abscisic acid analogs as antagonists of PYL-PP2C receptor interactions.

    PubMed

    Takeuchi, Jun; Okamoto, Masanori; Akiyama, Tomonori; Muto, Takuya; Yajima, Shunsuke; Sue, Masayuki; Seo, Mitsunori; Kanno, Yuri; Kamo, Tsunashi; Endo, Akira; Nambara, Eiji; Hirai, Nobuhiro; Ohnishi, Toshiyuki; Cutler, Sean R; Todoroki, Yasushi

    2014-06-01

    The plant stress hormone abscisic acid (ABA) is critical for several abiotic stress responses. ABA signaling is normally repressed by group-A protein phosphatases 2C (PP2Cs), but stress-induced ABA binds Arabidopsis PYR/PYL/RCAR (PYL) receptors, which then bind and inhibit PP2Cs. X-ray structures of several receptor-ABA complexes revealed a tunnel above ABA's 3' ring CH that opens at the PP2C binding interface. Here, ABA analogs with sufficiently long 3' alkyl chains were predicted to traverse this tunnel and block PYL-PP2C interactions. To test this, a series of 3'-alkylsulfanyl ABAs were synthesized with different alkyl chain lengths. Physiological, biochemical and structural analyses revealed that a six-carbon alkyl substitution produced a potent ABA antagonist that was sufficiently active to block multiple stress-induced ABA responses in vivo. This study provides a new approach for the design of ABA analogs, and the results validated structure-based design for this target class.

  11. Phosphatidic Acid Induces Ligand-independent Epidermal Growth Factor Receptor Endocytic Traffic through PDE4 Activation

    PubMed Central

    Norambuena, Andrés; Metz, Claudia; Jung, Juan E.; Silva, Antonia; Otero, Carolina; Cancino, Jorge; Retamal, Claudio; Valenzuela, Juan C.; Soza, Andrea

    2010-01-01

    Endocytosis modulates EGFR function by compartmentalizing and attenuating or enhancing its ligand-induced signaling. Here we show that it can also control the cell surface versus intracellular distribution of empty/inactive EGFR. Our previous observation that PKA inhibitors induce EGFR internalization prompted us to test phosphatidic acid (PA) generated by phospholipase D (PLD) as an endogenous down-regulator of PKA activity, which activates rolipram-sensitive type 4 phosphodiesterases (PDE4) that degrade cAMP. We found that inhibition of PA hydrolysis by propranolol, in the absence of ligand, provokes internalization of inactive (neither tyrosine-phosphorylated nor ubiquitinated) EGFR, accompanied by a transient increase in PA levels and PDE4s activity. This EGFR internalization is mimicked by PA micelles and is strongly counteracted by PLD2 silencing, rolipram or forskolin treatment, and PKA overexpression. Accelerated EGFR endocytosis seems to be mediated by clathrin-dependent and -independent pathways, leading to receptor accumulation in juxtanuclear recycling endosomes, also due to a decreased recycling. Internalized EGFR can remain intracellular without degradation for several hours or return rapidly to the cell surface upon discontinuation of the stimulus. This novel regulatory mechanism of EGFR, also novel function of signaling PA, can transmodulate receptor accessibility in response to heterologous stimuli. PMID:20554760

  12. Acid-sensing ion channels and transient-receptor potential ion channels in zebrafish taste buds.

    PubMed

    Levanti, M; Randazzo, B; Viña, E; Montalbano, G; Garcia-Suarez, O; Germanà, A; Vega, J A; Abbate, F

    2016-09-01

    Sensory information from the environment is required for life and survival, and it is detected by specialized cells which together make up the sensory system. The fish sensory system includes specialized organs that are able to detect mechanical and chemical stimuli. In particular, taste buds are small organs located on the tongue in terrestrial vertebrates that function in the perception of taste. In fish, taste buds occur on the lips, the flanks, and the caudal (tail) fins of some species and on the barbels of others. In fish taste receptor cells, different classes of ion channels have been detected which, like in mammals, presumably participate in the detection and/or transduction of chemical gustatory signals. However, since some of these ion channels are involved in the detection of additional sensory modalities, it can be hypothesized that taste cells sense stimuli other than those specific for taste. This mini-review summarizes current knowledge on the presence of transient-receptor potential (TRP) and acid-sensing (ASIC) ion channels in the taste buds of teleosts, especially adult zebrafish. Up to now ASIC4, TRPC2, TRPA1, TRPV1 and TRPV4 ion channels have been found in the sensory cells, while ASIC2 was detected in the nerves supplying the taste buds.

  13. Binding of thyroglobulin to bovine thyroid membranes. Role of specific amino acids in receptor recognition.

    PubMed

    Shifrin, S; Kohn, L D

    1981-10-25

    Bovine thyroglobulin was treated with increasing ratios of succinic anhydride, trinitrobenzene sulfonic acid, tetranitromethane, and N-acetylimidazole in an attempt to assess the role of lysine or tyrosine residues in binding to thyroid membrane receptors. Extensive succinylation results in dissociation to 12 S thyroglobulin with retention of a considerable portion of the three-dimensional structure. Only 25% of the lysine residues can be modified by trinitrophenylation without affecting inter-subunit interactions. Succinylation as well as trinitrophenylation increases the affinity of thyroglobulin for the membrane receptor by a factor of 2. The binding of thyroglobulin to the membrane was reduced after nitration of 30% of the tyrosyl residues with tetranitromethane. O-Acetylation of 40-70% of the tyrosyl residues by N-acetylimidazole nearly abolished the ability of thyroglobulin to bind to the membrane. Removal of the O-acetyl group with hydroxylamine restored the binding properties. The results indicate that tyrosyl residues play an important role in thyroglobulin interactions with thyroid membranes.

  14. Contamination with retinoic acid receptor agonists in two rivers in the Kinki region of Japan.

    PubMed

    Inoue, Daisuke; Nakama, Koki; Sawada, Kazuko; Watanabe, Taro; Takagi, Mai; Sei, Kazunari; Yang, Min; Hirotsuji, Junji; Hu, Jianying; Nishikawa, Jun-ichi; Nakanishi, Tsuyoshi; Ike, Michihiko

    2010-04-01

    This study was conducted to investigate the agonistic activity against human retinoic acid receptor (RAR) alpha in the Lake Biwa-Yodo River and the Ina River in the Kinki region of Japan. To accomplish this, a yeast two-hybrid assay was used to elucidate the spatial and temporal variations and potential sources of RARalpha agonist contamination in the river basins. RARalpha agonistic activity was commonly detected in the surface water samples collected along two rivers at different periods, with maximum all-trans retinoic acid (atRA) equivalents of 47.6 ng-atRA/L and 23.5 ng-atRA/L being observed in Lake Biwa-Yodo River and Ina River, respectively. The results indicated that RARalpha agonists are always present and widespread in the rivers. Comparative investigation of RARalpha and estrogen receptor alpha agonistic activities at 20 stations along each river revealed that the spatial variation pattern of RARalpha agonist contamination was entirely different from that of the estrogenic compound contamination. This suggests that the effluent from municipal wastewater treatment plants, a primary source of estrogenic compounds, seemed not to be the cause of RARalpha agonist contamination in the rivers. Fractionation using high performance liquid chromatography (HPLC) directed by the bioassay found two bioactive fractions from river water samples, suggesting the presence of at least two RARalpha agonists in the rivers. Although a trial conducted to identify RARalpha agonists in the major bioactive fraction was not completed as part of this study, comparison of retention times in HPLC analysis and quantification with liquid chromatography-mass spectrometry analysis revealed that the major causative contaminants responsible for the RARalpha agonistic activity were not RAs (natural RAR ligands) and 4-oxo-RAs, while 4-oxo-RAs were identified as the major RAR agonists in sewage in Beijing, China. These findings suggest that there are unknown RARalpha agonists with high

  15. Antagonizing Retinoic Acid Receptors Increases Myeloid Cell Production by Cultured Human Hematopoietic Stem Cells.

    PubMed

    Brown, Geoffrey; Marchwicka, Aleksandra; Cunningham, Alan; Toellner, Kai-Michael; Marcinkowska, Ewa

    2017-02-01

    Activities of the retinoic acid receptor (RAR)α and RARγ are important to hematopoiesis. Here, we have investigated the effects of receptor selective agonists and antagonists on the primitive human hematopoietic cell lines KG1 and NB-4 and purified normal human hematopoietic stem cells (HSCs). Agonizing RARα (by AGN195183) was effective in driving neutrophil differentiation of NB-4 cells and this agonist synergized with a low amount (10 nM) of 1α,25-dihydroxyvitamin D3 to drive monocyte differentiation of NB-4 and KG1 cells. Treatment of cultures of human HSCs (supplemented with stem cell factor ± interleukin 3) with an antagonist of all RARs (AGN194310) or of RARα (AGN196996) prolonged the lifespan of cultures, up to 55 days, and increased the production of neutrophils and monocytes. Slowing down of cell differentiation was not observed, and instead, hematopoietic stem and progenitor cells had expanded in number. Antagonism of RARγ (by AGN205728) did not affect cultures of HSCs. Studies of CV-1 and LNCaP cells transfected with RAR expression vectors and a reporter vector revealed that RARγ and RARβ are activated by sub-nM all-trans retinoic acid (EC50-0.3 nM): ~50-fold more is required for activation of RARα (EC50-16 nM). These findings further support the notion that the balance of expression and activity of RARα and RARγ are important to hematopoietic stem and progenitor cell expansion and differentiation.

  16. Lysophosphatidic acid induces vasodilation mediated by LPA1 receptors, phospholipase C, and endothelial nitric oxide synthase

    PubMed Central

    Ruisanchez, Éva; Dancs, Péter; Kerék, Margit; Németh, Tamás; Faragó, Bernadett; Balogh, Andrea; Patil, Renukadevi; Jennings, Brett L.; Liliom, Károly; Malik, Kafait U.; Smrcka, Alan V.; Tigyi, Gabor; Benyó, Zoltán

    2014-01-01

    Lysophosphatidic acid (LPA) has been implicated as a mediator of several cardiovascular functions, but its potential involvement in the control of vascular tone is obscure. Here, we show that both LPA (18:1) and VPC31143 (a synthetic agonist of LPA1–3 receptors) relax intact mouse thoracic aorta with similar Emax values (53.9 and 51.9% of phenylephrine-induced precontraction), although the EC50 of LPA- and VPC31143-induced vasorelaxations were different (400 vs. 15 nM, respectively). Mechanical removal of the endothelium or genetic deletion of endothelial nitric oxide synthase (eNOS) not only diminished vasorelaxation by LPA or VPC31143 but converted it to vasoconstriction. Freshly isolated mouse aortic endothelial cells expressed LPA1, LPA2, LPA4 and LPA5 transcripts. The LPA1,3 antagonist Ki16425, the LPA1 antagonist AM095, and the genetic deletion of LPA1, but not that of LPA2, abolished LPA-induced vasorelaxation. Inhibition of the phosphoinositide 3 kinase–protein kinase B/Akt pathway by wortmannin or MK-2206 failed to influence the effect of LPA. However, pharmacological inhibition of phospholipase C (PLC) by U73122 or edelfosine, but not genetic deletion of PLCε, abolished LPA-induced vasorelaxation and indicated that a PLC enzyme, other than PLCε, mediates the response. In summary, the present study identifies LPA as an endothelium-dependent vasodilator substance acting via LPA1, PLC, and eNOS.—Ruisanchez, É., Dancs, P., Kerék, M., Németh, T., Faragó, B., Balogh, A., Patil, R., Jennings, B. L., Liliom, K., Malik, K. U., Smrcka, A. V., Tigyi, G., Benyó, Z. Lysophosphatidic acid induces vasodilation mediated by LPA1 receptors, phospholipase C, and endothelial nitric oxide synthase. PMID:24249637

  17. Hydroxyindole Carboxylic Acid-Based Inhibitors for Receptor-Type Protein Tyrosine Protein Phosphatase Beta

    PubMed Central

    Zeng, Li-Fan; Zhang, Ruo-Yu; Bai, Yunpeng; Wu, Li; Gunawan, Andrea M.

    2014-01-01

    Abstract Aims: Protein tyrosine phosphatases (PTPs) play an important role in regulating a wide range of cellular processes. Understanding the role of PTPs within these processes has been hampered by a lack of potent and selective PTP inhibitors. Generating potent and selective probes for PTPs remains a significant challenge because of the highly conserved and positively charged PTP active site that also harbors a redox-sensitive Cys residue. Results: We describe a facile method that uses an appropriate hydroxyindole carboxylic acid to anchor the inhibitor to the PTP active site and relies on the secondary binding elements introduced through an amide-focused library to enhance binding affinity for the target PTP and to impart selectivity against off-target phosphatases. Here, we disclose a novel series of hydroxyindole carboxylic acid-based inhibitors for receptor-type tyrosine protein phosphatase beta (RPTPβ), a potential target that is implicated in blood vessel development. The representative RPTPβ inhibitor 8b-1 (L87B44) has an IC50 of 0.38 μM and at least 14-fold selectivity for RPTPβ over a large panel of PTPs. Moreover, 8b-1 also exhibits excellent cellular activity and augments growth factor signaling in HEK293, MDA-MB-468, and human umbilical vein endothelial cells. Innovation: The bicyclic salicylic acid pharmacophore-based focused library approach may provide a potential solution to overcome the bioavailability issue that has plagued the PTP drug discovery field for many years. Conclusion: A novel method is described for the development of bioavailable PTP inhibitors that utilizes bicyclic salicylic acid to anchor the inhibitors to the active site and peripheral site interactions to enhance binding affinity and selectivity. Antioxid. Redox Signal. 20, 2130–2140. PMID:24180557

  18. Cerebrospinal Fluid Steroidomics: Are Bioactive Bile Acids Present in Brain?*

    PubMed Central

    Ogundare, Michael; Theofilopoulos, Spyridon; Lockhart, Andrew; Hall, Leslie J.; Arenas, Ernest; Sjövall, Jan; Brenton, A. Gareth; Wang, Yuqin; Griffiths, William J.

    2010-01-01

    In this study we have profiled the free sterol content of cerebrospinal fluid by a combination of charge tagging and liquid chromatography-tandem mass spectrometry. Surprisingly, the most abundant cholesterol metabolites were found to be C27 and C24 intermediates of the bile acid biosynthetic pathways with structures corresponding to 7α-hydroxy-3-oxocholest-4-en-26-oic acid (7.170 ± 2.826 ng/ml, mean ± S.D., six subjects), 3β-hydroxycholest-5-en-26-oic acid (0.416 ± 0.193 ng/ml), 7α,x-dihydroxy-3-oxocholest-4-en-26-oic acid (1.330 ± 0.543 ng/ml), and 7α-hydroxy-3-oxochol-4-en-24-oic acid (0.172 ± 0.085 ng/ml), and the C26 sterol 7α-hydroxy-26-norcholest-4-ene-3,x-dione (0.204 ± 0.083 ng/ml), where x is an oxygen atom either on the CD rings or more likely on the C-17 side chain. The ability of intermediates of the bile acid biosynthetic pathways to activate the liver X receptors (LXRs) and the farnesoid X receptor was also evaluated. The acidic cholesterol metabolites 3β-hydroxycholest-5-en-26-oic acid and 3β,7α-dihydroxycholest-5-en-26-oic acid were found to activate LXR in a luciferase assay, but the major metabolite identified in this study, i.e. 7α-hydroxy-3-oxocholest-4-en-26-oic acid, was not an LXR ligand. 7α-Hydroxy-3-oxocholest-4-en-26-oic acid is formed from 3β,7α-dihydroxycholest-5-en-26-oic acid in a reaction catalyzed by 3β-hydroxy-Δ5-C27-steroid dehydrogenase (HSD3B7), which may thus represent a deactivation pathway of LXR ligands in brain. Significantly, LXR activation has been found to reduce the symptoms of Alzheimer disease (Fan, J., Donkin, J., and Wellington C. (2009) Biofactors 35, 239–248); thus, cholesterol metabolites may play an important role in the etiology of Alzheimer disease. PMID:19996111

  19. Identification of the putative binding pocket of valerenic acid on GABAA receptors using docking studies and site‐directed mutagenesis

    PubMed Central

    Luger, D; Poli, G; Wieder, M; Stadler, M; Ke, S; Ernst, M; Hohaus, A; Linder, T; Seidel, T; Langer, T; Hering, S

    2015-01-01

    Background and Purpose β2/3‐subunit‐selective modulation of GABAA receptors by valerenic acid (VA) is determined by the presence of transmembrane residue β2/3N265. Currently, it is not known whether β2/3N265 is part of VA's binding pocket or is involved in the transduction pathway of VA's action. The aim of this study was to clarify the localization of VA's binding pocket on GABAA receptors. Experimental Approach Docking and a structure‐based three‐dimensional pharmacophore were employed to identify candidate amino acid residues that are likely to interact with VA. Selected amino acid residues were mutated, and VA‐induced modulation of the resulting GABAA receptors expressed in Xenopus oocytes was analysed. Key Results A binding pocket for VA at the β+/α− interface encompassing amino acid β3N265 was predicted. Mutational analysis of suggested amino acid residues revealed a complete loss of VA's activity on β3M286W channels as well as significantly decreased efficacy and potency of VA on β3N265S and β3F289S receptors. In addition, reduced efficacy of VA‐induced I GABA enhancement was also observed for α1M235W, β3R269A and β3M286A constructs. Conclusions and Implications Our data suggest that amino acid residues β3N265, β3F289, β3M286, β3R269 in the β3 subunit, at or near the etomidate/propofol binding site(s), form part of a VA binding pocket. The identification of the binding pocket for VA is essential for elucidating its pharmacological effects and might also help to develop new selective GABAA receptor ligands. PMID:26375408

  20. Structural studies of the actions of anesthetic drugs on the γ-aminobutyric acid type A receptor.

    PubMed

    Akk, Gustav; Steinbach, Joe Henry

    2011-12-01

    The γ-aminobutyric acid type A receptor is the major transmitter-gated inhibitory channel in the central nervous system. The receptor is a target for anesthetics, anticonvulsants, anxiolytics, and sedatives whose actions facilitate the flow of chloride ions through the channel and enhance the inhibitory tone in the brain. Both the kinetic and structural aspects of the actions of modulators of the γ-aminobutyric acid type A receptor are of great importance to understanding the molecular mechanisms of general anesthesia. In this review, the structural rearrangements that take place in the γ-aminobutyric acid type A receptor during channel activation and modulation are described, focusing on data obtained using voltage-clamp fluorometry. Voltage-clamp fluorometry entails the binding of an environmentally sensitive fluorophore molecule to a site of interest in the receptor, and measurement of changes in the fluorescence signal resulting from activation- or modulation-elicited structural changes. Detailed investigations can provide a map of structural changes that underlie or accompany the functional effects of modulators.

  1. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    PubMed Central

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease. PMID:26796668

  2. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    NASA Astrophysics Data System (ADS)

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease.

  3. Altered systemic bile acid homeostasis contributes to liver disease in pediatric patients with intestinal failure

    PubMed Central

    Xiao, Yong-Tao; Cao, Yi; Zhou, Ke-Jun; Lu, Li-Na; Cai, Wei

    2016-01-01

    Intestinal failure (IF)-associated liver disease (IFALD), as a major complication, contributes to significant morbidity in pediatric IF patients. However, the pathogenesis of IFALD is still uncertain. We here investigate the roles of bile acid (BA) dysmetabolism in the unclear pathogenesis of IFALD. It found that the histological evidence of pediatric IF patients exhibited liver injury, which was characterized by liver bile duct proliferation, inflammatory infiltration, hepatocyte apoptosis and different stages of fibrosis. The BA compositions were altered in serum and liver of pediatric IF patients, as reflected by a primary BA dominant composition. In IF patients, the serum FGF19 levels decreased significantly, and were conversely correlated with ileal inflammation grades (r = −0.50, p < 0.05). In ileum, the inflammation grades were inversely associated with farnesoid X receptor (FXR) expression (r = −0.55, p < 0.05). In liver, the expression of induction of the rate-limiting enzyme in bile salt synthesis, cytochrome P450 7a1 (CYP7A1) increased evidently. In conclusion, ileum inflammation decreases FXR expression corresponding to reduce serum FGF19 concentration, along with increased hepatic bile acid synthesis, leading to liver damages in IF patients. PMID:27976737

  4. StAR-related lipid transfer domain protein 5 binds primary bile acids[S

    PubMed Central

    Létourneau, Danny; Lorin, Aurélien; Lefebvre, Andrée; Frappier, Vincent; Gaudreault, Francis; Najmanovich, Rafael; Lavigne, Pierre; LeHoux, Jean-Guy

    2012-01-01

    Steroidogenic acute regulatory-related lipid transfer (START) domain proteins are involved in the nonvesicular intracellular transport of lipids and sterols. The STARD1 (STARD1 and STARD3) and STARD4 subfamilies (STARD4–6) have an internal cavity large enough to accommodate sterols. To provide a deeper understanding on the structural biology of this domain, the binding of sterols to STARD5, a member of the STARD4 subfamily, was monitored. The SAR by NMR [1H-15N heteronuclear single-quantum coherence (HSQC)] approach, complemented by circular dichroism (CD) and isothermal titration calorimetry (ITC), was used. Titration of STARD5 with cholic (CA) and chenodeoxycholic acid (CDCA), ligands of the farnesoid X receptor (FXR), leads to drastic perturbation of the 1H-15N HSQC spectra and the identification of the residues in contact with those ligands. The most perturbed residues in presence of ligands are lining the internal cavity of the protein. Ka values of 1.8·10−4 M−1 and 6.3·104 M−1 were measured for CA and CDCA, respectively. This is the first report of a START domain protein in complex with a sterol ligand. Our original findings indicate that STARD5 may be involved in the transport of bile acids rather than cholesterol. PMID:23018617

  5. Metabotropic glutamate receptors, transmitter output and fatty acids: studies in rat brain slices.

    PubMed Central

    Lombardi, G.; Leonardi, P.; Moroni, F.

    1996-01-01

    1. The effects of (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD), a non-selective agonist of the metabotropic glutamate receptors (mGluRs), have been studied in rat cortical and striatal slices by measuring the depolarization-induced output of D-[3H]-aspartate (D-[3H]-Asp) and of [3H]-glutamate ([3H]-Glu), neosynthesized from [3H]-glutamine. 2. In cortical slices, 1S,3R-ACPD potentiated the depolarization-induced (KCl, 30 mM) output of both D-[3H]-Asp and [3H]-Glu. The potentiation, obtained at 300 microM 1S,3R-ACPD was 65 +/- 6% for D-[3H]-Asp and 56 +/- 10% for [3H]-Glu. Conversely, in striatal slices, 1S,3R-ACPD reduced the depolarization-induced transmitter output. The reduction, obtained at 300 microM of the agonist, was 60 +/- 8% for D-[3H]-Asp and 50 +/- 5% for neosynthesized [3H]-Glu. 3. Bovine serum albumin (BSA, 15 microM), which is able to bind locally produced fatty acids, completely eliminated the potentiating effect 1S,3R-ACPD had on D-[3H]-Asp output from cortical slices. Low concentrations of arachidonic acid (1-10 microM) or of oleic acid (1-10 microM) added to BSA-containing perfusion medium, restored this potentiating effect. BSA, however, had no effect on the inhibitory action of 1S,3R-ACPD in striatal slices. 4. Bromophenacyl bromide (100 microM), an inhibitor of phospholipase A2, and RG80267 (100 microM), an inhibitor of diacylglycerol lipase, have been shown to inhibit fatty acid production. These compounds prevented the potentiating effect of 1S,3R-ACPD on D-[3H]-Asp-output in cortical slices. 5. Indomethacin (100 microM), an inhibitor of cyclo-oxygenases, plus nordihydroguaiaretic acid (100 microM), an inhibitor of lipoxygenases, increased D-[3H]-Asp output in cortical slices perfused with BSA-containing medium. 6. These experiments suggest that the mGluR-mediated potentiation of transmitter output requires the availability of unsaturated fatty acids, such as arachidonic or oleic acids, in cortical slices. In contrast, the m

  6. Retinoic acid represses CYP7A1 expression in human hepatocytes and HepG2 cells by FXR/RXR-dependent and independent mechanisms.

    PubMed

    Cai, Shi-Ying; He, Hongwei; Nguyen, Trong; Mennone, Albert; Boyer, James L

    2010-08-01

    Cholesterol 7alpha-hydroxylase (CYP7A1) plays a key role in maintaining lipid and bile salt homeostasis as it is the rate-limiting enzyme converting cholesterol to bile acids. Deficiency of CYP7A1 leads to hyperlipidemia in man and mouse. Hyperlipidemia is often seen in patients when treated with high-dose retinoic acid (RA), but the molecular mechanisms remain elusive. Our present study revealed that CYP7A1 mRNA expression is greatly repressed by RA in both human hepatocytes and HepG2 cells where increased fibroblast growth factor 19 (FGF19) and small heterodimer partner (SHP) expressions were also observed, suggesting farnesoid X receptor (FXR) and retinoid X receptor (RXR) were activated. Promoter reporter assays demonstrate that all-trans RA (atRA) specifically activated FXR/RXR. However, detailed molecular analyses indicate that this activation is through RXR, whose ligand is 9-cis RA. Knocking down of FXR or RXRalpha by small interference RNA (siRNA) in human hepatocytes increased CYP7A1 basal expression, but the repressive effect of atRA persisted, suggesting there are also FXR/RXR-independent mechanisms mediating atRA repression of CYP7A1 expression. Chromatin immunoprecipitation (ChIP) assay and cell transfection results indicate that PGC-1alpha plays a role in the FXR/RXR-independent mechanism. Our findings may provide a potential explanation for hyperlipidemic side effects observed in some patients treated with high-dose RA.

  7. Targeting metastasis-initiating cells through the fatty acid receptor CD36.

    PubMed

    Pascual, Gloria; Avgustinova, Alexandra; Mejetta, Stefania; Martín, Mercè; Castellanos, Andrés; Attolini, Camille Stephan-Otto; Berenguer, Antoni; Prats, Neus; Toll, Agustí; Hueto, Juan Antonio; Bescós, Coro; Di Croce, Luciano; Benitah, Salvador Aznar

    2017-01-05

    The fact that the identity of the cells that initiate metastasis in most human cancers is unknown hampers the development of antimetastatic therapies. Here we describe a subpopulation of CD44(bright) cells in human oral carcinomas that do not overexpress mesenchymal genes, are slow-cycling, express high levels of the fatty acid receptor CD36 and lipid metabolism genes, and are unique in their ability to initiate metastasis. Palmitic acid or a high-fat diet specifically boosts the metastatic potential of CD36(+) metastasis-initiating cells in a CD36-dependent manner. The use of neutralizing antibodies to block CD36 causes almost complete inhibition of metastasis in immunodeficient or immunocompetent orthotopic mouse models of human oral cancer, with no side effects. Clinically, the presence of CD36(+) metastasis-initiating cells correlates with a poor prognosis for numerous types of carcinomas, and inhibition of CD36 also impairs metastasis, at least in human melanoma- and breast cancer-derived tumours. Together, our results indicate that metastasis-initiating cells particularly rely on dietary lipids to promote metastasis.

  8. Retinoic acid receptor regulation of epimorphic and homeostatic regeneration in the axolotl.

    PubMed

    Nguyen, Matthew; Singhal, Pankhuri; Piet, Judith W; Shefelbine, Sandra J; Maden, Malcolm; Voss, S Randal; Monaghan, James R

    2017-02-15

    Salamanders are capable of regenerating amputated limbs by generating a mass of lineage-restricted cells called a blastema. Blastemas only generate structures distal to their origin unless treated with retinoic acid (RA), which results in proximodistal (PD) limb duplications. Little is known about the transcriptional network that regulates PD duplication. In this study, we target specific retinoic acid receptors (RARs) to either PD duplicate (RA treatment or RARγ agonist) or truncate (RARβ antagonist) regenerating limbs. RARE-EGFP reporter axolotls showed divergent reporter activity in limbs undergoing PD duplication versus truncation, suggesting differences in patterning and skeletal regeneration. Transcriptomics identified expression patterns that explain PD duplication, including upregulation of proximal homeobox gene expression and silencing of distal-associated genes, whereas limb truncation was associated with disrupted skeletal differentiation. RARβ antagonism in uninjured limbs induced a loss of skeletal integrity leading to long bone regression and loss of skeletal turnover. Overall, mechanisms were identified that regulate the multifaceted roles of RARs in the salamander limb including regulation of skeletal patterning during epimorphic regeneration, skeletal tissue differentiation during regeneration, and homeostatic regeneration of intact limbs.

  9. Phosphorylation of histone H3 is functionally linked to retinoic acid receptor β promoter activation

    PubMed Central

    Lefebvre, Bruno; Ozato, Keiko; Lefebvre, Philippe

    2002-01-01

    Ligand-dependent transcriptional activation of retinoic acid receptors (RARs) is a multistep process culminating in the formation of a multimeric co-activator complex on regulated promoters. Several co-activator complexes harbor an acetyl transferase activity, which is required for retinoid-induced transcription of reporter genes. Using murine P19 embryonal carcinoma cells, we examined the relationship between histone post-translational modifications and activation of the endogenous RARβ2 promoter, which is under the control of a canonical retinoic acid response element and rapidly induced upon retinoid treatment. While histones H3 and H4 were constitutively acetylated at this promoter, retinoid agonists induced a rapid phosphorylation at Ser10 of histone H3. A retinoid antagonist, whose activity was independent of co-repressor binding to RAR, could oppose this agonist-induced H3 phosphorylation. Since such post-translational modifications were not observed at several other promoters, we conclude that histone H3 phosphorylation may be a molecular signature of the activated, retinoid-controlled mRARβ2 gene promoter. PMID:11897660

  10. Adiponectin receptor 1 conserves docosahexaenoic acid and promotes photoreceptor cell survival

    PubMed Central

    Rice, Dennis S.; Calandria, Jorgelina M.; Gordon, William C.; Jun, Bokkyoo; Zhou, Yongdong; Gelfman, Claire M.; Li, Songhua; Jin, Minghao; Knott, Eric J.; Chang, Bo; Abuin, Alex; Issa, Tawfik; Potter, David; Platt, Kenneth A.; Bazan, Nicolas G.

    2015-01-01

    The identification of pathways necessary for photoreceptor and retinal pigment epithelium (RPE) function is critical to uncover therapies for blindness. Here we report the discovery of adiponectin receptor 1 (AdipoR1) as a regulator of these cells’ functions. Docosahexaenoic acid (DHA) is avidly retained in photoreceptors, while mechanisms controlling DHA uptake and retention are unknown. Thus, we demonstrate that AdipoR1 ablation results in DHA reduction. In situ hybridization reveals photoreceptor and RPE cell AdipoR1 expression, blunted in AdipoR1−/− mice. We also find decreased photoreceptor-specific phosphatidylcholine containing very long-chain polyunsaturated fatty acids and severely attenuated electroretinograms. These changes precede progressive photoreceptor degeneration in AdipoR1−/− mice. RPE-rich eyecup cultures from AdipoR1−/− reveal impaired DHA uptake. AdipoR1 overexpression in RPE cells enhances DHA uptake, whereas AdipoR1 silencing has the opposite effect. These results establish AdipoR1 as a regulatory switch of DHA uptake, retention, conservation and elongation in photoreceptors and RPE, thus preserving photoreceptor cell integrity. PMID:25736573

  11. The G protein-coupled bile acid receptor, TGR5, stimulates gallbladder filling.

    PubMed

    Li, Tingting; Holmstrom, Sam R; Kir, Serkan; Umetani, Michihisa; Schmidt, Daniel R; Kliewer, Steven A; Mangelsdorf, David J

    2011-06-01

    TGR5 is a G protein-coupled bile acid receptor present in brown adipose tissue and intestine, where its agonism increases energy expenditure and lowers blood glucose. Thus, it is an attractive drug target for treating human metabolic disease. However, TGR5 is also highly expressed in gallbladder, where its functions are less well characterized. Here, we demonstrate that TGR5 stimulates the filling of the gallbladder with bile. Gallbladder volume was increased in wild-type but not Tgr5(-/-) mice by administration of either the naturally occurring TGR5 agonist, lithocholic acid, or the synthetic TGR5 agonist, INT-777. These effects were independent of fibroblast growth factor 15, an enteric hormone previously shown to stimulate gallbladder filling. Ex vivo analyses using gallbladder tissue showed that TGR5 activation increased cAMP concentrations and caused smooth muscle relaxation in a TGR5-dependent manner. These data reveal a novel, gallbladder-intrinsic mechanism for regulating gallbladder contractility. They further suggest that TGR5 agonists should be assessed for effects on human gallbladder as they are developed for treating metabolic disease.

  12. Sialyl Lewis(x) hybridized complement receptor type 1 moderates acid aspiration injury.

    PubMed

    Kyriakides, C; Wang, Y; Austen, W G; Favuzza, J; Kobzik, L; Moore, F D; Hechtman, H B

    2001-12-01

    The potentially enhanced anti-inflammatory effects of the sialyl Lewis(x) (sLe(x))-decorated version of soluble complement receptor type 1 (sCR1) in moderating acid aspiration injury are examined. HCl was instilled in tracheostomy tubes placed in mice, and extravasation of (125)I-labeled albumin in bronchoalveolar lavage (BAL) fluid was used to calculate the vascular permeability index (PI). Neutrophil counts in BAL fluid and immunohistochemistry were performed. PI was moderated by 82% after treatment with sCR1sLe(x) compared with 54% in sCR1-untreated mice (P < 0.05). Respective reductions in PI in mice treated 0.5 and 1 h after acid aspiration with sCR1sLe(x) of 70 and 57% were greater than the decreases in PI of 45 and 38% observed in respective sCR1-treated groups (P < 0.05). BAL fluid neutrophil counts in sCR1sLe(x)-treated mice were significantly less than those in sCR1-treated animals, which were similar to those in untreated mice. Immunohistochemistry stained for sCR1 only on the pulmonary vascular endothelium of sCR1sLe(x)- but not sCR1-treated mice. In conclusion, sCR1sLe(x) moderates permeability by antagonizing complement activation and neutrophil adhesion. Delayed complement and neutrophil antagonism significantly reduces injury.

  13. [EFfect of quinazolone-alkyl-carboxylic acid derivatives on the transmembrane Ca2+ ion flux mediated by AMPA receptors].

    PubMed

    Szárics, Eva; LaszTóczi, Bálint; Nyikos, Lajos; Barabás, Péter; Kovács, Ilona; Skuban, Nina; Nagy, Péter I; Kökösi, József; Takácsné, Novák Krisztina; Kardos, Julianna

    2002-01-01

    The excitatory neurotransmitter, Glu, plays a crucial role in many sensory and motor functions as well as in brain development, learning and memory and it is also involved in the pathogenesis of a number of neurological disorders, including epilepsy, Alzheimer's and Parkinson's diseases. Therefore, the study of Glu receptors (GluRs) is of therapeutical importance. We showed here by fluorescence monitoring of transmembrane Ca2+ ion fluxes in response to (S)-alpha-amino-3-hidroxi-5-metil-4-izoxazol propionic acid ((S)-AMPA) on the time scale of 0.00004-10 s that Ca2+ ion influx proceeds through faster and slower desensitizing receptors. Pharmacological isolation of the slower and faster desensitizing AMPA receptor was possible by fluorescence monitoring of Ca2+ ion translocation in response to (S)-AMPA in the presence and absence of various 2-methyl-4-oxo-3H-quinazoline-3-alkyl-carboxilic acid derivatives (Qxs): the acetic acid Q1 inhibits the slower desensitizing receptor response specifically, while the acetyl-piperidine Q5 is a more potent inhibitor of the faster desensitizing receptor response. In addition, spontaneous interictal activity, as induced by high [K+] conditions in hippocampal slices, was reduced significantly by Q5, suggesting a possible anticonvulsant property of Q5. Substitutions of Qxs into the GluR2 S1S2 binding core were consistent with their effect by causing variable degree of S1S2 bridging interaction as one of the main determinants of AMPA receptor agonist activity. The exploitation of differences between similar receptors will be important in the development and use of drugs with high pharmacological specificity.

  14. Succinate reverses in-vitro platelet inhibition by acetylsalicylic acid and P2Y receptor antagonists.

    PubMed

    Spath, Brigitte; Hansen, Arne; Bokemeyer, Carsten; Langer, Florian

    2012-01-01

    High on-treatment platelet reactivity has been associated with adverse cardiovascular events in patients receiving anti-platelet agents, but the molecular mechanisms underlying this phenomenon remain incompletely understood. Succinate, a citric acid cycle intermediate, is released into the circulation under conditions of mitochondrial dysfunction due to hypoxic organ damage, including sepsis, stroke, and myocardial infarction. Because the G protein-coupled receptor (GPCR) for succinate, SUCNR1 (GPR91), is present on human platelets, we hypothesized that succinate-mediated platelet stimulation may counteract the pharmacological effects of cyclooxygenase-1 and ADP receptor antagonists. To test this hypothesis in a controlled in-vitro study, washed platelets from healthy donors were treated with acetylsalicylic acid (ASA) or small-molecule P2Y(1) or P2Y(12) inhibitors and subsequently analyzed by light transmittance aggregometry using arachidonic acid (AA), ADP and succinate as platelet agonists. Aggregation in response to succinate alone was highly variable with only 29% of donors showing a (mostly delayed) platelet response. In contrast, succinate reproducibly and concentration-dependently (10-1000 µM) enhanced platelet aggregation in response to low concentrations of exogenous ADP. Furthermore, while succinate alone had no effect in the presence of platelet inhibitors, responsiveness of platelets to ADP after pretreatment with P2Y(1) or P2Y(12) antagonists was fully restored, when platelets were co-stimulated with 100 µM succinate. Similarly, succinate completely (at 1000 µM) or partially (at 100 µM) reversed the inhibitory effect of ASA on AA-induced platelet aggregation. In contrast, succinate failed to restore platelet responsiveness in the presence of both ASA and the P2Y(12) antagonist, suggesting that concomitant signaling via different GPCRs was required. Essentially identical results were obtained, when flow cytometric analysis of surface CD62P

  15. Regulation of Phospholipase D Activity and Phosphatidic Acid Production after Purinergic (P2Y6) Receptor Stimulation*

    PubMed Central

    Scott, Sarah A.; Xiang, Yun; Mathews, Thomas P.; Cho, Hyekyung P.; Myers, David S.; Armstrong, Michelle D.; Tallman, Keri A.; O'Reilly, Matthew C.; Lindsley, Craig W.; Brown, H. Alex

    2013-01-01

    Phosphatidic acid (PA) is a lipid second messenger located at the intersection of several lipid metabolism and cell signaling events including membrane trafficking, survival, and proliferation. Generation of signaling PA has long been primarily attributed to the activation of phospholipase D (PLD). PLD catalyzes the hydrolysis of phosphatidylcholine into PA. A variety of both receptor-tyrosine kinase and G-protein-coupled receptor stimulations have been shown to lead to PLD activation and PA generation. This study focuses on profiling the PA pool upon P2Y6 receptor signaling manipulation to determine the major PA producing enzymes. Here we show that PLD, although highly active, is not responsible for the majority of stable PA being produced upon UDP stimulation of the P2Y6 receptor and that PA levels are tightly regulated. By following PA flux in the cell we show that PLD is involved in an initial increase in PA upon receptor stimulation; however, when PLD is blocked, the cell compensates by increasing PA production from other sources. We further delineate the P2Y6 signaling pathway showing that phospholipase Cβ3 (PLCβ3), PLCδ1, DGKζ and PLD are all downstream of receptor activation. We also show that DGKζ is a novel negative regulator of PLD activity in this system that occurs through an inhibitory mechanism with PKCα. These results further define the downstream events resulting in PA production in the P2Y6 receptor signaling pathway. PMID:23723068

  16. The Free Fatty Acid Receptor G Protein-coupled Receptor 40 (GPR40) Protects from Bone Loss through Inhibition of Osteoclast Differentiation*

    PubMed Central

    Wauquier, Fabien; Philippe, Claire; Léotoing, Laurent; Mercier, Sylvie; Davicco, Marie-Jeanne; Lebecque, Patrice; Guicheux, Jérôme; Pilet, Paul; Miot-Noirault, Elisabeth; Poitout, Vincent; Alquier, Thierry; Coxam, Véronique; Wittrant, Yohann

    2013-01-01

    The mechanisms linking fat intake to bone loss remain unclear. By demonstrating the expression of the free fatty acid receptor G-coupled protein receptor 40 (GPR40) in bone cells, we hypothesized that this receptor may play a role in mediating the effects of fatty acids on bone remodeling. Using micro-CT analysis, we showed that GPR40−/− mice exhibit osteoporotic features suggesting a positive role of GPR40 on bone density. In primary cultures of bone marrow, we showed that GW9508, a GRP40 agonist, abolished bone-resorbing cell differentiation. This alteration of the receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation occurred via the inhibition of the nuclear factor κB (NF-κB) signaling pathway as demonstrated by decrease in gene reporter activity, inhibitor of κB kinase (IKKα/β) activation, inhibitor of κB (IkBα) phosphorylation, and nuclear factor of activated T cells 1 (NFATc1) expression. The GPR40-dependent effect of GW9508 was confirmed using shRNA interference in osteoclast precursors and GPR40−/− primary cell cultures. In addition, in vivo administration of GW9508 counteracted ovariectomy-induced bone loss in wild-type but not GPR40−/− mice, enlightening the obligatory role of the GPR40 receptor. Then, in a context of growing prevalence of metabolic and age-related bone disorders, our results demonstrate for the first time in translational approaches that GPR40 is a relevant target for the design of new nutritional and therapeutic strategies to counter bone complications. PMID:23335512

  17. Allosteric regulation of the discriminative responsiveness of retinoic acid receptor to natural and synthetic ligands by retinoid X receptor and DNA.

    PubMed

    Mouchon, A; Delmotte, M H; Formstecher, P; Lefebvre, P

    1999-04-01

    Transcriptional activation by retinoids is mediated through two families of nuclear receptors, all-trans-retinoic acid (RARs) and 9-cis retinoic acid receptors (RXRs). Conformationally restricted retinoids are used to achieve selective activation of RAR isotype alpha, beta or gamma, which reduces side effects in therapeutical applications. Synthetic retinoids mimic some of all-trans retinoic acid biological effects in vivo but interact differently with the ligand binding domain of RARalpha and induce distinct structural transitions of the receptor. In this report, we demonstrate that RAR-selective ligands have distinct quantitative activation properties which are reflected by their abilities to promote interaction of DNA-bound human RXRalpha (hRXRalpha)-hRARalpha heterodimers with the nuclear receptor coactivator (NCoA) SRC-1 in vitro. The hormone response element core motifs spacing defined the relative affinity of liganded heterodimers for two NCoAs, SRC-1 and RIP140. hRXRalpha activating function 2 was critical to confer hRARalpha full responsiveness but not differential sensitivity of hRARalpha to natural or synthetic retinoids. We also provide evidence showing that lysines located in helices 3 and 4, which define part of hRARalpha NCoA binding surface, contribute differently to (i) the transcriptional activity and (ii) the interaction of RXR-RAR heterodimers with SRC-1, when challenged by either natural or RAR-selective retinoids. Thus, ligand structure, DNA, and RXR exert allosteric regulations on hRARalpha conformation organized as a DNA-bound heterodimer. We suggest that the use of physically distinct NCoA binding interfaces may be important in controlling specific genes by conformationally restricted ligands.

  18. Allosteric Regulation of the Discriminative Responsiveness of Retinoic Acid Receptor to Natural and Synthetic Ligands by Retinoid X Receptor and DNA

    PubMed Central

    Mouchon, Arnaud; Delmotte, Marie-Hélène; Formstecher, Pierre; Lefebvre, Philippe

    1999-01-01

    Transcriptional activation by retinoids is mediated through two families of nuclear receptors, all-trans-retinoic acid (RARs) and 9-cis retinoic acid receptors (RXRs). Conformationally restricted retinoids are used to achieve selective activation of RAR isotype α, β or γ, which reduces side effects in therapeutical applications. Synthetic retinoids mimic some of all-trans retinoic acid biological effects in vivo but interact differently with the ligand binding domain of RARα and induce distinct structural transitions of the receptor. In this report, we demonstrate that RAR-selective ligands have distinct quantitative activation properties which are reflected by their abilities to promote interaction of DNA-bound human RXRα (hRXRα)-hRARα heterodimers with the nuclear receptor coactivator (NCoA) SRC-1 in vitro. The hormone response element core motifs spacing defined the relative affinity of liganded heterodimers for two NCoAs, SRC-1 and RIP140. hRXRα activating function 2 was critical to confer hRARα full responsiveness but not differential sensitivity of hRARα to natural or synthetic retinoids. We also provide evidence showing that lysines located in helices 3 and 4, which define part of hRARα NCoA binding surface, contribute differently to (i) the transcriptional activity and (ii) the interaction of RXR-RAR heterodimers with SRC-1, when challenged by either natural or RAR-selective retinoids. Thus, ligand structure, DNA, and RXR exert allosteric regulations on hRARα conformation organized as a DNA-bound heterodimer. We suggest that the use of physically distinct NCoA binding interfaces may be important in controlling specific genes by conformationally restricted ligands. PMID:10082574

  19. Retinoic acid receptor α1 variants, RARα1ΔB and RARα1ΔBC, define a new class of nuclear receptor isoforms

    PubMed Central

    Parrado, Antonio; Despouy, Gilles; Kraïba, Radhia; Pogam, Carole Le; Dupas, Stéphane; Choquette, Maryline; Robledo, Macarena; Larghero, Jérôme; Bui, Hung; Gall, Isabelle Le; Rochette-Egly, Cécile; Chomienne, Christine; Padua, Rose Ann

    2001-01-01

    Retinoic acid (RA) binds and activates retinoid X receptor (RXR)/retinoic acid receptor (RAR) heterodimers, which regulate the transcription of genes that have retinoic acid response elements (RARE). The RAR isotypes (α, β and γ) are comprised of six regions designated A–F. Two isoforms of RARα, 1 and 2, have been identified in humans, which have different A regions generated by differential promoter usage and alternative splicing. We have isolated two new splice variants of RARα1 from human B lymphocytes. In one of these variants, exon 2 is juxtaposed to exon 5, resulting in an altered reading frame and a stop codon. This variant, designated RARα1ΔB, does not code for a functional receptor. In the second variant, exon 2 is juxtaposed to exon 6, maintaining the reading frame. This isoform, designated RARα1ΔBC, retains most of the functional domains of RARα1, but omits the transactivation domain AF-1 and the DNA-binding domain. Consequently, it does not bind nor transactivate RARE on its own. Nevertheless, RARα1ΔBC interacts with RXRα and, as an RXRα/RARα1ΔBC heterodimer, transactivates the DR5 RARE upon all-trans-RA binding. The use of RAR- and RXR-specific ligands shows that, whereas transactivation of the DR5 RARE through the RXRα/RARα1 heterodimer is mediated only by RAR ligands, transactivation through the RXRα/RARα1ΔBC heterodimer is mediated by RAR and RXR ligands. Whilst RARα1 has a broad tissue distribution, RARα1ΔBC has a more heterogeneous distribution, but with significant expression in myeloid cells. RARα1ΔBC is an infrequent example of a functional nuclear receptor which deletes the DNA-binding domain. PMID:11812818

  20. Amino acid residues of bitter taste receptor TAS2R16 that determine sensitivity in primates to β-glycosides

    PubMed Central

    Imai, Hiroo; Suzuki-Hashido, Nami; Ishimaru, Yoshiro; Sakurai, Takanobu; Yin, Lijie; Pan, Wenshi; Ishiguro, Masaji; Masuda, Katsuyoshi; Abe, Keiko; Misaka, Takumi; Hirai, Hirohisa

    2016-01-01

    In mammals, bitter taste is mediated by TAS2Rs, which belong to the family of seven transmembrane G protein-coupled receptors. Since TAS2Rs are directly involved in the interaction between mammals and their dietary sources, it is likely that these genes evolved to reflect species-specific diets during mammalian evolution. Here, we analyzed the amino acids responsible for the difference in sensitivities of TAS2R16s of various primates using a cultured cell expression system. We found that the sensitivity of TAS2R16 varied due to several amino acid residues. Mutation of amino acid residues at E86T, L247M, and V260F in human and langur TAS2R16 for mimicking the macaque TAS2R16 decreased the sensitivity of the receptor in an additive manner, which suggests its contribution to the potency of salicin, possibly via direct interaction. However, mutation of amino acid residues 125 and 133 in human TAS2R16, which are situated in helix 4, to the macaque sequence increased the sensitivity of the receptor. These results suggest the possibility that bitter taste sensitivities evolved independently by replacing specific amino acid residues of TAS2Rs in different primate species to adapt to species-specific food. PMID:27924271

  1. Porcine Sapelovirus Uses α2,3-Linked Sialic Acid on GD1a Ganglioside as a Receptor

    PubMed Central

    Kim, Deok-Song; Son, Kyu-Yeol; Koo, Kyung-Min; Kim, Ji-Yun; Alfajaro, Mia Madel; Park, Jun-Gyu; Hosmillo, Myra; Soliman, Mahmoud; Baek, Yeong-Bin; Cho, Eun-Hyo; Lee, Ju-Hwan; Kang, Mun-Il

    2016-01-01

    ABSTRACT The receptor(s) for porcine sapelovirus (PSV), which causes diarrhea, pneumonia, polioencephalomyelitis, and reproductive disorders in pigs, remains largely unknown. Given the precedent for other picornaviruses which use terminal sialic acids (SAs) as receptors, we examined the role of SAs in PSV binding and infection. Using a variety of approaches, including treating cells with a carbohydrate-destroying chemical (NaIO4), mono- or oligosaccharides (N-acetylneuraminic acid, galactose, and 6′-sialyllactose), linkage-specific sialidases (neuraminidase and sialidase S), lectins (Maakia amurensis lectin and Sambucus nigra lectin), proteases (trypsin and chymotrypsin), and glucosylceramide synthase inhibitors (dl-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol and phospholipase C), we demonstrated that PSV could recognize α2,3-linked SA on glycolipids as a receptor. On the other hand, PSVs had no binding affinity for synthetic histo-blood group antigens (HBGAs), suggesting that PSVs could not use HBGAs as receptors. Depletion of cell surface glycolipids followed by reconstitution studies indicated that GD1a ganglioside, but not other gangliosides, could restore PSV binding and infection, further confirming α2,3-linked SA on GD1a as a PSV receptor. Our results could provide significant information on the understanding of the life cycle of sapelovirus and other picornaviruses. For the broader community in the area of pathogens and pathogenesis, these findings and insights could contribute to the development of affordable, useful, and efficient drugs for anti-sapelovirus therapy. IMPORTANCE The porcine sapelovirus (PSV) is known to cause enteritis, pneumonia, polioencephalomyelitis, and reproductive disorders in pigs. However, the receptor(s) that the PSV utilizes to enter host cells remains largely unknown. Using a variety of approaches, we showed that α2,3-linked terminal sialic acid (SA) on the cell surface GD1a ganglioside could be used for PSV

  2. Orally Administered Berberine Modulates Hepatic Lipid Metabolism by Altering Microbial Bile Acid Metabolism and the Intestinal FXR Signaling Pathway.

    PubMed

    Sun, Runbin; Yang, Na; Kong, Bo; Cao, Bei; Feng, Dong; Yu, Xiaoyi; Ge, Chun; Huang, Jingqiu; Shen, Jianliang; Wang, Pei; Feng, Siqi; Fei, Fei; Guo, Jiahua; He, Jun; Aa, Nan; Chen, Qiang; Pan, Yang; Schumacher, Justin D; Yang, Chung S; Guo, Grace L; Aa, Jiye; Wang, Guangji

    2017-02-01

    Previous studies suggest that the lipid-lowering effect of berberine (BBR) involves actions on the low-density lipoprotein receptor and the AMP-activated protein kinase signaling pathways. However, the implication of these mechanisms is unclear because of the low bioavailability of BBR. Because the main action site of BBR is the gut and intestinal farnesoid X receptor (FXR) plays a pivotal role in the regulation of lipid metabolism, we hypothesized that the effects of BBR on intestinal FXR signaling pathway might account for its pharmacological effectiveness. Using wild type (WT) and intestine-specific FXR knockout (FXR(int-/-)) mice, we found that BBR prevented the development of high-fat-diet-induced obesity and ameliorated triglyceride accumulation in livers of WT, but not FXR(int-/-) mice. BBR increased conjugated bile acids in serum and their excretion in feces. Furthermore, BBR inhibited bile salt hydrolase (BSH) activity in gut microbiota, and significantly increased the levels of tauro-conjugated bile acids, especially tauro-cholic acid(TCA), in the intestine. Both BBR and TCA treatment activated the intestinal FXR pathway and reduced the expression of fatty-acid translocase Cd36 in the liver. These results indicate that BBR may exert its lipid-lowering effect primarily in the gut by modulating the turnover of bile acids and subsequently the ileal FXR signaling pathway. In summary, we provide the first evidence to suggest a new mechanism of BBR action in the intestine that involves, sequentially, inhibiting BSH, elevating TCA, and activating FXR, which lead to the suppression of hepatic expression of Cd36 that results in reduced uptake of long-chain fatty acids in the liver.

  3. Expression and regulation of retinoic acid receptors in human breast cancer cells.

    PubMed

    Roman, S D; Clarke, C L; Hall, R E; Alexander, I E; Sutherland, R L

    1992-04-15

    Retinoic acid is known to inhibit mammary carcinogenesis in rodents and to inhibit proliferation and steroid hormone receptor gene expression in human breast cancer cells. Since these effects are likely to be mediated by nuclear retinoic acid receptors (RARs) the present study was initiated to determine the expression and regulation of RARs in human breast cancer cell lines. Differential cellular gene expression of the RARs was determined by Northern blot analysis of total RNA prepared from 5 ER+ and 6 ER- cell lines. RAR alpha was detected as mRNA species of 2.7 and 3.4 kilobases in all cell lines and the level of gene expression was greater in ER+ cell lines (P less than 0.001). RAR beta mRNA (3.7 kilobases) was detected in seven of the eleven lines tested and was expressed most commonly in ER- cell lines. RAR gamma mRNA was expressed in all cell lines as a transcript of 3.4 kilobases at levels that were similar in both ER+ and ER- cell lines. Retinoic acid failed to regulate the expression of the RAR alpha and RAR gamma genes. The effect of steroid hormones on RAR alpha and RAR gamma mRNA levels was also examined. In four PR+ cell lines (T-47D, BT 474, MCF-7M, and MDA-MB-361), progestins markedly decreased RAR alpha mRNA levels. The progestin effect on RAR alpha levels in T-47D cells was detectable at concentrations of 0.05 nM and was maximal at 1 nM 16 alpha-ethyl-21-hydroxy-19-nor-4-pregnene-3,20-dione ORG 2058, whereas dihydrotestosterone and dexamethasone were without effect. RAR alpha and RAR gamma mRNA levels were rapidly decreased by progestin, and the effect was maximal 3-6 h after ORG 2058 treatment. However, the mRNA loss was transient, and recovery of RAR alpha and RAR gamma mRNA levels was noted 12-24 h after retinoic acid treatment. Although RAR gamma mRNA returned to control levels by 24 h, the decrease in RAR alpha mRNA was maintained at around 50% control until at least 48 h. In summary, RAR alpha and RAR gamma were expressed in all human breast

  4. Activation of lysophosphatidic acid receptor by gintonin inhibits Kv1.2 channel activity: involvement of tyrosine kinase and receptor protein tyrosine phosphatase α.

    PubMed

    Lee, Jun-Ho; Choi, Sun-Hye; Lee, Byung-Hwan; Hwang, Sung-Hee; Kim, Hyeon-Joong; Rhee, Jeehae; Chung, Chihye; Nah, Seung-Yeol

    2013-08-26

    Gintonin is a novel ginseng-derived G protein-coupled lysophosphatidic acid (LPA) receptor ligand. The primary action of gintonin is to elicit a transient increase in [Ca(2+)]i via activation of LPA receptor subtypes. Voltage-gated potassium (Kv) channels play important roles in synaptic transmission in nervous systems. The previous reports have shown that Kv channels can be regulated by Gαq/11 protein-coupled receptor ligands. In the present study, we examined the effects of gintonin on Kv1.2 channel activity expressed in Xenopus oocytes after injection of RNA encoding the human Kv1.2 α subunit. Gintonin treatment inhibited Kv1.2 channel activity in reversible and concentration-dependent manners. The inhibitory effect of gintonin on Kv1.2 channel activity was blocked by active phospholipase C inhibitor, inositol 1,4,5-triphosphate receptor antagonist, and intracellular Ca(2+) chelator. The co-expression of active receptor protein tyrosine phosphatase α (RPTPα) with Kv1.2 channel greatly attenuated gintonin-mediated inhibition of Kv1.2 channel activity, but attenuation was not observed with catalytically inactive RPTPα. Furthermore, neither genistein, a tyrosine kinase inhibitor, nor site-directed mutation of a tyrosine residue (Y132 to Y132F), which is phosphorylated by tyrosine kinase of the N-terminal of the Kv1.2 channel α subunit, significantly attenuated gintonin-mediated inhibition of Kv1.2 channel activity. These results indicate that the gintonin-mediated Kv1.2 channel regulation involves the dual coordination of both tyrosine kinase and RPTPα coupled to this receptor. Finally, gintonin-mediated regulation of Kv1.2 channel activity might explain one of the modulations of gintonin-mediated neuronal activities in nervous systems.

  5. Genetic variants of the unsaturated fatty acid receptor GPR120 relating to obesity in dogs.

    PubMed

    Miyabe, Masahiro; Gin, Azusa; Onozawa, Eri; Daimon, Mana; Yamada, Hana; Oda, Hitomi; Mori, Akihiro; Momota, Yutaka; Azakami, Daigo; Yamamoto, Ichiro; Mochizuki, Mariko; Sako, Toshinori; Tamura, Katsutoshi; Ishioka, Katsumi

    2015-10-01

    G protein-coupled receptor (GPR) 120 is an unsaturated fatty acid receptor, which is associated with various physiological functions. It is reported that the genetic variant of GPR120, p.Arg270His, is detected more in obese people, and this genetic variation functionally relates to obesity in humans. Obesity is a common nutritional disorder also in dogs, but the genetic factors have not ever been identified in dogs. In this study, we investigated the molecular structure of canine GPR120 and searched for candidate genetic variants which may relate to obesity in dogs. Canine GPR120 was highly homologous to those of other species, and seven transmembrane domains and two N-glycosylation sites were conserved. GPR120 mRNA was expressed in lung, jejunum, ileum, colon, hypothalamus, hippocampus, spinal cord, bone marrow, dermis and white adipose tissues in dogs, as those in mice and humans. Genetic variants of GPR120 were explored in client-owned 141 dogs, resulting in that 5 synonymous and 4 non-synonymous variants were found. The variant c.595C>A (p.Pro199Thr) was found in 40 dogs, and the gene frequency was significantly higher in dogs with higher body condition scores, i.e. 0.320 in BCS4-5 dogs, 0.175 in BCS3 dogs and 0.000 in BCS2 dogs. We conclude that c.595C>A (p.Pro199Thr) is a candidate variant relating to obesity, which may be helpful for nutritional management of dogs.

  6. A honey bee odorant receptor for the queen substance 9-oxo-2-decenoic acid.

    PubMed

    Wanner, Kevin W; Nichols, Andrew S; Walden, Kimberly K O; Brockmann, Axel; Luetje, Charles W; Robertson, Hugh M

    2007-09-04

    By using a functional genomics approach, we have identified a honey bee [Apis mellifera (Am)] odorant receptor (Or) for the queen substance 9-oxo-2-decenoic acid (9-ODA). Honey bees live in large eusocial colonies in which a single queen is responsible for reproduction, several thousand sterile female worker bees complete a myriad of tasks to maintain the colony, and several hundred male drones exist only to mate. The "queen substance" [also termed the queen retinue pheromone (QRP)] is an eight-component pheromone that maintains the queen's dominance in the colony. The main component, 9-ODA, acts as a releaser pheromone by attracting workers to the queen and as a primer pheromone by physiologically inhibiting worker ovary development; it also acts as a sex pheromone, attracting drones during mating flights. However, the extent to which social and sexual chemical messages are shared remains unresolved. By using a custom chemosensory-specific microarray and qPCR, we identified four candidate sex pheromone Ors (AmOr10, -11, -18, and -170) from the honey bee genome based on their biased expression in drone antennae. We assayed the pheromone responsiveness of these receptors by using Xenopus oocytes and electrophysiology. AmOr11 responded specifically to 9-ODA (EC50=280+/-31 nM) and not to any of the other seven QRP components, other social pheromones, or floral odors. We did not observe any responses of the other three Ors to any of the eight QRP pheromone components, suggesting 9-ODA is the only QRP component that also acts as a long-distance sex pheromone.

  7. Receptor for Hyaluronic Acid-Mediated Motility is Associated with Poor Survival in Pancreatic Ductal Adenocarcinoma

    PubMed Central

    Cheng, Xiao-Bo; Sato, Norihiro; Kohi, Shiro; Koga, Atsuhiro; Hirata, Keiji

    2015-01-01

    Receptor for hyaluronic acid (HA)-mediated motility (RHAMM) is a nonintegral cell surface receptor involved in the aggressive phenotype in a wide spectrum of human malignancies, but the significance of RHAMM in pancreatic ductal adenocarcinoma (PDAC) remains unknown. In this study, we investigated the expression of RHAMM and its clinical relevance in PDAC. RHAMM mRNA expression was examined in 8 PDAC cell lines and in primary pancreatic cancer and adjacent non-tumor tissues from 14 patients using real-time RT-PCR. Western blotting was carried out to analyze the expression of RHAMM protein in PDAC cell lines. We also investigated the expression patterns of RHAMM protein in tissue samples from 70 PDAC patients using immunohistochemistry. The RHAMM mRNA expression was increased in some PDAC cell lines as compared to a non-tumorous pancreatic epithelial cell line HPDE. The RHAMM mRNA expression was significantly higher in PDAC tissues as compared to corresponding non-tumorous pancreatic tissues (P < 0.0001). The RHAMM protein expression was higher in the vast majority of PDAC cell lines relative to the expression in HPDE. The immunohistochemical analysis revealed strong expression of RHAMM in 52 (74%) PDAC tissues. Strong expression of RHAMM was significantly associated with a shorter survival time (P = 0.038). In multivariate analysis, tumor stage (P = 0.039), residual tumor (P = 0.015), and strong RHAMM expression (P = 0.034) were independent factors predicting poor survival. Strong expression of RHAMM may predict poor survival in PDAC patients and may provide prognostic and, possibly, therapeutic value. PMID:26516356

  8. The antiallodynic action target of intrathecal gabapentin: Ca2+ channels, KATP channels or N-methyl-d-aspartic acid receptors?

    PubMed

    Cheng, Jen-Kun; Chen, Chien-Chuan; Yang, Jia-Rung; Chiou, Lih-Chu

    2006-01-01

    Gabapentin is a novel analgesic whose mechanism of action is not known. We investigated in a postoperative pain model whether adenosine triphosphate (ATP)-sensitive K+ (K(ATP)) channels, N-methyl-d-aspartic acid (NMDA) receptors, and Ca2+ channels are involved in the antiallodynic effect of intrathecal gabapentin. Mechanical allodynia was induced by a paw incision in isoflurane-anesthetized rats. Withdrawal thresholds to von Frey filament stimulation near the incision site were measured before and after incision and after intrathecal drug administration. The antiallodynic effect of gabapentin (100 mug) was not affected by intrathecal pretreatment with antagonists of K(ATP) channels, NMDA receptors or gamma-aminobutyric acid (GABA)(A) receptors. K(ATP) channel openers and GABA(A) receptor agonist, per se, had little effect on the postincision allodynic response. The Ca2+ channel blocker of N-type (omega-conotoxin GVIA, 0.1-3 microg), but not of P/Q-type (omega-agatoxin IVA), L-type (verapamil, diltiazem or nimodipine), or T-type (mibefradil), attenuated the incision-induced allodynia, as did gabapentin. Both the antiallodynic effects of gabapentin and omega-conotoxin GVIA were attenuated by Bay K 8644, an L-type Ca2+ channel activator. These results provide correlative evidence to support the contention that N-type Ca2+ channels, but not K(ATP) channels or NMDA or GABA(A) receptors, might be involved in the antiallodynic effect of intrathecal gabapentin.

  9. Bile acid regulates c-Jun expression through the orphan nuclear receptor SHP induction in gastric cells

    SciTech Connect

    Park, Won Il; Park, Min Jung; An, Jin Kwang; Choi, Yung Hyun; Kim, Hye Young; Cheong, JaeHun Yang, Ung Suk

    2008-05-02

    Bile reflux is considered to be one of the most important causative factors in gastric carcinogenesis, due to the attendant inflammatory changes in the gastric mucosa. In this study, we have assessed the molecular mechanisms inherent to the contribution of bile acid to the transcriptional regulation of inflammatory-related genes. In this study, we demonstrated that bile acid induced the expression of the SHP orphan nuclear receptor at the transcriptional level via c-Jun activation. Bile acid also enhanced the protein interaction of NF-{kappa}B and SHP, thereby resulting in an increase in c-Jun expression and the production of the inflammatory cytokine, TNF{alpha}. These results indicate that bile acid performs a critical function in the regulation of the induction of inflammatory-related genes in gastric cells, and that bile acid-mediated gene expression provides a pre-clue for the development of gastric cellular malformation.

  10. Estrogen dissociates Tau and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor subunit in postischemic hippocampus.

    PubMed

    Cardona-Gómez, Gloria Patricia; Arango-Davila, Cesar; Gallego-Gómez, Juan Carlos; Barrera-Ocampo, Alvaro; Pimienta, Hernan; Garcia-Segura, Luis Miguel

    2006-08-21

    During cerebral ischemia, part of the damage associated with the hyperactivation of glutamate receptors results from the hyperphosphorylation of the microtubule-associated protein Tau. Previous studies have shown that estradiol treatment reduces neural damage after cerebral ischemia. Here, we show that transient occlusion of the middle cerebral artery results in the hyperphosphorylation of Tau and in a significant increase in the association of Tau with glycogen synthase kinase-3beta and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid type glutamate receptor subunits 2/3 in the hippocampus. Estradiol treatment decreased hippocampal injury, inhibited glycogen synthase kinase-3beta and decreased the hyperphosphorylation of Tau and the interaction of Tau with glycogen synthase kinase-3beta and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor. These findings suggest that ischemia produces a strong association between Tau and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor, and estradiol can exert at least part of its neuroprotective activity through inhibition of glycogen synthase kinase-3beta.

  11. Arachidonic Acid Metabolite 19(S)-HETE Induces Vasorelaxation and Platelet Inhibition by Activating Prostacyclin (IP) Receptor

    PubMed Central

    Chennupati, Ramesh; Nüsing, Rolf M.; Offermanns, Stefan

    2016-01-01

    19(S)-hydroxy-eicosatetraenoic acid (19(S)-HETE) belongs to a family of arachidonic acid metabolites produced by cytochrome P450 enzymes, which play critical roles in the regulation of cardiovascular, renal and pulmonary functions. Although it has been known for a long time that 19(S)-HETE has vascular effects, its mechanism of action has remained unclear. In this study we show that 19(S)-HETE induces cAMP accumulation in the human megakaryoblastic leukemia cell line MEG-01. This effect was concentration-dependent with an EC50 of 520 nM, insensitive to pharmacological inhibition of COX-1/2 and required the expression of the G-protein Gs. Systematic siRNA-mediated knock-down of each G-protein coupled receptor (GPCR) expressed in MEG-01 followed by functional analysis identified the prostacyclin receptor (IP) as the mediator of the effects of 19(S)-HETE, and the heterologously expressed IP receptor was also activated by 19(S)-HETE in a concentration-dependent manner with an EC50 of 567 nM. Pretreatment of isolated murine platelets with 19(S)-HETE blocked thrombin-induced platelets aggregation, an effect not seen in platelets from mice lacking the IP receptor. Furthermore, 19(S)-HETE was able to relax mouse mesenteric artery- and thoracic aorta-derived vessel segments. While pharmacological inhibition of COX-1/2 enzymes had no effect on the vasodilatory activity of 19(S)-HETE these effects were not observed in vessels from mice lacking the IP receptor. These results identify a novel mechanism of action for the CYP450-dependent arachidonic acid metabolite 19(S)-HETE and point to the existence of a broader spectrum of naturally occurring prostanoid receptor agonists. PMID:27662627

  12. Imidazoleacetic acid-ribotide induces depression of synaptic responses in hippocampus through activation of imidazoline receptors

    PubMed Central

    Bozdagi, O.; Wang, X. B.; Martinelli, G. P.; Prell, G.; Friedrich, V. L.; Huntley, G. W.

    2011-01-01

    Imidazole-4-acetic acid-ribotide (IAA-RP), an endogenous agonist at imidazoline receptors (I-Rs), is a putative neurotransmitter/regulator in mammalian brain. We studied the effects of IAA-RP on excitatory transmission by performing extracellular and whole cell recordings at Schaffer collateral-CA1 synapses in rat hippocampal slices. Bath-applied IAA-RP induced a concentration-dependent depression of synaptic transmission that, after washout, returned to baseline within 20 min. Maximal decrease occurred with 10 μM IAA-RP, which reduced the slope of field extracellular postsynaptic potentials (fEPSPs) to 51.2 ± 5.7% of baseline at 20 min of exposure. Imidazole-4-acetic acid-riboside (IAA-R; 10 μM), the endogenous dephosphorylated metabolite of IAA-RP, also produced inhibition of fEPSPs. This effect was smaller than that produced by IAA-RP (to 65.9 ± 3.8% of baseline) and occurred after a further 5- to 8-min delay. The frequency, but not the amplitude, of miniature excitatory postsynaptic currents was decreased, and paired-pulse facilitation (PPF) was increased after application of IAA-RP, suggesting a principally presynaptic site of action. Since IAA-RP also has low affinity for α2-adrenergic receptors (α2-ARs), we tested synaptic depression induced by IAA-RP in the presence of α2-ARs, I1-R, or I3-R antagonists. The α2-AR antagonist rauwolscine (100 nM), which blocked the actions of the α2-AR agonist clonidine, did not affect either the IAA-RP-induced synaptic depression or the increase in PPF. In contrast, efaroxan (50 μM), a mixed I1-R and α2-AR antagonist, abolished the synaptic depression induced by IAA-RP and abolished the related increase in PPF. KU-14R, an I3-R antagonist, partially attenuated responses to IAA-RP. Taken together, these data support a role for IAA-RP in modulating synaptic transmission in the hippocampus through activation of I-Rs. PMID:21228308

  13. Blockade of lysophosphatidic acid receptors LPAR1/3 ameliorates lung fibrosis induced by irradiation

    SciTech Connect

    Gan, Lu; Xue, Jian-Xin; Li, Xin; Liu, De-Song; Ge, Yan; Ni, Pei-Yan; Deng, Lin; Lu, You; Jiang, Wei

    2011-05-27

    Highlights: {yields} Lysophosphatidic acid (LPA) levels and its receptors LPAR1/3 transcripts were elevated during the development of radiation-induced lung fibrosis. {yields} Lung fibrosis was obviously alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. {yields} VPC12249 administration effectively inhibited radiation-induced fibroblast accumulation in vivo, and suppressed LPA-induced fibroblast proliferation in vitro. {yields} LPA-LPAR1/3 signaling regulated TGF{beta}1 and CTGF expressions in radiation-challenged lungs, but only influenced CTGF expression in cultured fibroblasts. {yields} LPA-LPAR1/3 signaling induced fibroblast proliferation through a CTGF-dependent pathway, rather than through TGF{beta}1 activation. -- Abstract: Lung fibrosis is a common and serious complication of radiation therapy for lung cancer, for which there are no efficient treatments. Emerging evidence indicates that lysophosphatidic acid (LPA) and its receptors (LPARs) are involved in the pathogenesis of fibrosis. Here, we reported that thoracic radiation with 16 Gy in mice induced development of radiation lung fibrosis (RLF) accompanied by obvious increases in LPA release and LPAR1 and LPAR3 (LPAR1/3) transcripts. RLF was significantly alleviated in mice treated with the dual LPAR1/3 antagonist, VPC12249. VPC12249 administration effectively prolonged animal survival, restored lung structure, inhibited fibroblast accumulation and reduced collagen deposition. Moreover, profibrotic cytokines in radiation-challenged lungs obviously decreased following administration of VPC12249, including transforming growth factor {beta}1 (TGF{beta}1) and connective tissue growth factor (CTGF). In vitro, LPA induced both fibroblast proliferation and CTGF expression in a dose-dependent manner, and both were suppressed by blockade of LPAR1/3. The pro-proliferative activity of LPA on fibroblasts was inhibited by siRNA directed against CTGF. Together, our data suggest that the LPA-LPAR1

  14. Central phencyclidine (PCP) receptor binding is glutamate dependent: evidence for a PCP/excitatory amino acid receptor (EAAR) complex

    SciTech Connect

    Loo, P.; Braunwalder, A.; Lehmann, J.; Williams, M.

    1986-03-01

    PCP and other dissociative anesthetica block the increase in neuronal firing rate evoked by the EAAR agonist, N-methyl-Daspartate. NMDA and other EAAs such as glutamate (glu) have not been previously shown to affect PCP ligand binding. In the present study, using once washed rat forebrain membranes, 10 ..mu..M-glu was found to increase the binding of (/sup 3/H)TCP, a PCP analog, to defined PCP recognition sites by 20%. Removal of glu and aspartate (asp) by extensive washing decreased TCP binding by 75-90%. In these membranes, 10 ..mu..M L-glu increased TCP binding 3-fold. This effect was stereospecific and evoked by other EAAs with the order of activity, L-glu > D-asp > L- asp > NMDA > D-glu > quisqualate. Kainate, GABA, NE, DA, 5-HT, 2-chloroadenosine, oxotremorine and histamine had no effect on TCP binding at concentrations up to 100 ..mu..M. The effects of L-glu were attenuated by the NMDA-type receptor antagonist, 2-amino-7--phosphonoheptanoate (AP7; 10 ..mu..M-1 mM). These findings indicate that EAAS facilitate TCP binding, possibly through NMDA-type receptors. The observed interaction between the PCP receptor and EAARs may reflect the existence of a macromolecular receptor complex similar to that demonstrated for the benzodiazepines and GABA.

  15. PERFLUOROPHOSPHONIC ACID ACTIVATES PEROXISOME PROUFERATOR-ACTIVATED RECEPTOR-ALPHA BUT NOT CONSTITUTIVE ANDROSTANE RECEPTOR IN THE MURINE LIVER

    EPA Science Inventory

    Masurf FS-780 is a commercial perfluoro-chemical mixture that contains C612-perfluoroalkylphosphonic acid (PFPA) derivatives. PFPAs have received recent attention as a previously under recognized subclass of perfluoroalkyl acids (PFAAs) that are found in the environment. The curr...

  16. Retinol-binding protein 4 and its membrane receptor STRA6 control adipogenesis by regulating cellular retinoid homeostasis and retinoic acid receptor α activity.

    PubMed

    Muenzner, Matthias; Tuvia, Neta; Deutschmann, Claudia; Witte, Nicole; Tolkachov, Alexander; Valai, Atijeh; Henze, Andrea; Sander, Leif E; Raila, Jens; Schupp, Michael

    2013-10-01

    Retinoids are vitamin A (retinol) derivatives and complex regulators of adipogenesis by activating specific nuclear receptors, including the retinoic acid receptor (RAR) and retinoid X receptor (RXR). Circulating retinol-binding protein 4 (RBP4) and its membrane receptor STRA6 coordinate cellular retinol uptake. It is unknown whether retinol levels and the activity of RAR and RXR in adipocyte precursors are linked via RBP4/STRA6. Here, we show that STRA6 is expressed in precursor cells and, dictated by the apo- and holo-RBP4 isoforms, mediates bidirectional retinol transport that controls RARα activity and subsequent adipocyte differentiation. Mobilization of retinoid stores in mice by inducing RBP4 secretion from the liver activated RARα signaling in the precursor cell containing the stromal-vascular fraction of adipose tissue. Retinol-loaded holo-RBP4 blocked adipocyte differentiation of cultured precursors by activating RARα. Remarkably, retinol-free apo-RBP4 triggered retinol efflux that reduced cellular retinoids, RARα activity, and target gene expression and enhanced adipogenesis synergistically with ectopic STRA6. Thus, STRA6 in adipocyte precursor cells links nuclear RARα activity to the circulating RBP4 isoforms, whose ratio in obese mice was shifted toward limiting the adipogenic potential of their precursors. This novel cross talk identifies a retinol-dependent metabolic function of RBP4 that may have important implications for the treatment of obesity.

  17. Inhibition of testicular embryonal carcinoma cell tumorigenicity by peroxisome proliferator-activated receptor-β/δ- and retinoic acid receptor-dependent mechanisms.

    PubMed

    Yao, Pei-Li; Chen, Li Ping; Dobrzański, Tomasz P; Phillips, Dylan A; Zhu, Bokai; Kang, Boo-Hyon; Gonzalez, Frank J; Peters, Jeffrey M

    2015-11-03

    Peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) has important physiological functions in control of cell growth, lipid and glucose homeostasis, differentiation and inflammation. To investigate the role of PPARβ/δ in cancer, stable human testicular embryonal carcinoma cell lines were developed that constitutively express PPARβ/δ. Expression of PPARβ/δ caused enhanced activation of the receptor, and this significantly decreased proliferation, migration, invasion, anchorage-independent growth, and also reduced tumor mass and volume of ectopic xenografts derived from NT2/D1 cells compared to controls. The changes observed in xenografts were associated with decreased PPARβ/δ-dependent expression of proliferating cell nuclear antigen and octamer-binding transcription factor-3/4, suggesting suppressed tumor proliferation and induction of differentiation. Inhibition of migration and invasion was mediated by PPARβ/δ competing with formation of the retinoic acid receptor (RAR)/retinoid X receptor (RXR) complex, resulting in attenuation of RARα-dependent matrix metalloproteinase-2 expression and activity. These results demonstrate that PPARβ/δ mediates attenuation of human testicular embryonal carcinoma cell progression through a novel RAR-dependent mechanism and suggest that activation of PPARβ/δ inhibits RAR/RXR dimerization and represents a new therapeutic strategy.

  18. Induction of CYP3A4 and MDR1 gene expression by baicalin, baicalein, chlorogenic acid, and ginsenoside Rf through constitutive androstane receptor- and pregnane X receptor-mediated pathways.

    PubMed

    Li, Yue; Wang, Qi; Yao, Xiaomin; Li, Yan

    2010-08-25

    The herbal products baicalin, baicalein, chlorogenic acid, and ginsenoside Rf have multiple pharmacological effects and are extensively used in alternative and/or complementary therapies. The present study investigated whether baicalin, baicalein, chlorogenic acid, and ginsenoside Rf induced the expression of the cytochrome P450 3A4 (CYP3A4) and multi-drug resistance 1 (MDR1) genes through the pregnane X receptor and constitutive androstane receptor pathways. Real time PCR, western blotting, and a luminescent assay were used to assess the induction of gene expression and activity of CYP3A4 and MDR1 by the test compounds. The interactions of baicalein/chlorogenic acid/ginsenoside Rf with constitutive androstane receptor and pregnane X receptor were evaluated using luciferase reporter and gel shift assays. Baicalein induced the expression of CYP3A4 and MDR1 mRNA by activating pregnane X receptor and constitutive androstane receptor. Chlorogenic acid and ginsenoside Rf showed a relatively weak effect on CYP3A4 promoter activation only in HepG2 cells cotransfected with constitutive androstane receptor and demonstrated no effects on MDR1 via either the constitutive androstane receptor or pregnane X receptor pathway. Baicalin had no effect on either CYP3A4 or MDR1 gene expression. In conclusion, baicalein has the potential to up-regulate CYP3A4 and MDR1 through the direct activation of the constitutive androstane receptor and pregnane X receptor pathways. Chlorogenic acid and ginsenoside Rf only induced constitutive androstane receptor-mediated CYP3A4 expression.

  19. Retinoic acid-related orphan receptor α regulates diurnal rhythm and fasting induction of sterol 12α-hydroxylase in bile acid synthesis.

    PubMed

    Pathak, Preeti; Li, Tiangang; Chiang, John Y L

    2013-12-27

    Sterol 12α-hydroxylase (CYP8B1) is required for cholic acid synthesis and plays a critical role in intestinal cholesterol absorption and pathogenesis of cholesterol gallstone, dyslipidemia, and diabetes. In this study we investigated the underlying mechanism of fasting induction and circadian rhythm of CYP8B1 by a cholesterol-activated nuclear receptor and core clock gene retinoic acid-related orphan receptor α (RORα). Fasting stimulated, whereas restricted-feeding reduced expression of CYP8B1 mRNA and protein. However, fasting and feeding had little effect on the diurnal rhythm of RORα mRNA expression, but fasting increased RORα protein levels by cAMP-activated protein kinase A-mediated phosphorylation and stabilization of the protein. Adenovirus-mediated gene transduction of RORα to mice strongly induced CYP8B1 expression, and increased liver cholesterol and 12α-hydroxylated bile acids in the bile acid pool and serum. A reporter assay identified a functional RORα response element in the CYP8B1 promoter. RORα recruited cAMP response element-binding protein-binding protein (CBP) to stimulate histone acetylation on the CYP8B1 gene promoter. In conclusion, RORα is a key regulator of diurnal rhythm and fasting induction of CYP8B1, which regulates bile acid composition and serum and liver cholesterol levels. Antagonizing RORα activity may be a therapeutic strategy for treating inflammatory diseases such as non-alcoholic fatty liver disease and type 2 diabetes.

  20. Non-classical mechanism of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor channel block by fluoxetine.

    PubMed

    Barygin, Oleg I; Komarova, Margarita S; Tikhonova, Tatiana B; Tikhonov, Denis B

    2015-04-01

    Antidepressants have many targets in the central nervous system. A growing body of data demonstrates the influence of antidepressants on glutamatergic neurotransmission. In the present work, we studied the inhibition of native Ca(2+)-permeable and Ca(2+)-impermeable α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors in rat brain neurons by fluoxetine. The Ca(2+)-impermeable AMPA receptors in CA1 hippocampal pyramidal neurons were weakly affected. The IC50 value for the inhibition of Ca(2+)-permeable AMPA receptors in giant striatal interneurons was 43 ± 7 μM. The inhibition of Ca(2+)-permeable AMPA receptors was voltage dependent, suggesting deep binding in the pore. However, the use dependence of fluoxetine action differed markedly from that of classical AMPA receptor open-channel blockers. Moreover, fluoxetine did not compete with other channel blockers. In contrast to fluoxetine, its membrane-impermeant quaternary analog demonstrated all of the features of channel inhibition typical for open-channel blockers. It is suggested that fluoxetine reaches the binding site through a hydrophobic access pathway. Such a mechanism of block is described for ligands of sodium and calcium channels, but was never found in AMPA receptors. Molecular modeling suggests binding of fluoxetine in the subunit interface; analogous binding was proposed for local anesthetics in closed sodium channels and for benzothiazepines in calcium channels.

  1. Multiplex detection of functional G protein-coupled receptors harboring site-specifically modified unnatural amino acids.

    PubMed

    Naganathan, Saranga; Ray-Saha, Sarmistha; Park, Minyoung; Tian, He; Sakmar, Thomas P; Huber, Thomas

    2015-01-27

    We developed a strategy for identifying positions in G protein-coupled receptors that are amenable to bioorthogonal modification with a peptide epitope tag under cell culturing conditions. We introduced the unnatural amino acid p-azido-l-phenylalanine (azF) into human CC chemokine receptor 5 (CCR5) at site-specific amber codon mutations. We then used strain-promoted azide-alkyne [3+2] cycloaddition to label the azF-CCR5 variants with a FLAG peptide epitope-conjugated aza-dibenzocyclooctyne (DBCO) reagent. A microtiter plate-based sandwich fluorophore-linked immunosorbent assay was used to probe simultaneously the FLAG epitope and the receptor using infrared dye-conjugated antibodies so that the extent of DBCO incorporation, corresponding nominally to labeling efficiency, could be quantified ratiometrically. The extent of incorporation of DBCO at the various sites was evaluated in the context of a recent crystal structure of maraviroc-bound CCR5. We observed that labeling efficiency varied dramatically depending on the topological location of the azF in CCR5. Interestingly, position 109 in transmembrane helix 3, located in a hydrophobic cavity on the extracellular side of the receptor, was labeled most efficiently. Because the bioorthogonal labeling and detection strategy described might be used to introduce a variety of different peptide epitopes or fluorophores into engineered expressed receptors, it might prove to be useful for a wide range of applications, including single-molecule detection studies of receptor trafficking and signaling mechanism.

  2. Multiplex Detection of Functional G Protein-Coupled Receptors Harboring Site-Specifically Modified Unnatural Amino Acids

    PubMed Central

    2015-01-01

    We developed a strategy for identifying positions in G protein-coupled receptors that are amenable to bioorthogonal modification with a peptide epitope tag under cell culturing conditions. We introduced the unnatural amino acid p-azido-l-phenylalanine (azF) into human CC chemokine receptor 5 (CCR5) at site-specific amber codon mutations. We then used strain-promoted azide–alkyne [3+2] cycloaddition to label the azF-CCR5 variants with a FLAG peptide epitope-conjugated aza-dibenzocyclooctyne (DBCO) reagent. A microtiter plate-based sandwich fluorophore-linked immunosorbent assay was used to probe simultaneously the FLAG epitope and the receptor using infrared dye-conjugated antibodies so that the extent of DBCO incorporation, corresponding nominally to labeling efficiency, could be quantified ratiometrically. The extent of incorporation of DBCO at the various sites was evaluated in the context of a recent crystal structure of maraviroc-bound CCR5. We observed that labeling efficiency varied dramatically depending on the topological location of the azF in CCR5. Interestingly, position 109 in transmembrane helix 3, located in a hydrophobic cavity on the extracellular side of the receptor, was labeled most efficiently. Because the bioorthogonal labeling and detection strategy described might be used to introduce a variety of different peptide epitopes or fluorophores into engineered expressed receptors, it might prove to be useful for a wide range of applications, including single-molecule detection studies of receptor trafficking and signaling mechanism. PMID:25524496

  3. Fatty acid synthase regulates estrogen receptor-α signaling in breast cancer cells

    PubMed Central

    Menendez, J A; Lupu, R

    2017-01-01

    Fatty acid synthase (FASN), the key enzyme for endogenous synthesis of fatty acids, is overexpressed and hyperactivated in a biologically aggressive subset of sex steroid-related tumors, including breast carcinomas. Using pharmacological and genetic approaches, we assessed the molecular relationship between FASN signaling and estrogen receptor alpha (ERα) signaling in breast cancer. The small compound C75, a synthetic slow-binding inhibitor of FASN activity, induced a dramatic augmentation of estradiol (E2)-stimulated, ERα-driven transcription. FASN and ERα were both necessary for the synergistic activation of ERα transcriptional activity that occurred following co-exposure to C75 and E2: first, knockdown of FASN expression using RNAi (RNA interference) drastically lowered (>100 fold) the amount of E2 required for optimal activation of ERα-mediated transcriptional activity; second, FASN blockade synergistically increased E2-stimulated ERα-mediated transcriptional activity in ERα-negative breast cancer cells stably transfected with ERα, but not in ERα-negative parental cells. Non-genomic, E2-regulated cross-talk between the ERα and MAPK pathways participated in these phenomena. Thus, treatment with the pure antiestrogen ICI 182 780 or the potent and specific inhibitor of MEK/ERK, U0126, was sufficient to abolish the synergistic nature of the interaction between FASN blockade and E2-stimulated ERα transactivation. FASN inhibition suppressed E2-stimulated breast cancer cell proliferation and anchorage-independent colony formation while promoting the reduction of ERα protein. FASN blockade resulted in the increased expression and nuclear accumulation of the cyclin-dependent kinase inhibitors p21WAF1/CIP1 and p27Kip1, two critical mediators of the therapeutic effects of antiestrogen in breast cancer, while inactivating AKT, a key mediator of E2-promoted anchorage-independent growth. The ability of FASN to regulate E2/ERα signaling may represent a

  4. New Clothes for the Jasmonic Acid Receptor COI1: Delayed Abscission, Meristem Arrest and Apical Dominance

    PubMed Central

    Kim, Joonyup; Dotson, Bradley; Rey, Camila; Lindsey, Joshua; Bleecker, Anthony B.; Binder, Brad M.; Patterson, Sara E.

    2013-01-01

    In a screen for delayed floral organ abscission in Arabidopsis, we have identified a novel mutant of CORONATINE INSENSITIVE 1 (COI1), the F-box protein that has been shown to be the jasmonic acid (JA) co-receptor. While JA has been shown to have an important role in senescence, root development, pollen dehiscence and defense responses, there has been little focus on its critical role in floral organ abscission. Abscission, or the detachment of organs from the main body of a plant, is an essential process during plant development and a unique type of cell separation regulated by endogenous and exogenous signals. Previous studies have indicated that auxin and ethylene are major plant hormones regulating abscission; and here we show that regulation of floral organ abscission is also controlled by jasmonic acid in Arabidopsis thaliana. Our characterization of coi1-1 and a novel allele (coi1-37) has also revealed an essential role in apical dominance and floral meristem arrest. In this study we provide genetic evidence indicating that delayed abscission 4 (dab4-1) is allelic to coi1-1 and that meristem arrest and apical dominance appear to be evolutionarily divergent functions for COI1 that are governed in an ecotype-dependent manner. Further characterizations of ethylene and JA responses of dab4-1/coi1-37 also provide new information suggesting separate pathways for ethylene and JA that control both floral organ abscission and hypocotyl growth in young seedlings. Our study opens the door revealing new roles for JA and its interaction with other hormones during plant development. PMID:23573263

  5. Robust GLP-1 secretion by basic L-amino acids does not require the GPRC6A receptor.

    PubMed

    Clemmensen, Christoffer; Jørgensen, Christinna V; Smajilovic, Sanela; Bräuner-Osborne, Hans

    2017-04-01

    The G protein-coupled receptor GPRC6A (GPCR, Class C, group 6, subtype A) has been proposed to be a sensor for basic L-amino acids that are hypothesized to translate ingestive behaviour to endocrine information. However, the contribution of the GPRC6A receptor to L-amino acid-induced glucagon-like peptide 1 (GLP-1) secretion is unclear. Therefore, to discover whether the GPRC6A receptor is indispensible for amino acid-induced secretion of GLP-1, we treated, with oral gavage, GPRC6A knock-out (KO) and wild-type (WT) littermate mice with GPRC6A ligands (L-arginine and L-ornithine) and assessed GLP-1 levels in circulation. We found that oral administration of both L-arginine and L-ornithine significantly increased total plasma GLP-1 levels to a similar level in GPRC6A KO and WT mice 15 minutes after gavage (both amino acids) and accumulated up to 60 minutes after gavage (L-arginine). Conversely, GLP-1 secretion at the 30- and 60-minute time points in the KO mice was attenuated and did not reach statistical significance. In summary, these data confirm that L-arginine is a potent GLP-1 secretagogue and show that the main effect occurs independently of GPRC6A. In addition, this is the first study to show that also L-ornithine powerfully elicits GLP-1 release in vivo.

  6. Biphasic effects of baclofen on phrenic motoneurons: possible involvement of two types of gamma-aminobutyric acid (GABA) receptors.

    PubMed

    Lalley, P M

    1983-08-01

    Intravenous injections of baclofen have two general dose-dependent effects on phrenic motoneurons in anesthetized cats. Small doses (0.5-1.5 mg/kg) increase the frequency of action potentials recorded from single motoneurons and from the phrenic nerve, whereas large doses (2-10 mg/kg) reduce or abolish action potentials. The increase in frequency produced by small doses is accompanied by membrane depolarization and, in most experiments, by increased input resistance. Large doses hyperpolarize phrenic motoneurons and produce greater increases in input resistance. Extracellular recording during microelectrophoretic application of baclofen reveals only one effect, depression of cell firing, at all effective current strengths. The low dose stimulatory effect of i.v. baclofen is attributed to disinhibition, whereas the depression by large doses is attributed to disfacilitation. During incomplete inhibition by baclofen, CO2 administration further depresses phrenic nerve activity. Bicuculline (100-600 micrograms/kg i.v.) and picrotoxin (900 micrograms/kg i.v.) restore firing depressed by baclofen, whereas strychnine (80-1280 micrograms/kg) does not. 3-Aminopropanesulfonic acid (5-75 mg/kg i.v.) an agonist at gamma-aminobutyric acid-A receptor sites, depresses phrenic nerve activity. It is suggested that the low dose stimulatory effects are related to actions at gamma-aminobutyric acid-B receptors, whereas the high dose depressant effects are related, at least in part, to activation of gamma-aminobutyric acid-A receptors.

  7. Effects of curcumin and tannic acid on the aluminum- and lead-induced oxidative neurotoxicity and alterations in NMDA receptors.

    PubMed

    Tüzmen, Münire Nalan; Yücel, Nilgün Candan; Kalburcu, Tülden; Demiryas, Nazan

    2015-02-01

    Exposure to aluminum (Al) and lead (Pb) can cause brain damage. Also, Pb and Al exposure alte