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Sample records for acid receptor fxr

  1. Bile acid nuclear receptor FXR and digestive system diseases

    PubMed Central

    Ding, Lili; Yang, Li; Wang, Zhengtao; Huang, Wendong

    2015-01-01

    Bile acids (BAs) are not only digestive surfactants but also important cell signaling molecules, which stimulate several signaling pathways to regulate some important biological processes. The bile-acid-activated nuclear receptor, farnesoid X receptor (FXR), plays a pivotal role in regulating bile acid, lipid and glucose homeostasis as well as in regulating the inflammatory responses, barrier function and prevention of bacterial translocation in the intestinal tract. As expected, FXR is involved in the pathophysiology of a wide range of diseases of gastrointestinal tract, including inflammatory bowel disease, colorectal cancer and type 2 diabetes. In this review, we discuss current knowledge of the roles of FXR in physiology of the digestive system and the related diseases. Better understanding of the roles of FXR in digestive system will accelerate the development of FXR ligands/modulators for the treatment of digestive system diseases. PMID:26579439

  2. Bile acid nuclear receptor FXR and digestive system diseases.

    PubMed

    Ding, Lili; Yang, Li; Wang, Zhengtao; Huang, Wendong

    2015-03-01

    Bile acids (BAs) are not only digestive surfactants but also important cell signaling molecules, which stimulate several signaling pathways to regulate some important biological processes. The bile-acid-activated nuclear receptor, farnesoid X receptor (FXR), plays a pivotal role in regulating bile acid, lipid and glucose homeostasis as well as in regulating the inflammatory responses, barrier function and prevention of bacterial translocation in the intestinal tract. As expected, FXR is involved in the pathophysiology of a wide range of diseases of gastrointestinal tract, including inflammatory bowel disease, colorectal cancer and type 2 diabetes. In this review, we discuss current knowledge of the roles of FXR in physiology of the digestive system and the related diseases. Better understanding of the roles of FXR in digestive system will accelerate the development of FXR ligands/modulators for the treatment of digestive system diseases. PMID:26579439

  3. Bile acid-activated nuclear receptor FXR suppresses apolipoprotein A-I transcription via a negative FXR response element

    PubMed Central

    Claudel, Thierry; Sturm, Ekkehard; Duez, Hélène; Torra, Inés Pineda; Sirvent, Audrey; Kosykh, Vladimir; Fruchart, Jean-Charles; Dallongeville, Jean; Hum, Dean W.; Kuipers, Folkert; Staels, Bart

    2002-01-01

    Serum levels of HDL are inversely correlated with the risk of coronary heart disease. The anti-atherogenic effect of HDL is partially mediated by its major protein constituent apoA-I. In this study, we identify bile acids that are activators of the nuclear receptor farnesoid X receptor (FXR) as negative regulators of human apoA-I expression. Intrahepatocellular accumulation of bile acids, as seen in patients with progressive familial intrahepatic cholestasis and biliary atresia, was associated with diminished apoA-I serum levels. In human apoA-I transgenic mice, treatment with the FXR agonist taurocholic acid strongly decreased serum concentrations and liver mRNA levels of human apoA-I, which was associated with reduced serum HDL levels. Incubation of human primary hepatocytes and hepatoblastoma HepG2 cells with bile acids resulted in a dose-dependent downregulation of apoA-I expression. Promoter mutation analysis and gel-shift experiments in HepG2 cells demonstrated that bile acid–activated FXR decreases human apoA-I promoter activity by a negative FXR response element mapped to the C site. FXR bound this site and repressed transcription in a manner independent of retinoid X receptor. The nonsteroidal synthetic FXR agonist GW4064 likewise decreased apoA-I mRNA levels and promoter activity in HepG2 cells. PMID:11927623

  4. 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

  5. Ligand-dependent coactivation of the human bile acid receptor FXR by the peroxisome proliferator-activated receptor gamma coactivator-1alpha.

    PubMed

    Savkur, Rajesh S; Thomas, Jeffrey S; Bramlett, Kelli S; Gao, Yunling; Michael, Laura F; Burris, Thomas P

    2005-01-01

    Peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1alpha) has been shown to play an important role in energy metabolism by coordinating transcriptional programs involved in mitochondrial biogenesis, adaptive thermogenesis, gluconeogenesis, and fatty acid oxidation. PGC-1alpha also plays a crucial role in cholesterol metabolism by serving as a coactivator of the liver X receptor-alpha and inducing the expression of cholesterol 7-alpha-hydroxylase. Here, we demonstrate that PGC-1alpha also functions as an effective coactivator of farnesoid X receptor (FXR), the bile acid receptor. Transient cotransfection assays demonstrate that PGC-1alpha enhances ligand-mediated FXR transcription when either full-length FXR or Gal4 DNA binding domain-FXR-ligand binding domain chimeras were analyzed. Mammalian two-hybrid analyses, glutathione S-transferase affinity chromatography and biochemical coactivator recruitment assays demonstrate ligand-dependent interaction between the two proteins both in vivo and in vitro. PGC-1alpha-mediated coactivation of FXR was highly ligand-dependent and absolutely required an intact activation function-2 (AF-2) domain of FXR and the LXXLL motif in PGC-1alpha. The integrity of the charge clamp was required, further illustrating the role of the ligand binding domain of FXR in PGC-1alpha recognition. Together, these results indicate that PGC-1alpha functions as a potent coactivator for FXR and further implicates its role in the regulation of genes that are involved in bile acid and lipid metabolism.

  6. Conformationally constrained farnesoid X receptor (FXR) agonists: Naphthoic acid-based analogs of GW 4064.

    PubMed

    Akwabi-Ameyaw, Adwoa; Bass, Jonathan Y; Caldwell, Richard D; Caravella, Justin A; Chen, Lihong; Creech, Katrina L; Deaton, David N; Jones, Stacey A; Kaldor, Istvan; Liu, Yaping; Madauss, Kevin P; Marr, Harry B; McFadyen, Robert B; Miller, Aaron B; Iii, Frank Navas; Parks, Derek J; Spearing, Paul K; Todd, Dan; Williams, Shawn P; Wisely, G Bruce

    2008-08-01

    Starting from the known FXR agonist GW 4064 1a, a series of stilbene replacements were prepared. The 6-substituted 1-naphthoic acid 1b was an equipotent FXR agonist with improved developability parameters relative to 1a. Analog 1b also reduced the severity of cholestasis in the ANIT acute cholestatic rat model.

  7. Effects of bile acids and the bile acid receptor FXR agonist on the respiratory rhythm in the in vitro brainstem medulla slice of neonatal Sprague-Dawley rats.

    PubMed

    Zhao, Cong; Wang, Xianbao; Cong, Yuling; Deng, Yi; Xu, Yijun; Chen, Aihua; Yin, Yanru

    2014-01-01

    Intrahepatic cholestasis of pregnancy is always accompanied by adverse fetal outcomes such as malfunctions of respiration. Farnesoid X receptor (FXR) plays a critical role in the homeostasis of bile acids. Thus, we are determined to explore the effects of farnesoid X receptor (FXR) and five bile acids on respiratory rhythm generation and modulation of neonatal rats. Spontaneous periodic respiratory-related rhythmical discharge activity (RRDA) was recorded from hypoglossal nerves during the perfusion of modified Krebs solution. Group 1-6 was each given GW4064 and five bile acids of chenodeoxycholic acid (CDCA), deoxycholic acid (DCA), lithocholic acid (LCA), cholic acid (CA) as well as ursodeoxycholic acid (UDCA) at different concentrations to identify their specific functions on respiratory rhythm modulations. Group 7 was applied to receive FXR blocker Z-guggulsterone and Z-guggulsterone with the above bile acids separately to explore the role of FXR in the respiratory rhythm modulation. Group 8 was given dimethyl sulfoxide (DMSO) as controls. Apart from UDCA, CDCA, DCA LCA and CA all exerted effects on RRDA recorded from hypoglossal nerves in a concentration-dependent manner. Respiratory cycle (RC), Inspiratory time (TI), Expiratory Time (TE) and Integral Amplitude (IA) were influenced and such effects could be reversed by Z-guggulsterone. FXR may contribute to the effects on the modulation of respiratory rhythm exerted by bile acids.

  8. 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.

  9. Design, Synthesis, and Biological Evaluation of Novel Nonsteroidal Farnesoid X Receptor (FXR) Antagonists: Molecular Basis of FXR Antagonism.

    PubMed

    Huang, Huang; Si, Pei; Wang, Lei; Xu, Yong; Xu, Xin; Zhu, Jin; Jiang, Hualiang; Li, Weihua; Chen, Lili; Li, Jian

    2015-07-01

    Farnesoid X receptor (FXR) plays an important role in the regulation of cholesterol, lipid, and glucose metabolism. Recently, several studies on the molecular basis of FXR antagonism have been reported. However, none of these studies employs an FXR antagonist with nonsteroidal scaffold. On the basis of our previously reported FXR antagonist with a trisubstituted isoxazole scaffold, a novel nonsteroidal FXR ligand was designed and used as a lead for structural modification. In total, 39 new trisubstituted isoxazole derivatives were designed and synthesized, which led to pharmacological profiles ranging from agonist to antagonist toward FXR. Notably, compound 5s (4'-[(3-{[3-(2-chlorophenyl)-5-(2-thienyl)isoxazol-4-yl]methoxy}-1H-pyrazol-1-yl)methyl]biphenyl-2-carboxylic acid), containing a thienyl-substituted isoxazole ring, displayed the best antagonistic activity against FXR with good cellular potency (IC50 =12.2 ± 0.2 μM). Eventually, this compound was used as a probe in a molecular dynamics simulation assay. Our results allowed us to propose an essential molecular basis for FXR antagonism, which is consistent with a previously reported antagonistic mechanism; furthermore, E467 on H12 was found to be a hot-spot residue and may be important for the future design of nonsteroidal antagonists of FXR.

  10. 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

  11. Farnesoid X receptor (FXR) gene deficiency impairs urine concentration in mice

    PubMed Central

    Zhang, Xiaoyan; Huang, Shizheng; Gao, Min; Liu, Jia; Jia, Xiao; Han, Qifei; Zheng, Senfeng; Miao, Yifei; Li, Shuo; Weng, Haoyu; Xia, Xuan; Du, Shengnan; Wu, Wanfu; Gustafsson, Jan-Åke; Guan, Youfei

    2014-01-01

    The farnesoid X receptor (FXR) is a ligand-activated transcription factor belonging to the nuclear receptor superfamily. FXR is mainly expressed in liver and small intestine, where it plays an important role in bile acid, lipid, and glucose metabolism. The kidney also has a high FXR expression level, with its physiological function unknown. Here we demonstrate that FXR is ubiquitously distributed in renal tubules. FXR agonist treatment significantly lowered urine volume and increased urine osmolality, whereas FXR knockout mice exhibited an impaired urine concentrating ability, which led to a polyuria phenotype. We further found that treatment of C57BL/6 mice with chenodeoxycholic acid, an FXR endogenous ligand, significantly up-regulated renal aquaporin 2 (AQP2) expression, whereas FXR gene deficiency markedly reduced AQP2 expression levels in the kidney. In vitro studies showed that the AQP2 gene promoter contained a putative FXR response element site, which can be bound and activated by FXR, resulting in a significant increase of AQP2 transcription in cultured primary inner medullary collecting duct cells. In conclusion, the present study demonstrates that FXR plays a critical role in the regulation of urine volume, and its activation increases urinary concentrating capacity mainly via up-regulating its target gene AQP2 expression in the collecting ducts. PMID:24464484

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

    PubMed

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

    2006-01-24

    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.

  13. The Gut Microbiota as a Therapeutic Target in IBD and Metabolic Disease: A Role for the Bile Acid Receptors FXR and TGR5

    PubMed Central

    Baars, Annemarie; Oosting, Annemarie; Knol, Jan; Garssen, Johan; van Bergenhenegouwen, Jeroen

    2015-01-01

    The gut microbiota plays a crucial role in regulating many physiological systems of the host, including the metabolic and immune system. Disturbances in microbiota composition are increasingly correlated with disease; however, the underlying mechanisms are not well understood. Recent evidence suggests that changes in microbiota composition directly affect the metabolism of bile salts. Next to their role in digestion of dietary fats, bile salts function as signaling molecules for bile salt receptors such as Farnesoid X receptor (FXR) and G protein-coupled bile acid receptor (TGR5). Complementary to their role in metabolism, FXR and TGR5 are shown to play a role in intestinal homeostasis and immune regulation. This review presents an overview of evidence showing that changes in bile salt pool and composition due to changes in gut microbial composition contribute to the pathogenesis of inflammatory bowel disease and metabolic disease, possibly through altered activation of TGR5 and FXR. We further discuss how dietary interventions, such as pro- and synbiotics, may be used to treat metabolic disease and inflammatory bowel disease (IBD) through normalization of bile acid dysregulation directly or indirectly through normalization of the intestinal microbiota.

  14. The Gut Microbiota as a Therapeutic Target in IBD and Metabolic Disease: A Role for the Bile Acid Receptors FXR and TGR5

    PubMed Central

    Baars, Annemarie; Oosting, Annemarie; Knol, Jan; Garssen, Johan; van Bergenhenegouwen, Jeroen

    2015-01-01

    The gut microbiota plays a crucial role in regulating many physiological systems of the host, including the metabolic and immune system. Disturbances in microbiota composition are increasingly correlated with disease; however, the underlying mechanisms are not well understood. Recent evidence suggests that changes in microbiota composition directly affect the metabolism of bile salts. Next to their role in digestion of dietary fats, bile salts function as signaling molecules for bile salt receptors such as Farnesoid X receptor (FXR) and G protein-coupled bile acid receptor (TGR5). Complementary to their role in metabolism, FXR and TGR5 are shown to play a role in intestinal homeostasis and immune regulation. This review presents an overview of evidence showing that changes in bile salt pool and composition due to changes in gut microbial composition contribute to the pathogenesis of inflammatory bowel disease and metabolic disease, possibly through altered activation of TGR5 and FXR. We further discuss how dietary interventions, such as pro- and synbiotics, may be used to treat metabolic disease and inflammatory bowel disease (IBD) through normalization of bile acid dysregulation directly or indirectly through normalization of the intestinal microbiota. PMID:27682110

  15. FXR and liver carcinogenesis

    PubMed Central

    Huang, Xiong-fei; Zhao, Wei-yu; Huang, Wen-dong

    2015-01-01

    Farnesoid X receptor (FXR) is a member of the nuclear receptor family and a ligand-modulated transcription factor. In the liver, FXR has been considered a multi-functional cell protector and a tumor suppressor. FXR can suppress liver carcinogenesis via different mechanisms: 1) FXR maintains the normal liver metabolism of bile acids, glucose and lipids; 2) FXR promotes liver regeneration and repair after injury; 3) FXR protects liver cells from death and enhances cell survival; 4) FXR suppresses hepatic inflammation, thereby preventing inflammatory damage; and 5) FXR can directly increase the expression of some tumor-suppressor genes and repress the transcription of several oncogenes. However, inflammation and epigenetic silencing are known to decrease FXR expression during tumorigenesis. The reactivation of FXR function in the liver may be a potential therapeutic approach for patients with liver cancer. PMID:25500874

  16. Activation of the nuclear receptor FXR enhances hepatocyte chemoprotection and liver tumor chemoresistance against genotoxic compounds.

    PubMed

    Vaquero, Javier; Briz, Oscar; Herraez, Elisa; Muntané, Jordi; Marin, Jose J G

    2013-10-01

    The success of pharmacological treatments in primary liver cancers is limited by the marked efficacy of mechanisms of chemoresistance already present in hepatocytes. The role of the nuclear receptor FXR is unclear. Although, in non-treated liver tumors, its expression is reduced, the refractoriness to anticancer drugs is high. Moreover, the treatment with cisplatin up-regulates FXR. The aim of this study was to investigate whether FXR is involved in stimulating chemoprotection/chemoresistance in healthy and tumor liver cells. In human hepatocytes, the activation of FXR with the agonist GW4064 resulted in a significant protection against cisplatin-induced toxicity. In human hepatoma Alexander cells, with negligible endogenous expression of FXR, GW4064 also protected against cisplatin-induced toxicity, but only if they were previously transfected with FXR/RXR. Investigation of 109 genes potentially involved in chemoresistance revealed that only ABCB4, TCEA2, CCL14, CCL15 and KRT13 were up-regulated by FXR activation both in human hepatocytes and FXR/RXR-expressing hepatoma cells. In both models, cisplatin, even in the absence of FXR agonists, such as bile acids and GW4064, was able to up-regulate FXR targets genes, which was due to FXR-mediated trans-activation of response elements in the promoter region. FXR-dependent chemoprotection was also efficient against other DNA-damaging compounds, such as doxorubicin, mitomycin C and potassium dichromate, but not against non-genotoxic drugs, such as colchicine, paclitaxel, acetaminophen, artesunate and sorafenib. In conclusion, ligand-dependent and independent activation of FXR stimulates mechanisms able to enhance the chemoprotection of hepatocytes against genotoxic compounds and to reduce the response of liver tumor cells to certain pharmacological treatments.

  17. Lowering bile acid pool size with a synthetic farnesoid X receptor (FXR) agonist induces obesity and diabetes through reduced energy expenditure.

    PubMed

    Watanabe, Mitsuhiro; Horai, Yasushi; Houten, Sander M; Morimoto, Kohkichi; Sugizaki, Taichi; Arita, Eri; Mataki, Chikage; Sato, Hiroyuki; Tanigawara, Yusuke; Schoonjans, Kristina; Itoh, Hiroshi; Auwerx, Johan

    2011-07-29

    We evaluated the metabolic impact of farnesoid X receptor (FXR) activation by administering a synthetic FXR agonist (GW4064) to mice in which obesity was induced by a high fat diet. Administration of GW4064 accentuated body weight gain and glucose intolerance induced by the high fat diet and led to a pronounced worsening of the changes in liver and adipose tissue. Mechanistically, treatment with GW4064 decreased bile acid (BA) biosynthesis, BA pool size, and energy expenditure, whereas reconstitution of the BA pool in these GW4064-treated animals by BA administration dose-dependently reverted the metabolic abnormalities. Our data therefore suggest that activation of FXR with synthetic agonists is not useful for long term management of the metabolic syndrome, as it reduces the BA pool size and subsequently decreases energy expenditure, translating as weight gain and insulin resistance. In contrast, expansion of the BA pool size, which can be achieved by BA administration, could be an interesting strategy to manage the metabolic syndrome.

  18. Bioenergetic cues shift FXR splicing towards FXRα2 to modulate hepatic lipolysis and fatty acid metabolism

    PubMed Central

    Correia, Jorge C.; Massart, Julie; de Boer, Jan Freark; Porsmyr-Palmertz, Margareta; Martínez-Redondo, Vicente; Agudelo, Leandro Z.; Sinha, Indranil; Meierhofer, David; Ribeiro, Vera; Björnholm, Marie; Sauer, Sascha; Dahlman-Wright, Karin; Zierath, Juleen R.; Groen, Albert K.; Ruas, Jorge L.

    2015-01-01

    Objective Farnesoid X receptor (FXR) plays a prominent role in hepatic lipid metabolism. The FXR gene encodes four proteins with structural differences suggestive of discrete biological functions about which little is known. Methods We expressed each FXR variant in primary hepatocytes and evaluated global gene expression, lipid profile, and metabolic fluxes. Gene delivery of FXR variants to Fxr−/− mouse liver was performed to evaluate their role in vivo. The effects of fasting and physical exercise on hepatic Fxr splicing were determined. Results We show that FXR splice isoforms regulate largely different gene sets and have specific effects on hepatic metabolism. FXRα2 (but not α1) activates a broad transcriptional program in hepatocytes conducive to lipolysis, fatty acid oxidation, and ketogenesis. Consequently, FXRα2 decreases cellular lipid accumulation and improves cellular insulin signaling to AKT. FXRα2 expression in Fxr−/− mouse liver activates a similar gene program and robustly decreases hepatic triglyceride levels. On the other hand, FXRα1 reduces hepatic triglyceride content to a lesser extent and does so through regulation of lipogenic gene expression. Bioenergetic cues, such as fasting and exercise, dynamically regulate Fxr splicing in mouse liver to increase Fxrα2 expression. Conclusions Our results show that the main FXR variants in human liver (α1 and α2) reduce hepatic lipid accumulation through distinct mechanisms and to different degrees. Taking this novel mechanism into account could greatly improve the pharmacological targeting and therapeutic efficacy of FXR agonists. PMID:26909306

  19. Mice with hepatocyte-specific FXR deficiency are resistant to spontaneous but susceptible to cholic acid-induced hepatocarcinogenesis.

    PubMed

    Kong, Bo; Zhu, Yan; Li, Guodong; Williams, Jessica A; Buckley, Kyle; Tawfik, Ossama; Luyendyk, James P; Guo, Grace L

    2016-03-01

    Farnesoid X receptor (FXR) belongs to the nuclear receptor superfamily with its endogenous ligands bile acids. Mice with whole body FXR deficiency develop liver tumors spontaneously, but the underlying mechanism is unclear. Moreover, it is unknown whether FXR deficiency in liver alone serves as a tumor initiator or promoter during liver carcinogenesis. This study aims to evaluate the effects of hepatocyte-specific FXR deficiency (FXR(hep-/-)) in liver tumor formation. The results showed that FXR(hep-/-) mice did not show spontaneous liver tumorigenesis with aging (up to 24 mo of age). Therefore FXR(hep-/-) mice were fed a bile acid (cholic acid)-containing diet alone or along with a liver tumor initiator, diethylnitrosamine (DEN). Thirty weeks later, no tumors were found in wild-type or FXR(hep-/-) mice without any treatment or with DEN only. However, with cholic acid, while only some wild-type mice developed tumors, all FXR(hep-/-) mice presented with severe liver injury and tumors. Interestingly, FXR(hep-/-) mouse livers increased basal expression of tumor suppressor p53 protein, apoptosis, and decreased basal cyclin D1 expression, which may prevent tumor development in FXR(hep-/-) mice. However, cholic acid feeding reversed these effects in FXR(hep-/-) mice, which is associated with an increased cyclin D1 and decreased cell cycle inhibitors. More in-depth analysis indicates that the increased in cell growth might result from disturbance of the MAPK and JAK/Stat3 signaling pathways. In conclusion, this study shows that hepatic FXR deficiency may only serve as a tumor initiator, and increased bile acids is required for tumor formation likely by promoting cell proliferation. PMID:26744468

  20. Knockdown of ATP8B1 expression leads to specific downregulation of the bile acid sensor FXR in HepG2 cells: effect of the FXR agonist GW4064.

    PubMed

    Martínez-Fernández, Pilar; Hierro, Loreto; Jara, Paloma; Alvarez, Luis

    2009-05-01

    Farnesoid X receptor (FXR) is a bile acid-sensing nuclear receptor that controls bile acid homeostasis. It has been suggested that downregulation of FXR contributes to the pathogenesis of an inherited disorder of bile secretion caused by mutations in ATP8B1. We have investigated the relationship between ATP8B1 knockdown and FXR downregulation in the human hepatoblastoma cell line HepG2. Transfection of HepG2 cells with ATP8B1 small interfering RNA (siRNA) duplexes led to a 60% reduction in the endogenous levels of ATP8B1 mRNA and protein and a concomitant decrease in FXR mRNA and protein content, as well as in FXR phosphorylation. This decrease was accompanied by a marked reduction in mRNA levels of a subset of FXR targets, such as bile salt export pump (ABCB11), small heterodimer partner, and uridine 5'-diphosphate-glucuronosyltransferase. ATP8B1 inhibition specifically targeted FXR since mRNA expression of other prominent nuclear receptors, such as pregnane X receptor and constitutive androstane receptor, or liver-enriched transcription factors, such as hepatocyte nuclear factor 1alpha (HNF-1alpha) and HNF-4alpha, was not altered. The expression of other key genes involved in bile acid synthesis, detoxification, and transport also remained unchanged upon ATP8B1 knockdown. Supporting the specificity of the effect, siRNA-mediated silencing of ABCB11, whose defect is associated with another inherited disorder of bile secretion, did not affect FXR expression. Treatment with the synthetic FXR agonist GW4064 was able to partially neutralize ATP8B1 siRNA-mediated FXR downregulation and fully counteract inhibition of FXR target genes. Collectively these findings indicate that ATP8B1 knockdown specifically downregulates FXR, and this action can be circumvented by treatment with FXR agonists.

  1. Navigation in bile acid chemical space: discovery of novel FXR and GPBAR1 ligands

    PubMed Central

    Finamore, Claudia; Festa, Carmen; Renga, Barbara; Sepe, Valentina; Carino, Adriana; Masullo, Dario; Biagioli, Michele; Marchianò, Silvia; Capolupo, Angela; Monti, Maria Chiara; Fiorucci, Stefano; Zampella, Angela

    2016-01-01

    Bile acids are signaling molecules interacting with nuclear receptors and membrane G-protein-coupled receptors. Among these receptors, the farnesoid X receptor (FXR) and the membrane G-coupled receptor (GPBAR1) have gained increasing consideration as druggable receptors and their exogenous dual regulation represents an attractive strategy in the treatment of enterohepatic and metabolic disorders. However, the therapeutic use of dual modulators could be associated to severe side effects and therefore the discovery of selective GPBAR1 and FXR agonists is an essential step in the medicinal chemistry optimization of bile acid scaffold. In this study, a new series of 6-ethylcholane derivatives modified on the tetracyclic core and on the side chain has been designed and synthesized and their in vitro activities on FXR and GPBAR1 were assayed. This speculation resulted in the identification of compound 7 as a potent and selective GPBAR1 agonist and of several derivatives showing potent dual agonistic activity. PMID:27381677

  2. 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.

  3. Bile Acids as Hormones: The FXR-FGF15/19 Pathway.

    PubMed

    Kliewer, Steven A; Mangelsdorf, David J

    2015-01-01

    While it has long been recognized that bile acids are essential for solubilizing lipophilic nutrients in the small intestine, the discovery in 1999 that bile acids serve as ligands for the nuclear receptor farnesoid X receptor (FXR) opened the floodgates in terms of characterizing their actions as selective signaling molecules. Bile acids act on FXR in ileal enterocytes to induce the expression of fibroblast growth factor (FGF)15/19, an atypical FGF that functions as a hormone. FGF15/19 subsequently acts on a cell surface receptor complex in hepatocytes to repress bile acid synthesis and gluconeogenesis, and to stimulate glycogen and protein synthesis. FGF15/19 also stimulates gallbladder filling. Thus, the bile acid-FXR-FGF15/19 signaling pathway regulates diverse aspects of the postprandial enterohepatic response. Pharmacologically, this endocrine pathway provides exciting new opportunities for treating metabolic disease and bile acid-related disorders such as primary biliary cirrhosis and bile acid diarrhea. Both FXR agonists and FGF19 analogs are currently in clinical trials. PMID:26045265

  4. Bile Acids as Hormones: The FXR-FGF15/19 Pathway

    PubMed Central

    Kliewer, Steven A.; Mangelsdorf, David J.

    2015-01-01

    While it has long been recognized that bile acids are essential for solubilizing lipophilic nutrients in the small intestine, the discovery in 1999 that bile acids serve as ligands for the nuclear receptor FXR opened the floodgates in terms of characterizing their actions as selective signaling molecules. Bile acids act on FXR in ileal enterocytes to induce the expression of fibroblast growth factor (FGF) 15/19, an atypical FGF that functions as a hormone. FGF15/19 subsequently acts on a cell surface receptor complex in hepatocytes to repress bile acid synthesis and gluconeogenesis and to stimulate glycogen and protein synthesis. FGF15/19 also stimulates gallbladder filling. Thus, the bile acid-FXR-FGF15/19 signaling pathway regulates diverse aspects of the postprandial enterohepatic response. Pharmacologically, this endocrine pathway provides exciting new opportunities for treating metabolic disease and bile acid-related disorders such as primary biliary cirrhosis and bile acid diarrhea. Both FXR agonists and FGF19 analogs are currently in clinical trials. PMID:26045265

  5. 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

  6. Bile acid signaling through FXR induces intracellular adhesion molecule-1 expression in mouse liver and human hepatocytes.

    PubMed

    Qin, Pu; Borges-Marcucci, Lisa A; Evans, Mark J; Harnish, Douglas C

    2005-08-01

    Previous studies have demonstrated a dramatic induction of inflammatory gene expression in livers from mice fed a high-fat, high-cholesterol diet containing cholate after 3-5 wk. To determine the contribution of cholate in mediating these inductions, C57BL/6 mice were fed a chow diet supplemented with increasing concentrations of cholic acid (CA) for 5 days. A dose-dependent induction in the hepatic levels of TNF-alpha, VCAM-1, ICAM-1, and SAA-2 mRNA were observed. As positive controls, a dose-dependent repression of cholesterol 7alpha-hydroxylase and a dose-dependent induction of small heterodimer partner (SHP) expression were also observed, suggesting that farnesoid X receptor (FXR) was activated. In addition, ICAM-1 and SHP mRNA levels were also induced in primary human hepatocytes when treated with chenodeoxycholic acid or GW4064, a FXR-selective agonist. The involvement of FXR in CA-induced inflammatory gene expression was further investigated in the human hepatic cell line HepG2. Both ICAM-1 and SHP expression were induced in a dose- and time-dependent manner by treatment with the FXR-selective agonist GW4064. Moreover, the induction of ICAM-1 by GW4064 was inhibited by the FXR antagonist guggulsterone or with transfection of FXR siRNA. Finally, the activity of FXR was mapped to a retinoic acid response element (RARE) site containing an imbedded farnesoid X response element (FXRE) on the human ICAM-1 promoter and FXR and retinoid X receptor were demonstrated to bind to this site. Finally, FXR-mediated activation of ICAM-1 could be further enhanced by TNF-alpha cotreatment in hepatocytes, suggesting a potential cooperation between cytokine and bile acid-signaling pathways during hepatic inflammatory events.

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

    PubMed

    Dossa, Avafia Y; Escobar, Oswaldo; Golden, Jamie; Frey, Mark R; Ford, Henri R; Gayer, Christopher P

    2016-01-15

    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.

  8. FXR-dependent and -independent interaction of glucocorticoids with the regulatory pathways involved in the control of bile acid handling by the liver.

    PubMed

    Rosales, R; Romero, M R; Vaquero, J; Monte, M J; Requena, P; Martinez-Augustin, O; Sanchez de Medina, F; Marin, J J G

    2013-03-15

    Treatment with glucocorticoids (GCs) may cause adverse effects, including cholestasis. The ability of dexamethasone, prednisolone and budesonide to affect the liver handling of bile acids (BAs) has been investigated. In rats treated with GCs for 4 days, altered serum and bile BA levels, changed conjugation pattern, and delayed and decreased ability to conjugate/secrete exogenously administered deoxycholate, were found using HPLC-MS/MS. RT-QPCR analyses revealed that GC treatment also induced a down-regulation of liver nuclear receptors (Fxr, Gr and Shp), transporters (Ntcp, Mrp4 and Bcrp) and enzymes (Cyp7a1 and Baat), whereas Bsep, Mrp2 and Cyp27a1 were up-regulated. Human HepG2 and Alexander cell lines were used as in vitro models of liver cells with and without constitutive FXR expression, respectively. In HepG2 cells, GCs induced a decreased expression of FXR and SHP, and inhibited the regulatory effect of GW4064 on FXR-target genes. In Alexander cells, only when they were transfected with FXR+RXR, GW4064 caused up-regulation of SHP and OSTβ, and a down-regulation of CYP27A1. GCs had the opposite effect on these genes, both in the absence and in the presence of FXR expression. Co-transfection of Alexander cells with IR-1-Luc and FXR+RXR revealed that GCs did not inhibit but moderately enhanced FXR activity. Moreover, GCs have a synergistic effect on GW4064-induced FXR activation, whereas chenodeoxycholate and GW4064 have an additive effect. In conclusion, GCs are able to directly or indirectly activate FXR but they also antagonize, through FXR-independent mechanisms, the expression of FXR and FXR target genes involved in the hepatic handling of BAs.

  9. Cysteine Sulfinic Acid Decarboxylase Regulation: A Role for FXR and SHP in Murine Hepatic Taurine Metabolism

    PubMed Central

    Kerr, Thomas A.; Matsumoto, Yuri; Matsumoto, Hitoshi; Xie, Yan; Hirschberger, Lawrence L.; Stipanuk, Martha H.; Anakk, Sayeepriyadarshini; Moore, David D.; Watanabe, Mitsuhiro; Kennedy, Susan

    2014-01-01

    Background Bile acid synthesis is regulated by nuclear receptors including farnesoid X receptor (FXR) and small heterodimer partner (SHP), and by fibroblast growth factor15/19 (FGF15/19). Because bile acid synthesis involves amino acid conjugation, we hypothesized that hepatic cysteine sulfinic acid decarboxylase (CSAD) (a key enzyme in taurine synthesis) is regulated by bile acids. Aims To investigate CSAD regulation by bile acids and CSAD regulatory mechanisms. Methods Mice were fed a control diet or a diet supplemented with either 0.5% cholate or 2% cholestyramine. To gain mechanistic insight into CSAD regulation, we utilized GW4064 (FXR agonist), FGF19, or T-0901317 (LXR agonist) and Shp−/− mice. Tissue mRNA expression was determined by qRT-PCR. Amino acids were measured by HPLC. Results Mice supplemented with dietary cholate exhibited reduced hepatic CSAD mRNA expression while those receiving cholestyramine exhibited increased hepatic CSAD mRNA expression. Activation of FXR suppressed CSAD mRNA expression whereas hepatic CSAD mRNA expression was increased in Shp−/− mice. Hepatic hypotaurine concentration (the product of CSAD) was higher in Shp−/− mice with a corresponding increase in serum (but not hepatic) taurine-conjugated bile acids. FGF19 administration suppressed hepatic CYP7A1 mRNA but did not change CSAD mRNA expression. LXR activation induced CYP7A1 mRNA yet failed to induce CSAD mRNA expression. Conclusion CSAD mRNA expression is physiologically regulated by bile acids in a feedback fashion via mechanisms involving SHP and FXR but not FGF15/19 or LXR. These novel findings implicate bile acids as regulators of CSAD mRNA via mechanisms shared in part with CYP7A1. PMID:24033844

  10. Structural Investigation for Optimization of Anthranilic Acid Derivatives as Partial FXR Agonists by in Silico Approaches

    PubMed Central

    Chen, Meimei; Yang, Xuemei; Lai, Xinmei; Kang, Jie; Gan, Huijuan; Gao, Yuxing

    2016-01-01

    In this paper, a three level in silico approach was applied to investigate some important structural and physicochemical aspects of a series of anthranilic acid derivatives (AAD) newly identified as potent partial farnesoid X receptor (FXR) agonists. Initially, both two and three-dimensional quantitative structure activity relationship (2D- and 3D-QSAR) studies were performed based on such AAD by a stepwise technology combined with multiple linear regression and comparative molecular field analysis. The obtained 2D-QSAR model gave a high predictive ability (R2train = 0.935, R2test = 0.902, Q2LOO = 0.899). It also uncovered that number of rotatable single bonds (b_rotN), relative negative partial charges (RPC−), oprea's lead-like (opr_leadlike), subdivided van der Waal’s surface area (SlogP_VSA2) and accessible surface area (ASA) were important features in defining activity. Additionally, the derived3D-QSAR model presented a higher predictive ability (R2train = 0.944, R2test = 0.892, Q2LOO = 0.802). Meanwhile, the derived contour maps from the 3D-QSAR model revealed the significant structural features (steric and electronic effects) required for improving FXR agonist activity. Finally, nine newly designed AAD with higher predicted EC50 values than the known template compound were docked into the FXR active site. The excellent molecular binding patterns of these molecules also suggested that they can be robust and potent partial FXR agonists in agreement with the QSAR results. Overall, these derived models may help to identify and design novel AAD with better FXR agonist activity. PMID:27070594

  11. Activators of the nuclear hormone receptors PPARalpha and FXR accelerate the development of the fetal epidermal permeability barrier.

    PubMed Central

    Hanley, K; Jiang, Y; Crumrine, D; Bass, N M; Appel, R; Elias, P M; Williams, M L; Feingold, K R

    1997-01-01

    Members of the superfamily of nuclear hormone receptors which are obligate heterodimeric partners of the retinoid X receptor may be important in epidermal development. Here, we examined the effects of activators of the receptors for vitamin D3 and retinoids, and of the peroxisome proliferator activated receptors (PPARs) and the farnesoid X-activated receptor (FXR), on the development of the fetal epidermal barrier in vitro. Skin explants from gestational day 17 rats (term is 22 d) are unstratified and lack a stratum corneum (SC). After incubation in hormone-free media for 3-4 d, a multilayered SC replete with mature lamellar membranes in the interstices and a functionally competent barrier appear. 9-cis or all-trans retinoic acid, 1,25 dihydroxyvitamin D3, or the PPARgamma ligands prostaglandin J2 or troglitazone did not affect the development of barrier function or epidermal morphology. In contrast, activators of the PPARalpha, oleic acid, linoleic acid, and clofibrate, accelerated epidermal development, resulting in mature lamellar membranes, a multilayered SC, and a competent barrier after 2 d of incubation. The FXR activators, all-trans farnesol and juvenile hormone III, also accelerated epidermal barrier development. Activities of beta-glucocerebrosidase and steroid sulfatase, enzymes previously linked to barrier maturation, also increased after treatment with PPARalpha and FXR activators. In contrast, isoprenoids, such as nerolidol, cis-farnesol, or geranylgeraniol, or metabolites in the cholesterol pathway, such as mevalonate, squalene, or 25-hydroxycholesterol, did not alter barrier development. Finally, additive effects were observed in explants incubated with clofibrate and farnesol together in suboptimal concentrations which alone did not affect barrier development. These data indicate a putative physiologic role for PPARalpha and FXR in epidermal barrier development. PMID:9239419

  12. Semisynthetic bile acid FXR and TGR5 agonists: physicochemical properties, pharmacokinetics, and metabolism in the rat.

    PubMed

    Roda, Aldo; Pellicciari, Roberto; Gioiello, Antimo; Neri, Flavia; Camborata, Cecilia; Passeri, Daniela; De Franco, Francesca; Spinozzi, Silvia; Colliva, Carolina; Adorini, Luciano; Montagnani, Marco; Aldini, Rita

    2014-07-01

    We report on the relationship between the structure-pharmacokinetics, metabolism, and therapeutic activity of semisynthetic bile acid analogs, including 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid (a selective farnesoid X receptor [FXR] receptor agonist), 6α-ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid (a specific Takeda G protein-coupled receptor 5 [TGR5] receptor agonist), and 6α-ethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate (a dual FXR/TGR5 agonist). We measured the main physicochemical properties of these molecules, including ionization constants, water solubility, lipophilicity, detergency, and protein binding. Biliary secretion and metabolism and plasma and hepatic concentrations were evaluated by high-pressure liquid chromatography-electrospray-mass spectrometry/mass spectrometry in bile fistula rat and compared with natural analogs chenodeoxycholic, cholic acid, and taurochenodexycholic acid and intestinal bacteria metabolism was evaluated in terms of 7α-dehydroxylase substrate-specificity in anaerobic human stool culture. The semisynthetic derivatives detergency, measured in terms of their critical micellar concentration, was quite similar to the natural analogs. They were slightly more lipophilic than the corresponding natural analogs, evaluated by their 1-octanol water partition coefficient (log P), because of the ethyl group in 6 position, which makes these molecules very stable toward bacterial 7-dehydroxylation. The hepatic metabolism and biliary secretion were different: 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid, as chenodeoxycholic acid, was efficiently conjugated with taurine in the liver and, only in this form, promptly and efficiently secreted in bile. 6α-Ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid was poorly conjugated with taurine because of the steric hindrance of the methyl at C23(S) position metabolized to the C23(R) isomer and partly conjugated with taurine. Conversely, 6

  13. Semisynthetic bile acid FXR and TGR5 agonists: physicochemical properties, pharmacokinetics, and metabolism in the rat.

    PubMed

    Roda, Aldo; Pellicciari, Roberto; Gioiello, Antimo; Neri, Flavia; Camborata, Cecilia; Passeri, Daniela; De Franco, Francesca; Spinozzi, Silvia; Colliva, Carolina; Adorini, Luciano; Montagnani, Marco; Aldini, Rita

    2014-07-01

    We report on the relationship between the structure-pharmacokinetics, metabolism, and therapeutic activity of semisynthetic bile acid analogs, including 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid (a selective farnesoid X receptor [FXR] receptor agonist), 6α-ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid (a specific Takeda G protein-coupled receptor 5 [TGR5] receptor agonist), and 6α-ethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate (a dual FXR/TGR5 agonist). We measured the main physicochemical properties of these molecules, including ionization constants, water solubility, lipophilicity, detergency, and protein binding. Biliary secretion and metabolism and plasma and hepatic concentrations were evaluated by high-pressure liquid chromatography-electrospray-mass spectrometry/mass spectrometry in bile fistula rat and compared with natural analogs chenodeoxycholic, cholic acid, and taurochenodexycholic acid and intestinal bacteria metabolism was evaluated in terms of 7α-dehydroxylase substrate-specificity in anaerobic human stool culture. The semisynthetic derivatives detergency, measured in terms of their critical micellar concentration, was quite similar to the natural analogs. They were slightly more lipophilic than the corresponding natural analogs, evaluated by their 1-octanol water partition coefficient (log P), because of the ethyl group in 6 position, which makes these molecules very stable toward bacterial 7-dehydroxylation. The hepatic metabolism and biliary secretion were different: 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid, as chenodeoxycholic acid, was efficiently conjugated with taurine in the liver and, only in this form, promptly and efficiently secreted in bile. 6α-Ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid was poorly conjugated with taurine because of the steric hindrance of the methyl at C23(S) position metabolized to the C23(R) isomer and partly conjugated with taurine. Conversely, 6

  14. Synthesis and evaluation of (18)F-labeled bile acid compound: a potential PET imaging agent for FXR-related diseases.

    PubMed

    Jia, Lina; Jiang, Dawei; Hu, Pengcheng; Li, Xiao; Shi, Hongcheng; Cheng, Dengfeng; Zhang, Lan

    2014-07-01

    The farnesoid-X-receptor (FXR) is a member of the nuclear hormone receptor superfamily. The FXR has critical functions in maintaining bile acid synthesis and homeostasis, liver regeneration and tumorigenesis, intestinal diseases, intestinal tumorigenesis, cholesterol gallstone disease, cholestasis, and atherosclerosis. FXR expression is strongly downregulated in liver fibrosis, hepatocellular adenoma and hepatocellular carcinoma compared to expression levels in adjacent normal tissues. Chenodeoxycholic acid (CDCA) is the most potent physiological ligand for FXR. CDCA was radiolabeled with (18)F based on the efficiency click reaction of 1,3-dipolar cycloaddition of terminal alkynes and organic azides for noninvasively evaluating the relationship between FXR and FXR-related disease. The PET tracer [(18)F]8 was produced by 'click' labeling and showed a high non-decay corrected radiochemical yield (end of synthesis (EOS) yield=42±3% (n=5) from aqueous [(18)F]fluoride), high radiochemical purity ( >99%), and high specific activity (>320GBq/μmol). [(18)F]8 had a high metabolic stability in vitro and in vivo. PET imaging studies in nude mice indicated a rapid uptake of the tracer into liver tissue with uniform distribution of radioactivity in the liver. Significant accumulation of radioactivity was found in the liver, gallbladder, and intestine, while no obvious uptake was observed in other organs, such as the bladder, heart, and brain. Thus, this PET tracer represents a novel tool for early detection of abnormalities in the liver and staging of neoplasms.

  15. 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

  16. FXR agonist obeticholic acid reduces hepatic inflammation and fibrosis in a rat model of toxic cirrhosis.

    PubMed

    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

  17. DAX1 suppresses FXR transactivity as a novel co-repressor

    SciTech Connect

    Li, Jin; Lu, Yan; Liu, Ruya; Xiong, Xuelian; Zhang, Zhijian; Zhang, Xianfeng; Ning, Guang; Li, Xiaoying

    2011-09-09

    Highlights: {yields} DAX1 is co-localized with FXR and interacts with FXR. {yields} DAX1 acts as a negative regulator of FXR. {yields} Three LXXLL motifs in the N-terminus of DAX1 were required. {yields} DAX1 suppresses FXR transactivation by competing with co-activators. -- Abstract: Bile acid receptor FXR (farnesoid X receptor) is a key regulator of hepatic bile acid, glucose and lipid homeostasis through regulation of numerous genes involved in the process of bile acid, triglyceride and glucose metabolism. DAX1 (dosage-sensitive sex reversal adrenal hypoplasia congenital critical region on X chromosome, gene 1) is an atypical member of the nuclear receptor family due to lack of classical DNA-binding domains and acts primarily as a co-repressor of many nuclear receptors. Here, we demonstrated that DAX1 is co-localized with FXR in the nucleus and acted as a negative regulator of FXR through a physical interaction with FXR. Our study showed that over-expression of DAX1 down-regulated the expression of FXR target genes, whereas knockdown of DAX1 led to their up-regulation. Furthermore, three LXXLL motifs in the N-terminus of DAX1 were required for the full repression of FXR transactivation. In addition, our study characterized that DAX1 suppresses FXR transactivation via competing with co-activators such as SRC-1 and PGC-1{alpha}. In conclusion, DAX1 acts as a co-repressor to negatively modulate FXR transactivity.

  18. Selective targeting of nuclear receptor FXR by avermectin analogues with therapeutic effects on nonalcoholic fatty liver disease

    PubMed Central

    Jin, Lihua; Wang, Rui; Zhu, Yanlin; Zheng, Weili; Han, Yaping; Guo, Fusheng; Ye, Frank Bin; Li, Yong

    2015-01-01

    Non-alcoholic fatty liver disease (NAFLD) has become a predictive factor of death from many diseases. Farnesoid X receptor (FXR) is an ideal target for NAFLD drug development due to its crucial roles in lipid metabolism. The aim of this work is to examine the molecular mechanisms and functional roles of FXR modulation by avermectin analogues in regulating metabolic syndromes like NAFLD. We found that among avermectin analogues studied, the analogues that can bind and activate FXR are effective in regulating metabolic parameters tested, including reducing hepatic lipid accumulation, lowering serum cholesterol and glucose levels, and improving insulin sensitivity, in a FXR dependent manner. Mechanistically, the avermectin analogues that interact with FXR exhibited features as partial agonists, with distinctive properties in modulating coregulator recruitment. Structural features critical for avermectin analogues to selectively bind to FXR were also revealed. This study indicated that in addition to antiparasitic activity, avermectin analogues are promising drug candidates to treat metabolism syndrome including NAFLD by directly targeting FXR. Additionally, the structural features that discriminate the selective binding of FXR by avermectin analogues may provide a unique safe approach to design drugs targeting FXR signaling. PMID:26620317

  19. Deletion of mouse FXR gene disturbs multiple neurotransmitter systems and alters neurobehavior

    PubMed Central

    Huang, Fei; Wang, Tingting; Lan, Yunyi; Yang, Li; Pan, Weihong; Zhu, Yonghui; Lv, Boyang; Wei, Yuting; Shi, Hailian; Wu, Hui; Zhang, Beibei; Wang, Jie; Duan, Xiaofeng; Hu, Zhibi; Wu, Xiaojun

    2015-01-01

    Farnesoid X receptor (FXR) is a nuclear hormone receptor involved in bile acid synthesis and homeostasis. Dysfunction of FXR is involved in cholestasis and atherosclerosis. FXR is prevalent in liver, gallbladder, and intestine, but it is not yet clear whether it modulates neurobehavior. In the current study, we tested the hypothesis that mouse FXR deficiency affects a specific subset of neurotransmitters and results in an unique behavioral phenotype. The FXR knockout mice showed less depressive-like and anxiety-related behavior, but increased motor activity. They had impaired memory and reduced motor coordination. There were changes of glutamatergic, GABAergic, serotoninergic, and norepinephrinergic neurotransmission in either hippocampus or cerebellum. FXR deletion decreased the amount of the GABA synthesis enzyme GAD65 in hippocampus but increased GABA transporter GAT1 in cerebral cortex. FXR deletion increased serum concentrations of many bile acids, including taurodehydrocholic acid, taurocholic acid, deoxycholic acid (DCA), glycocholic acid (GCA), tauro-α-muricholic acid, tauro-ω-muricholic acid, and hyodeoxycholic acid (HDCA). There were also changes in brain concentrations of taurocholic acid, taurodehydrocholic acid, tauro-ω-muricholic acid, tauro-β-muricholic acid, deoxycholic acid, and lithocholic acid (LCA). Taken together, the results from studies with FXR knockout mice suggest that FXR contributes to the homeostasis of multiple neurotransmitter systems in different brain regions and modulates neurobehavior. The effect appears to be at least partially mediated by bile acids that are known to cross the blood-brain barrier (BBB) inducing potential neurotoxicity. PMID:25870546

  20. Differential regulation of bile acid homeostasis by the farnesoid X receptor in liver and intestine.

    PubMed

    Kim, Insook; Ahn, Sung-Hoon; Inagaki, Takeshi; Choi, Mihwa; Ito, Shinji; Guo, Grace L; Kliewer, Steven A; Gonzalez, Frank J

    2007-12-01

    Bile acid concentrations are controlled by a feedback regulatory pathway whereby activation of the farnesoid X receptor (FXR) represses transcription of both the CYP7A1 gene, encoding the rate-limiting enzyme in the classic bile acid synthesis pathway, and the CYP8B1 gene, required for synthesis of cholic acid. The tissue-specific roles of FXR were examined using liver- and intestine-specific FXR-null models. FXR deficiency in either liver (Fxr DeltaL) or intestine (Fxr DeltaIE) increased bile acid pool size. Treatment with the FXR-selective agonist GW4064 significantly repressed CYP7A1 in Fxr DeltaL mice but not Fxr DeltaIE mice, demonstrating that activation of FXR in intestine but not liver is required for short-term repression of CYP7A1 in liver. This intestinal-specific effect of FXR is likely mediated through induction of the hormone FGF15, which suppresses CYP7A1. In comparison to CYP7A1, FXR-mediated repression of CYP8B1 was more dependent on the presence of FXR in liver and less dependent on its presence in intestine. Consistent with these findings, recombinant FGF15 repressed CYP7A1 mRNA levels without affecting CYP8B1 expression. These data provide evidence that FXR-mediated repression of bile acid synthesis requires the complementary actions of FXR in both liver and intestine and reveal mechanistic differences in feedback repression of CYP7A1 and CYP8B1.

  1. 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.

  2. Bile acid receptor agonist GW4064 regulates PPARγ coactivator-1α expression through estrogen receptor-related receptor α.

    PubMed

    Dwivedi, Shailendra Kumar Dhar; Singh, Nidhi; Kumari, Rashmi; Mishra, Jay Sharan; Tripathi, Sarita; Banerjee, Priyam; Shah, Priyanka; Kukshal, Vandana; Tyagi, Abdul Malik; Gaikwad, Anil Nilkanth; Chaturvedi, Rajnish Kumar; Mishra, Durga Prasad; Trivedi, Arun Kumar; Sanyal, Somali; Chattopadhyay, Naibedya; Ramachandran, Ravishankar; Siddiqi, Mohammad Imran; Bandyopadhyay, Arun; Arora, Ashish; Lundåsen, Thomas; Anakk, Sayee Priyadarshini; Moore, David D; Sanyal, Sabyasachi

    2011-06-01

    Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) is induced in energy-starved conditions and is a key regulator of energy homeostasis. This makes PGC-1α an attractive therapeutic target for metabolic syndrome and diabetes. In our effort to identify new regulators of PGC-1α expression, we found that GW4064, a widely used synthetic agonist for the nuclear bile acid receptor [farnesoid X receptor (FXR)] strongly enhances PGC-1α promoter reporter activity, mRNA, and protein expression. This induction in PGC-1α concomitantly enhances mitochondrial mass and expression of several PGC-1α target genes involved in mitochondrial function. Using FXR-rich or FXR-nonexpressing cell lines and tissues, we found that this effect of GW4064 is not mediated directly by FXR but occurs via activation of estrogen receptor-related receptor α (ERRα). Cell-based, biochemical and biophysical assays indicate GW4064 as an agonist of ERR proteins. Interestingly, FXR disruption alters GW4064 induction of PGC-1α mRNA in a tissue-dependent manner. Using FXR-null [FXR knockout (FXRKO)] mice, we determined that GW4064 induction of PGC-1α expression is not affected in oxidative soleus muscles of FXRKO mice but is compromised in the FXRKO liver. Mechanistic studies to explain these differences revealed that FXR physically interacts with ERR and protects them from repression by the atypical corepressor, small heterodimer partner in liver. Together, this interplay between ERRα-FXR-PGC-1α and small heterodimer partner offers new insights into the biological functions of ERRα and FXR, thus providing a knowledge base for therapeutics in energy balance-related pathophysiology.

  3. FXR-induced lysine-specific histone demethylase, LSD1, reduces hepatic bile acid levels and protects the liver against bile acid toxicity

    PubMed Central

    Kim, Young-Chae; Fang, Sungsoon; Byun, Sangwon; Seok, Sunmi; Kemper, Byron; Kemper, Jongsook Kim

    2015-01-01

    Bile acids (BAs) function as endocrine signaling molecules that activate multiple nuclear and membrane receptor signaling pathways to control fed-state metabolism. Since the detergent-like property of BAs causes liver damage at high concentrations, hepatic BA levels must be tightly regulated. BA homeostasis is regulated largely at the level of transcription by nuclear receptors, particularly the primary bile acid receptor, farnesoid X receptor (FXR), and small heterodimer partner (SHP) that inhibits BA synthesis by recruiting repressive histone-modifying enzymes. Although histone modifiers have been shown to regulate BA-responsive genes, their in vivo functions remain unclear. Here we show that lysine-specific histone demethylase1 (LSD1) is directly induced by BA-activated FXR, is recruited to BA synthetic genes, Cyp7a1 and Cyp8b1, and the BA uptake transporter gene, Ntcp, and removes a gene-activation mark, tri-methylated histone H3 lysine-4, leading to gene repression. LSD1 recruitment was dependent on SHP, and LSD1-mediated demethylation of H3K4-me3 was required for additional repressive histone modifications, H3K9/K14 deacetylation and H3K9 methylation. BA overload, feeding 0.5% cholic acid chow for 6 days, resulted in adaptive responses of altered expression of hepatic genes involved in BA synthesis, transport, and detoxification/conjugation. In contrast, adenoviral-mediated downregulation of hepatic LSD1 blunted these responses, which led to substantial increases in liver and serum BA levels, serum AST/ALT levels, and hepatic inflammation. This study identifies LSD1 as a novel histone-modifying enzyme in the orchestrated regulation mediated by the FXR and SHP that reduces hepatic BA levels and protects the liver against BA toxicity. PMID:25545350

  4. 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.

  5. Farnesoid X receptor, the bile acid sensing nuclear receptor, in liver regeneration

    PubMed Central

    Li, Guodong; L. Guo, Grace

    2015-01-01

    The liver is unique in regenerative potential, which could recover the lost mass and function after injury from ischemia and resection. The underlying molecular mechanisms of liver regeneration have been extensively studied in the past using the partial hepatectomy (PH) model in rodents, where 2/3 PH is carried out by removing two lobes. The whole process of liver regeneration is complicated, orchestrated event involving a network of connected interactions, which still remain fully elusive. Bile acids (BAs) are ligands of farnesoid X receptor (FXR), a nuclear receptor of ligand-activated transcription factor. FXR has been shown to be highly involved in liver regeneration. BAs and FXR not only interact with each other but also regulate various downstream targets independently during liver regeneration. Moreover, recent findings suggest that tissue-specific FXR also contributes to liver regeneration significantly. These novel findings suggest that FXR has much broader role than regulating BA, cholesterol, lipid and glucose metabolism. Therefore, these researches highlight FXR as an important pharmaceutical target for potential use of FXR ligands to regulate liver regeneration in clinic. This review focuses on the roles of BAs and FXR in liver regeneration and the current underlying molecular mechanisms which contribute to liver regeneration. PMID:26579433

  6. Substituted isoxazole analogs of farnesoid X receptor (FXR) agonist GW4064

    SciTech Connect

    Bass, Jonathan Y.; Caldwell, Richard D.; Caravella, Justin A.; Chen, Lihong; Creech, Katrina L.; Deaton, David N.; Madauss, Kevin P.; Marr, Harry B.; McFadyen, Robert B.; Miller, Aaron B.; Parks, Derek J.; Todd, Dan; Williams, Shawn P.; Wisely, G. Bruce

    2010-09-27

    Starting from the known FXR agonist GW 4064 1a, a series of alternately 3,5-substituted isoxazoles was prepared. Several of these analogs were potent full FXR agonists. A subset of this series, with a tether between the isoxazole ring and the 3-position aryl substituent, were equipotent FXR agonists to GW 4064 1a, with the 2,6-dimethyl phenol analog 1t having greater FRET FXR potency than GW 4064 1a.

  7. Substituted isoxazole analogs of farnesoid X receptor (FXR) agonist GW4064.

    PubMed

    Bass, Jonathan Y; Caldwell, Richard D; Caravella, Justin A; Chen, Lihong; Creech, Katrina L; Deaton, David N; Madauss, Kevin P; Marr, Harry B; McFadyen, Robert B; Miller, Aaron B; Parks, Derek J; Todd, Dan; Williams, Shawn P; Wisely, G Bruce

    2009-06-01

    Starting from the known FXR agonist GW 4064 1a, a series of alternately 3,5-substituted isoxazoles was prepared. Several of these analogs were potent full FXR agonists. A subset of this series, with a tether between the isoxazole ring and the 3-position aryl substituent, were equipotent FXR agonists to GW 4064 1a, with the 2,6-dimethyl phenol analog 1t having greater FRET FXR potency than GW 4064 1a.

  8. Curcumin protects ANIT-induced cholestasis through signaling pathway of FXR-regulated bile acid and inflammation.

    PubMed

    Yang, Fan; Tang, Xiaowen; Ding, Lili; Zhou, Yue; Yang, Qiaoling; Gong, Junting; Wang, Guangyun; Wang, Zhengtao; Yang, Li

    2016-01-01

    Cholestasis is a clinically significant symptom and widely associated with liver diseases, however, there are very few effective therapies for cholestasis. Danning tablet (DNT, a Chinese patent medicine preparation) has been clinically used to treat human liver and gallbladder diseases for more than 20 years in China. However, which ingredients of DNT contributed to this beneficial effect and their mechanistic underpinnings have been largely unknown. In the present study, we discovered that DNT not only demonstrated greater benefits for cholecystitis patients after cholecystectomy surgery in clinic but also showed protective effect against alpha-naphthylisothiocyanate (ANIT)-induced cholestasis model in rodent. Curcumin, one major compound derived from DNT, exerted the protective effect against cholestasis through farnesoid X receptor (FXR), which has been focused as potential therapeutic targets for treating cholestasis. The underlying mechanism of curcumin against cholestasis was restoring bile acid homeostasis and antagonizing inflammatory responses in a FXR-dependent manner and in turn contributed to overall cholestasis attenuation. Collectively, curcumin can be served as a potential treatment option for liver injury with cholestasis. PMID:27624003

  9. Curcumin protects ANIT-induced cholestasis through signaling pathway of FXR-regulated bile acid and inflammation

    PubMed Central

    Yang, Fan; Tang, Xiaowen; Ding, Lili; zhou, Yue; Yang, Qiaoling; Gong, Junting; Wang, Guangyun; Wang, Zhengtao; Yang, Li

    2016-01-01

    Cholestasis is a clinically significant symptom and widely associated with liver diseases, however, there are very few effective therapies for cholestasis. Danning tablet (DNT, a Chinese patent medicine preparation) has been clinically used to treat human liver and gallbladder diseases for more than 20 years in China. However, which ingredients of DNT contributed to this beneficial effect and their mechanistic underpinnings have been largely unknown. In the present study, we discovered that DNT not only demonstrated greater benefits for cholecystitis patients after cholecystectomy surgery in clinic but also showed protective effect against alpha-naphthylisothiocyanate (ANIT)-induced cholestasis model in rodent. Curcumin, one major compound derived from DNT, exerted the protective effect against cholestasis through farnesoid X receptor (FXR), which has been focused as potential therapeutic targets for treating cholestasis. The underlying mechanism of curcumin against cholestasis was restoring bile acid homeostasis and antagonizing inflammatory responses in a FXR-dependent manner and in turn contributed to overall cholestasis attenuation. Collectively, curcumin can be served as a potential treatment option for liver injury with cholestasis. PMID:27624003

  10. FXR regulates organic solute transporters alpha and beta in the adrenal gland, kidney, and intestine.

    PubMed

    Lee, Hans; Zhang, Yanqiao; Lee, Florence Y; Nelson, Stanley F; Gonzalez, Frank J; Edwards, Peter A

    2006-01-01

    Expression of the farnesoid X receptor (FXR; NR1H4) is limited to the liver, intestine, kidney, and adrenal gland. However, the role of FXR in the latter two organs is unknown. In the current study, we performed microarray analysis using RNA from H295R cells infected with constitutively active FXR. Several putative FXR target genes were identified, including the organic solute transporters alpha and beta (OSTalpha and OSTbeta). Electromobility shift assays and promoter-reporter studies identified functional farnesoid X receptor response elements (FXREs) in the promoters of both human genes. These FXREs are conserved in both mouse genes. Treatment of wild-type mice with 3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chloro-stilben-4-yl)-oxymethyl-5-isopropyl-isoxazole (GW4064), a synthetic FXR agonist, induced OSTalpha and OSTbeta mRNAs in the intestine and kidney. Both mRNAs were also induced when wild-type, but not FXR-deficient (FXR-/-), adrenals were cultured in the presence of GW4064. OSTalpha and OSTbeta mRNA levels were also induced in the adrenals and kidneys of wild-type, but not FXR-/-, mice after the increase of plasma bile acids in response to the hepatotoxin alpha-naphthylisothiocyanate. Finally, overexpression of human OSTalpha and OSTbeta facilitated the uptake of conjugated chenodeoxycholate and the activation of FXR target genes. These results demonstrate that OSTalpha and OSTbeta are novel FXR target genes that are expressed in the adrenal gland, kidney, and intestine.

  11. Differential activation of the human farnesoid X receptor depends on the pattern of expressed isoforms and the bile acid pool composition.

    PubMed

    Vaquero, Javier; Monte, Maria J; Dominguez, Mercedes; Muntané, Jordi; Marin, Jose J G

    2013-10-01

    The farnesoid X receptor (FXR) is a key sensor in bile acid homeostasis. Although four human FXR isoforms have been identified, the physiological role of this diversity is poorly understood. Here we investigated their subcellular localization, agonist sensitivity and response of target genes. Measurement of mRNA revealed that liver predominantly expressed FXRα1(+/-), whereas FXRα2(+/-) were the most abundant isoforms in kidney and intestine. In all cases, the proportion of FXRα(1/2)(+) and FXRα(1/2)(-) isoforms, i.e., with and without a 12bp insert, respectively, was approximately 50%. When FXR was expressed in liver and intestinal cells the magnitude of the response to GW4064 and bile acids differs among FXR isoforms. In both cell types the strongest response was that of FXRα1(-). Different efficacy of bile acids species to activate FXR was found. The four FXR isoforms shared the order of sensitivity to bile acids species. When in FXR-deficient cells FXR was transfected, unconjugated, but not taurine- and glycine-amidated bile acids, were able to activate FXR. In contrast, human hepatocytes and cell lines showing an endogenous expression of FXR were sensitive to both unconjugated and conjugated bile acids. This suggests that to activate FXR conjugated, but not unconjugated, bile acids require additional component(s) of the intracellular machinery not related with uptake processes, which are missing in some tumor cells. In conclusion, cell-specific pattern of FXR isoforms determine the overall tissue sensitivity to FXR agonists and may be involved in the differential response of FXR target genes to FXR activation.

  12. Alisol B 23-acetate protects against ANIT-induced hepatotoxity and cholestasis, due to FXR-mediated regulation of transporters and enzymes involved in bile acid homeostasis

    SciTech Connect

    Meng, Qiang; Chen, Xin-li; Wang, Chang-yuan; Liu, Qi; Sun, Hui-jun; Sun, Peng-yuan; Huo, Xiao-kui; Liu, Zhi-hao; Yao, Ji-hong; Liu, Ke-xin

    2015-03-15

    Intrahepatic cholestasis is a clinical syndrome with systemic and intrahepatic accumulation of excessive toxic bile acids that ultimately cause hepatobiliary injury. Appropriate regulation of bile acids in hepatocytes is critically important for protection against liver injury. In the present study, we characterized the protective effect of alisol B 23-acetate (AB23A), a natural triterpenoid, on alpha-naphthylisothiocyanate (ANIT)-induced liver injury and intrahepatic cholestasis in mice and further elucidated the mechanisms in vivo and in vitro. AB23A treatment dose-dependently protected against liver injury induced by ANIT through reducing hepatic uptake and increasing efflux of bile acid via down-regulation of hepatic uptake transporters (Ntcp) and up-regulation of efflux transporter (Bsep, Mrp2 and Mdr2) expression. Furthermore, AB23A reduced bile acid synthesis through repressing Cyp7a1 and Cyp8b1, increased bile acid conjugation through inducing Bal, Baat and bile acid metabolism through an induction in gene expression of Sult2a1. We further demonstrate the involvement of farnesoid X receptor (FXR) in the hepatoprotective effect of AB23A. The changes in transporters and enzymes, as well as ameliorative liver histology in AB23A-treated mice were abrogated by FXR antagonist guggulsterone in vivo. In vitro evidences also directly demonstrated the effect of AB23A on FXR activation in a dose-dependent manner using luciferase reporter assay in HepG2 cells. In conclusion, AB23A produces protective effect against ANIT-induced hepatotoxity and cholestasis, due to FXR-mediated regulation of transporters and enzymes. - Highlights: • AB23A has at least three roles in protection against ANIT-induced liver injury. • AB23A decreases Ntcp, and increases Bsep, Mrp2 and Mdr2 expression. • AB23A represses Cyp7a1 and Cyp8b1 through inducing Shp and Fgf15 expression. • AB23A increases bile acid metabolism through inducing Sult2a1 expression. • FXR activation is involved

  13. The Nuclear Receptor FXR Uncouples the Actions of miR-33 from SREBP-2

    PubMed Central

    Tarling, Elizabeth J.; Ahn, Hannah; de Aguiar Vallim, Thomas Q.

    2015-01-01

    Objective To determined whether activation of FXR alters cellular and plasma cholesterol homeostasis as a result of regulation of Srebp-2 and miR-33. Approach and Results ChIP-seq data identified an FXR-response element within intron 10 of the Srebp-2 gene. Consistent with this observation, treatment of mice with FXR-specific agonists (GSK2324 or GW4064) rapidly increased hepatic levels of Srebp-2 mRNA, pSREBP-2 protein, and miR-33. Further, miR-33 targets, that include ABCA1, NSF and CPT1, were all reduced in GSK2324-treated mice. In contrast, neither nSREBP-2 protein, nor SREBP-2 target genes were induced following FXR activation. The inability to process pSREBP-2 to nSREBP-2 is likely a consequence of induction of INSIG-2a by FXR agonists. Finally, we show that FXR-dependent induction of both Srebp-2 and miR-33 is ablated in Scap−/− mice that lack nSREBP-2. Conclusions We demonstrate that activation of FXR uncouples the expression of nSREBP-2 and miR-33, and the regulation of their respective target genes. Further, we conclude that the FXR agonist-dependent increase in miR-33 requires transcription of the Srebp-2 gene. PMID:25593129

  14. FXR silencing in human colon cancer by DNA methylation and KRAS signaling.

    PubMed

    Bailey, Ann M; Zhan, Le; Maru, Dipen; Shureiqi, Imad; Pickering, Curtis R; Kiriakova, Galina; Izzo, Julie; He, Nan; Wei, Caimiao; Baladandayuthapani, Veerabhadran; Liang, Han; Kopetz, Scott; Powis, Garth; Guo, Grace L

    2014-01-01

    Farnesoid X receptor (FXR) is a bile acid nuclear receptor described through mouse knockout studies as a tumor suppressor for the development of colon adenocarcinomas. This study investigates the regulation of FXR in the development of human colon cancer. We used immunohistochemistry of FXR in normal tissue (n = 238), polyps (n = 32), and adenocarcinomas, staged I-IV (n = 43, 39, 68, and 9), of the colon; RT-quantitative PCR, reverse-phase protein array, and Western blot analysis in 15 colon cancer cell lines; NR1H4 promoter methylation and mRNA expression in colon cancer samples from The Cancer Genome Atlas; DNA methyltransferase inhibition; methyl-DNA immunoprecipitation (MeDIP); bisulfite sequencing; and V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) knockdown assessment to investigate FXR regulation in colon cancer development. Immunohistochemistry and quantitative RT-PCR revealed that expression and function of FXR was reduced in precancerous lesions and silenced in a majority of stage I-IV tumors. FXR expression negatively correlated with phosphatidylinositol-4, 5-bisphosphate 3 kinase signaling and the epithelial-to-mesenchymal transition. The NR1H4 promoter is methylated in ~12% colon cancer The Cancer Genome Atlas samples, and methylation patterns segregate with tumor subtypes. Inhibition of DNA methylation and KRAS silencing both increased FXR expression. FXR expression is decreased early in human colon cancer progression, and both DNA methylation and KRAS signaling may be contributing factors to FXR silencing. FXR potentially suppresses epithelial-to-mesenchymal transition and other oncogenic signaling cascades, and restoration of FXR activity, by blocking silencing mechanisms or increasing residual FXR activity, represents promising therapeutic options for the treatment of colon cancer.

  15. Obeticholic acid, a synthetic bile acid agonist of the farnesoid X receptor, attenuates experimental autoimmune encephalomyelitis

    PubMed Central

    Ho, Peggy P.; Steinman, Lawrence

    2016-01-01

    Bile acids are ligands for the nuclear hormone receptor, farnesoid X receptor (FXR). The bile acid–FXR interaction regulates bile acid synthesis, transport, and cholesterol metabolism. Recently, bile acid–FXR regulation has been reported to play an integral role in both hepatic and intestinal inflammation, and in atherosclerosis. In this study, we found that FXR knockout mice had more disease severity in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Obeticholic acid (6α-ethyl-chenodeoxycholic acid, 6-ECDCA), a synthetic FXR agonist, is an orally available drug that is currently in clinical trials for the treatment of inflammatory diseases such as alcoholic hepatitis, nonalcoholic steatohepatitis, and primary biliary cirrhosis. When we treated mice exhibiting established EAE with 6-ECDCA, or the natural FXR ligand chenodeoxycholic acid (CDCA), clinical disease was ameliorated by (i) suppressing lymphocyte activation and proinflammatory cytokine production; (ii) reducing CD4+ T cells and CD19+ B cell populations and their expression of negative checkpoint regulators programmed cell death protein 1 (PD1), programmed death-ligand 1 (PD-L1), and B and T lymphocyte attenuator (BTLA); (iii) increasing CD8+ T cells and PD1, PDl-1, and BTLA expression; and (iv) reducing VLA-4 expression in both the T- and B-cell populations. Moreover, adoptive transfer of 6-ECDCA– or CDCA-treated donor cells failed to transfer disease in naive recipients. Thus, we show that FXR functions as a negative regulator in neuroinflammation and we highlight that FXR agonists represent a potential previously unidentified therapy for MS. PMID:26811456

  16. Recent advances in non-steroidal FXR antagonists development for therapeutic applications.

    PubMed

    Huang, Huang; Xu, Yong; Zhu, Jin; Li, Jian

    2014-01-01

    Farnesoid X receptor (FXR, NR1H4), a nuclear receptor (NR) highly expressed in the liver, intestine, kidney, adrenal glands and other cholesterol-rich tissues, functions as the master regulator for bile acid homeostasis. FXR, which regulates the expression of genes encoding proteins involved in cholesterol homeostasis, plays an essential role in regulating cholesterol, lipid, and glucose metabolism. Recently, some FXR agonists are reported to have low selectivity on NRs, which forces the researchers to move their eyes onto the development of FXR antagonists with high selectivity. The development of non-steroidal FXR antagonists with different scaffolds including AGN34, tuberatolides, atractylenolides, andrographolides, GW4064 derivatives and 1,3,4-trisubstitutedpyrazolones, provides us a prospect for the therapy of in ammation, metabolic syndrome, diabetes, cholesterol gallstones, and cancer. PMID:25388534

  17. 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.

  18. Intestinal FXR agonism promotes adipose tissue browning and reduces obesity and insulin resistance

    PubMed Central

    Fang, Sungsoon; Suh, Jae Myoung; Reilly, Shannon M; Yu, Elizabeth; Osborn, Olivia; Lackey, Denise; Yoshihara, Eiji; Perino, Alessia; Jacinto, Sandra; Lukasheva, Yelizaveta; Atkins, Annette R; Khvat, Alexander; Schnabl, Bernd; Yu, Ruth T; Brenner, David A; Coulter, Sally; Liddle, Christopher; Schoonjans, Kristina; Olefsky, Jerrold M; Saltiel, Alan R; Downes, Michael; Evans, Ronald M

    2015-01-01

    The systemic expression of the bile acid (BA) sensor farnesoid X receptor (FXR) has led to promising new therapies targeting cholesterol metabolism, triglyceride production, hepatic steatosis and biliary cholestasis. In contrast to systemic therapy, bile acid release during a meal selectively activates intestinal FXR. By mimicking this tissue-selective effect, the gut-restricted FXR agonist fexaramine (Fex) robustly induces enteric fibroblast growth factor 15 (FGF15), leading to alterations in BA composition, but does so without activating FXR target genes in the liver. However, unlike systemic agonism, we find that Fex reduces diet-induced weight gain, body-wide inflammation and hepatic glucose production, while enhancing thermogenesis and browning of white adipose tissue (WAT). These pronounced metabolic improvements suggest tissue-restricted FXR activation as a new approach in the treatment of obesity and metabolic syndrome. PMID:25559344

  19. FXR Primes the Liver for Intestinal FGF15 Signaling by Transient Induction of β-Klotho.

    PubMed

    Fu, Ting; Kim, Young-Chae; Byun, Sangwon; Kim, Dong-Hyun; Seok, Sunmi; Suino-Powell, Kelly; Xu, H Eric; Kemper, Byron; Kemper, Jongsook Kim

    2016-01-01

    The bile acid (BA)-sensing nuclear receptor, farnesoid X receptor (FXR), regulates postprandial metabolic responses, including inhibition of BA synthesis, by inducing the intestinal hormone, fibroblast growth factor (FGF)15 (FGF19 in human). In this study, we tested a novel hypothesis that FXR not only induces intestinal FGF15 but also primes the liver for effectively responding to the signal by transcriptional induction of the obligate coreceptor for FGF15, β-Klotho (βKL). Activation of FXR by a synthetic agonist, GW4064, in mice increased occupancy of FXR and its DNA-binding partner, retinoid X receptor-α, at FGF15-signaling component genes, particularly βKL, and induced expression of these genes. Interestingly, mRNA levels of Fgfr4, the FGF15 receptor, were not increased by GW4064, but protein levels increased as a result of βKL-dependent increased protein stability. Both FGF receptor 4 and βKL protein levels were substantially decreased in FXR-knockout (KO) mice, and FGF19 signaling, monitored by phosphorylated ERK, was blunted in FXR-KO mice, FXR-KO mouse hepatocytes, and FXR-down-regulated human hepatocytes. Overexpression of βKL in FXR-lacking hepatocytes partially restored FGF19 signaling and inhibition by FGF19 of Cyp7a1, which encodes the rate-limiting BA biosynthetic enzyme. In mice, transient inductions of intestinal Fgf15 and hepatic βKL were temporally correlated after GW4064 treatment, and pretreatment of hepatocytes with GW4064 before FGF19 treatment enhanced FGF19 signaling, which was abolished by transcriptional inhibition or βKL down-regulation. This study identifies FXR as a gut-liver metabolic coordinator for FGF15/19 action that orchestrates transient induction of hepatic βKL and intestinal Fgf15/19 in a temporally correlated manner.

  20. Conformational dynamics of human FXR-LBD ligand interactions studied by hydrogen/deuterium exchange mass spectrometry: insights into the antagonism of the hypolipidemic agent Z-guggulsterone.

    PubMed

    Yang, Liping; Broderick, David; Jiang, Yuan; Hsu, Victor; Maier, Claudia S

    2014-09-01

    Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily of transcription factors that plays a key role in the regulation of bile acids, lipid and glucose metabolisms. The regulative function of FXR is governed by conformational changes of the ligand binding domain (LBD) upon ligand binding. Although FXR is a highly researched potential therapeutic target, only a limited number of FXR-agonist complexes have been successfully crystallized and subsequently yielded high resolution structures. There is currently no structural information of any FXR-antagonist complexes publically available. We therefore explored the use of amide hydrogen/deuterium exchange (HDX) coupled with mass spectrometry for characterizing conformational changes in the FXR-LBD upon ligand binding. Ligand-specific deuterium incorporation profiles were obtained for three FXR ligand chemotypes: GW4064, a synthetic non-steroidal high affinity agonist; the bile acid chenodeoxycholic acid (CDCA), the endogenous low affinity agonist of FXR; and Z-guggulsterone (GG), an in vitro antagonist of the steroid chemotype. A comparison of the HDX profiles of their ligand-bound FXR-LBD complexes revealed a unique mode of interaction for GG. The conformational features of the FXR-LBD-antagonist interaction are discussed.

  1. Discovery That Theonellasterol a Marine Sponge Sterol Is a Highly Selective FXR Antagonist That Protects against Liver Injury in Cholestasis

    PubMed Central

    Renga, Barbara; Mencarelli, Andrea; D'Amore, Claudio; Cipriani, Sabrina; D'Auria, Maria Valeria; Sepe, Valentina; Chini, Maria Giovanna; Monti, Maria Chiara; Bifulco, Giuseppe; Zampella, Angela; Fiorucci, Stefano

    2012-01-01

    Background The farnesoid-x-receptor (FXR) is a bile acid sensor expressed in the liver and gastrointestinal tract. Despite FXR ligands are under investigation for treatment of cholestasis, a biochemical condition occurring in a number of liver diseases for which available therapies are poorly effective, mice harboring a disrupted FXR are protected against liver injury caused by bile acid overload in rodent models of cholestasis. Theonellasterol is a 4-methylene-24-ethylsteroid isolated from the marine sponge Theonella swinhoei. Here, we have characterized the activity of this theonellasterol on FXR-regulated genes and biological functions. Principal Findings Interrogation of HepG2 cells, a human hepatocyte cell line, by microarray analysis and transactivation assay shows that theonellasterol is a selective FXR antagonist, devoid of any agonistic or antagonistic activity on a number of human nuclear receptors including the vitamin D receptor, PPARs, PXR, LXRs, progesterone, estrogen, glucorticoid and thyroid receptors, among others. Exposure of HepG2 cells to theonellasterol antagonizes the effect of natural and synthetic FXR agonists on FXR-regulated genes, including SHP, OSTα, BSEP and MRP4. A proof-of-concept study carried out to investigate whether FXR antagonism rescues mice from liver injury caused by the ligation of the common bile duct, a model of obstructive cholestasis, demonstrated that theonellasterol attenuates injury caused by bile duct ligation as measured by assessing serum alanine aminostrasferase levels and extent of liver necrosis at histopathology. Analysis of genes involved in bile acid uptake and excretion by hepatocytes revealed that theonellasterol increases the liver expression of MRP4, a basolateral transporter that is negatively regulated by FXR. Administering bile duct ligated mice with an FXR agonist failed to rescue from liver injury and downregulated the expression of MRP4. Conclusions FXR antagonism in vivo results in a positive

  2. The farnesoid X receptor induces very low density lipoprotein receptor gene expression.

    PubMed

    Sirvent, Audrey; Claudel, Thierry; Martin, Geneviève; Brozek, John; Kosykh, Vladimir; Darteil, Raphaël; Hum, Dean W; Fruchart, Jean-Charles; Staels, Bart

    2004-05-21

    The farnesoid X receptor (FXR) is a nuclear receptor activated by bile acids (BAs). In response to ligand-binding, FXR regulates many genes involved in BA, lipid, and lipoprotein metabolism. To identify new FXR target genes, microarray technology was used to profile total RNA extracted from HepG2 cells treated with the natural FXR agonist chenodeoxycholic acid (CDCA). Interestingly, a significant increase of transcript level of the very low density lipoprotein receptor (VLDLR) was observed. Our data, resulting from selective FXR activation, FXR RNA silencing and FXR-deficient mice, clearly demonstrate that BAs up-regulate VLDLR transcript levels via a FXR-dependent mechanism in vitro in human and in vivo in mouse liver cells.

  3. 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.

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

    PubMed Central

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

    2015-01-01

    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. PMID:26567894

  5. Role of FXR in β-cells of lean and obese mice.

    PubMed

    Schittenhelm, Björn; Wagner, Rebecca; Kähny, Verena; Peter, Andreas; Krippeit-Drews, Peter; Düfer, Martina; Drews, Gisela

    2015-04-01

    We have recently shown that the bile acid (BA) taurochenodeoxycholate (TCDC) acutely stimulates insulin secretion via activation of the farnesoid X receptor (FXR). Aims of the current investigation were to discriminate between nongenomic (≤1 h) and genomic effects (24-48 h) of BAs on β-cells and to evaluate whether FXR can modulate the adverse effects of a high-fat diet (HFD). TCDC (500 nM) as well as glycine-conjugated and unconjugated CDC (chenodeoxycholate) increased insulin secretion in acute incubations but did not evoke additional effects after 1-2 days of preincubation. The BAs did not stimulate β-cells of FXR-knockout (KO) mice and activation of the G protein-coupled BA receptor TGR5 was ineffective, suggesting that FXR is the sole BA receptor in β-cells activated by TCDC and its analogues. As opposed to lean mice, obese FXR-KO mice did not show HFD-induced glucose intolerance and increased fasting glucose. The beneficial impact of FXR-KO on glucose metabolism cannot be explained by an adaptive compensation of insulin secretion or β-cell mass. Interestingly, in contrast to its effect on islets from lean mice, the FXR agonist GW4064 was ineffective in stimulating insulin secretion of islets from wild type mice fed a HFD or isolated islets kept in a glucolipotoxic medium. Additional feeding of CDC restored the effect of GW4064. CDC prevented HFD-induced impairment of glucose tolerance and in vitro effects of glucolipotoxicity. The data show that the FXR is the most important BA receptor in β-cells and that FXR signaling in β-cells is impaired by overnutrition, which alters activatability of the FXR.

  6. 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

  7. 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.

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

    PubMed

    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(50), 1μM) 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.

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

    PubMed Central

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

    2011-01-01

    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 (IC50, 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. PMID:22178739

  11. Ageing Fxr deficient mice develop increased energy expenditure, improved glucose control and liver damage resembling NASH.

    PubMed

    Bjursell, Mikael; Wedin, Marianne; Admyre, Therése; Hermansson, Majlis; Böttcher, Gerhard; Göransson, Melker; Lindén, Daniel; Bamberg, Krister; Oscarsson, Jan; Bohlooly-Y, Mohammad

    2013-01-01

    Nuclear receptor subfamily 1, group H, member 4 (Nr1h4, FXR) is a bile acid activated nuclear receptor mainly expressed in the liver, intestine, kidney and adrenal glands. Upon activation, the primary function is to suppress cholesterol 7 alpha-hydroxylase (Cyp7a1), the rate-limiting enzyme in the classic or neutral bile acid synthesis pathway. In the present study, a novel Fxr deficient mouse line was created and studied with respect to metabolism and liver function in ageing mice fed chow diet. The Fxr deficient mice were similar to wild type mice in terms of body weight, body composition, energy intake and expenditure as well as behaviours at a young age. However, from 15 weeks of age and onwards, the Fxr deficient mice had almost no body weight increase up to 39 weeks of age mainly because of lower body fat mass. The lower body weight gain was associated with increased energy expenditure that was not compensated by increased food intake. Fasting levels of glucose and insulin were lower and glucose tolerance was improved in old and lean Fxr deficient mice. However, the Fxr deficient mice displayed significantly increased liver weight, steatosis, hepatocyte ballooning degeneration and lobular inflammation together with elevated plasma levels of ALT, bilirubin and bile acids, findings compatible with non-alcoholic steatohepatitis (NASH) and cholestasis. In conclusion, ageing Fxr deficient mice display late onset leanness associated with elevated energy expenditure and improved glucose control but develop severe NASH-like liver pathology.

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

    NASA Astrophysics Data System (ADS)

    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.

    2006-03-01

    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. bacteria | biliary obstruction | epithelial barrier | ileum

  13. 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.

  14. Genome-wide profiling to analyze the effects of FXR activation on mouse renal proximal tubular cells

    PubMed Central

    Gui, Ting; Gai, Zhibo

    2015-01-01

    To assess the effect of farnesoid X receptor (FXR), a bile acid nuclear receptor, on renal proximal tubular cells, primary cultured mouse kidney proximal tubular cells were treated with GW4064 (a FXR agonist) or DMSO (as controls) overnight. Analysis of gene expression in the proximal tubular cells by whole genome microarrays indicated that FXR activation induced genes involved in fatty acid degradation and oxidation reduction. Among them, genes involved in glutathione metabolism were mostly induced. Here we describe in details the contents and quality controls for the gene expression and related results associated with the data uploaded to Gene Expression Omnibus (accession number GSE70296). PMID:26697325

  15. Pyrazinamide Induced Rat Cholestatic Liver Injury through Inhibition of FXR Regulatory Effect on Bile Acid Synthesis and Transport.

    PubMed

    Guo, Hong-Li; Hassan, Hozeifa M; Zhang, Yun; Dong, Si-Zhe; Ding, Ping-Ping; Wang, Tao; Sun, Li-Xin; Zhang, Lu-Yong; Jiang, Zhen-Zhou

    2016-08-01

    Pyrazinamide (PZA) is an indispensable first-line drug used for the treatment of tuberculosis which may cause serious hepatotoxicity; however, the mechanisms underlying these toxicities are poorly understood. Cholestasis plays an important role in drug-induced liver injury. Since there were no previous published works reported cholestasis and PZA hepatotoxicity relationship, this study aimed to identify whether PZA can induce liver injury with characterized evidences of cholestasis and to clarify expression changes of proteins related to both bile acid synthesis and transport in PZA-induced liver injury. PZA (2 g/kg) was administered for 7 consecutive days by oral gavage. Results showed there were 2-fold elevation in both ALT and AST serum levels in PZA-treated rats. In addition, a 10-fold increment in serum total bile acid was observed after PZA administration. The mRNA and protein expressions of bile acid synthesis and transport parameters were markedly altered, in which FXR, Bsep, Mrp2, Mdr2, Ostα/β, Oatp1a1, Oatp1b2, and Cyp8b1 were decreased (P < .05), while Mrp3, Ntcp, Oatp1a4, and Cyp7a1 were increased (P < .05). Moreover, treatment with the FXR agonist obeticholic acid (OCA) generated obvious reductions in serum ALT, AST, and TBA levels in PZA-treated rats. Those effects were due to transcriptional regulation of pre-mentioned target genes by OCA. Taken together, these results suggested that PZA-induced cholestatic liver injury was related to FXR inhibition, leading to the dysfunction in bile acid synthesis and transport. PMID:27255380

  16. Identification of a potent synthetic FXR agonist with an unexpected mode of binding and activation

    SciTech Connect

    Soisson, Stephen M.; Parthasarathy, Gopalakrishnan; Adams, Alan D.; Sahoo, Soumya; Sitlani, Ayesha; Sparrow, Carl; Cui, Jisong; Becker, Joseph W.

    2008-07-08

    The farnesoid X receptor (FXR), a member of the nuclear hormone receptor family, plays important roles in the regulation of bile acid and cholesterol homeostasis, glucose metabolism, and insulin sensitivity. There is intense interest in understanding the mechanisms of FXR regulation and in developing pharmaceutically suitable synthetic FXR ligands that might be used to treat metabolic syndrome. We report here the identification of a potent FXR agonist (MFA-1) and the elucidation of the structure of this ligand in ternary complex with the human receptor and a coactivator peptide fragment using x-ray crystallography at 1.9-{angstrom} resolution. The steroid ring system of MFA-1 binds with its D ring-facing helix 12 (AF-2) in a manner reminiscent of hormone binding to classical steroid hormone receptors and the reverse of the pose adopted by naturally occurring bile acids when bound to FXR. This binding mode appears to be driven by the presence of a carboxylate on MFA-1 that is situated to make a salt-bridge interaction with an arginine residue in the FXR-binding pocket that is normally used to neutralize bound bile acids. Receptor activation by MFA-1 differs from that by bile acids in that it relies on direct interactions between the ligand and residues in helices 11 and 12 and only indirectly involves a protonated histidine that is part of the activation trigger. The structure of the FXR:MFA-1 complex differs significantly from that of the complex with a structurally distinct agonist, fexaramine, highlighting the inherent plasticity of the receptor.

  17. Atorvastatin induces bile acid-synthetic enzyme Cyp7a1 by suppressing FXR signaling in both liver and intestine in mice.

    PubMed

    Fu, Zidong Donna; Cui, Julia Yue; Klaassen, Curtis D

    2014-12-01

    Statins are effective cholesterol-lowering drugs to treat CVDs. Bile acids (BAs), the end products of cholesterol metabolism in the liver, are important nutrient and energy regulators. The present study aims to investigate how statins affect BA homeostasis in the enterohepatic circulation. Male C57BL/6 mice were treated with atorvastatin (100 mg/kg/day po) for 1 week, followed by BA profiling by ultra-performance LC-MS/MS. Atorvastatin decreased BA pool size, mainly due to less BA in the intestine. Surprisingly, atorvastatin did not alter total BAs in the serum or liver. Atorvastatin increased the ratio of 12α-OH/non12α-OH BAs. Atorvastatin increased the mRNAs of the BA-synthetic enzymes cholesterol 7α-hydroxylase (Cyp7a1) (over 10-fold) and cytochrome P450 27a1, the BA uptake transporters Na⁺/taurocholate cotransporting polypeptide and organic anion transporting polypeptide 1b2, and the efflux transporter multidrug resistance-associated protein 2 in the liver. Noticeably, atorvastatin suppressed the expression of BA nuclear receptor farnesoid X receptor (FXR) target genes, namely small heterodimer partner (liver) and fibroblast growth factor 15 (ileum). Furthermore, atorvastatin increased the mRNAs of the organic cation uptake transporter 1 and cholesterol efflux transporters Abcg5 and Abcg8 in the liver. The increased expression of BA-synthetic enzymes and BA transporters appear to be a compensatory response to maintain BA homeostasis after atorvastatin treatment. The Cyp7a1 induction by atorvastatin appears to be due to suppressed FXR signaling in both the liver and intestine.

  18. Atorvastatin induces bile acid-synthetic enzyme Cyp7a1 by suppressing FXR signaling in both liver and intestine in mice[S

    PubMed Central

    Fu, Zidong Donna; Cui, Julia Yue; Klaassen, Curtis D.

    2014-01-01

    Statins are effective cholesterol-lowering drugs to treat CVDs. Bile acids (BAs), the end products of cholesterol metabolism in the liver, are important nutrient and energy regulators. The present study aims to investigate how statins affect BA homeostasis in the enterohepatic circulation. Male C57BL/6 mice were treated with atorvastatin (100 mg/kg/day po) for 1 week, followed by BA profiling by ultra-performance LC-MS/MS. Atorvastatin decreased BA pool size, mainly due to less BA in the intestine. Surprisingly, atorvastatin did not alter total BAs in the serum or liver. Atorvastatin increased the ratio of 12α-OH/non12α-OH BAs. Atorvastatin increased the mRNAs of the BA-synthetic enzymes cholesterol 7α-hydroxylase (Cyp7a1) (over 10-fold) and cytochrome P450 27a1, the BA uptake transporters Na+/taurocholate cotransporting polypeptide and organic anion transporting polypeptide 1b2, and the efflux transporter multidrug resistance-associated protein 2 in the liver. Noticeably, atorvastatin suppressed the expression of BA nuclear receptor farnesoid X receptor (FXR) target genes, namely small heterodimer partner (liver) and fibroblast growth factor 15 (ileum). Furthermore, atorvastatin increased the mRNAs of the organic cation uptake transporter 1 and cholesterol efflux transporters Abcg5 and Abcg8 in the liver. The increased expression of BA-synthetic enzymes and BA transporters appear to be a compensatory response to maintain BA homeostasis after atorvastatin treatment. The Cyp7a1 induction by atorvastatin appears to be due to suppressed FXR signaling in both the liver and intestine. PMID:25278499

  19. VPAC1 expression is regulated by FXR agonists in the human gallbladder epithelium.

    PubMed

    Chignard, Nicolas; Mergey, Martine; Barbu, Véronique; Finzi, Laetitia; Tiret, Emmanuel; Paul, Annick; Housset, Chantal

    2005-09-01

    Vasoactive intestinal peptide receptor-1 (VPAC1) is the high-affinity receptor of vasoactive intestinal peptide (VIP), a major regulator of bile secretion. To better define the level at which VPAC1 stimulates bile secretion, we examined its expression in the different cell types participating in bile formation (i.e., hepatocytes, bile duct, and gallbladder epithelial cells). Because VPAC1 expression was previously shown to be regulated by nuclear receptors, we tested the hypothesis that it may be regulated by the farnesoid X receptor (FXR). Quantitative RT-PCR and immunoblot analyses of cell isolates indicated that VPAC1 is expressed in all three cell types lining the human biliary tree, with predominant expression in the gallbladder. In primary cultures of human gallbladder epithelial cells, VIP induced cAMP production and chloride secretion. Analysis of the VPAC1 gene revealed the presence of potential FXR response element sequences, and both FXR and RXRalpha expressions were detected in gallbladder epithelial cells. In these cells, the FXR pharmacological agonist GW4064 upregulated VPAC1 expression in a dose-dependent manner, and this effect was antagonized by the RXRalpha ligand, 9-cis retinoic acid. Chenodeoxycholate activated endogenous FXR in gallbladder epithelial cells, as ascertained by electromobility shift assay and upregulation of the FXR target gene, small heterodimer partner. Chenodeoxycholate also provoked an increase in VPAC1 mRNA and protein content in these cells. In conclusion, FXR agonists may increase gallbladder fluid secretion through transcriptional activation of VPAC1, which may contribute to the regulation of bile secretion by bile salts and to a protective effect of FXR pharmacological agonists in gallstone disease.

  20. Phosphorylation of farnesoid X receptor by protein kinase C promotes its transcriptional activity.

    PubMed

    Gineste, Romain; Sirvent, Audrey; Paumelle, Réjane; Helleboid, Stéphane; Aquilina, Alexis; Darteil, Raphaël; Hum, Dean W; Fruchart, Jean-Charles; Staels, Bart

    2008-11-01

    The farnesoid X receptor (FXR, NR1H4) belongs to the nuclear receptor superfamily and is activated by bile acids such as chenodeoxycholic acid, or synthetic ligands such as GW4064. FXR is implicated in the regulation of bile acid, lipid, and carbohydrate metabolism. Posttranslational modifications regulating its activity have not been investigated yet. Here, we demonstrate that calcium-dependent protein kinase C (PKC) inhibition impairs ligand-mediated regulation of FXR target genes. Moreover, in a transactivation assay, we show that FXR transcriptional activity is modulated by PKC. Furthermore, phorbol 12-myristate 13-acetate , a PKC activator, induces the phosphorylation of endogenous FXR in HepG2 cells and PKCalpha phosphorylates in vitro FXR in its DNA-binding domain on S135 and S154. Mutation of S135 and S154 to alanine residues reduces in cell FXR phosphorylation. In contrast to wild-type FXR, mutant FXRS135AS154A displays an impaired PKCalpha-induced transactivation and a decreased ligand-dependent FXR transactivation. Finally, phosphorylation of FXR by PKC promotes the recruitment of peroxisomal proliferator-activated receptor gamma coactivator 1alpha. In conclusion, these findings show that the phosphorylation of FXR induced by PKCalpha directly modulates the ability of agonists to activate FXR.

  1. Statins and transcriptional regulation: The FXR connection

    SciTech Connect

    Habeos, Ioannis; Ziros, Panos G.; Psyrogiannis, Agathoklis; Vagenakis, Apostolos G.; Papavassiliou, Athanasios G. . E-mail: papavas@med.upatras.gr

    2005-08-26

    Farnesoid X receptor (FXR) is a nuclear receptor involved in lipoprotein as well as glucose metabolism. Statins are widely used hypolipidemic agents with many pleiotropic actions. It is known that statins affect other nuclear hormone receptors, but no reports are available on the effect of these drugs on FXR. Employing an animal model (Syrian hamsters), we hereby present evidence to demonstrate that Simvastatin, a broadly prescribed statin, decreases the expression of FXR at both the RNA and protein levels and down-regulates its DNA-binding activity. This novel property may have important implications on the mode statins influence on lipoprotein and carbohydrate homeostasis in the organism.

  2. Role of farnesoid X receptor and bile acids in alcoholic liver disease

    PubMed Central

    Manley, Sharon; Ding, Wenxing

    2015-01-01

    Alcoholic liver disease (ALD) is one of the major causes of liver morbidity and mortality worldwide. Chronic alcohol consumption leads to development of liver pathogenesis encompassing steatosis, inflammation, fibrosis, cirrhosis, and in extreme cases, hepatocellular carcinoma. Moreover, ALD may also associate with cholestasis. Emerging evidence now suggests that farnesoid X receptor (FXR) and bile acids also play important roles in ALD. In this review, we discuss the effects of alcohol consumption on FXR, bile acids and gut microbiome as well as their impacts on ALD. Moreover, we summarize the findings on FXR, FoxO3a (forkhead box-containing protein class O3a) and PPARα (peroxisome proliferator-activated receptor alpha) in regulation of autophagy-related gene transcription program and liver injury in response to alcohol exposure. PMID:26579442

  3. Role of farnesoid X receptor and bile acids in alcoholic liver disease.

    PubMed

    Manley, Sharon; Ding, Wenxing

    2015-03-01

    Alcoholic liver disease (ALD) is one of the major causes of liver morbidity and mortality worldwide. Chronic alcohol consumption leads to development of liver pathogenesis encompassing steatosis, inflammation, fibrosis, cirrhosis, and in extreme cases, hepatocellular carcinoma. Moreover, ALD may also associate with cholestasis. Emerging evidence now suggests that farnesoid X receptor (FXR) and bile acids also play important roles in ALD. In this review, we discuss the effects of alcohol consumption on FXR, bile acids and gut microbiome as well as their impacts on ALD. Moreover, we summarize the findings on FXR, FoxO3a (forkhead box-containing protein class O3a) and PPARα (peroxisome proliferator-activated receptor alpha) in regulation of autophagy-related gene transcription program and liver injury in response to alcohol exposure. PMID:26579442

  4. Inhibitory effects of bile acids and synthetic farnesoid X receptor agonists on rotavirus replication.

    PubMed

    Kim, Yunjeong; Chang, Kyeong-Ok

    2011-12-01

    Rotaviruses (group A rotaviruses) are the most important cause of severe gastroenteritis in infants and children worldwide. Currently, an antiviral drug is not available and information on therapeutic targets for antiviral development is limited for rotavirus infection. Previously, it was shown that lipid homeostasis is important in rotavirus replication. Since farnesoid X receptor (FXR) and its natural ligands bile acids (such as chenodeoxycholic acid [CDCA]) play major roles in cholesterol and lipid homeostasis, we examined the effects of bile acids and synthetic FXR agonists on rotavirus replication in association with cellular lipid levels. In a mouse model of rotavirus infection, effects of oral administration of CDCA on fecal rotavirus shedding were investigated. The results demonstrate the following. First, the intracellular contents of triglycerides were significantly increased by rotavirus infection. Second, CDCA, deoxycholic acid (DCA), and other synthetic FXR agonists, such as GW4064, significantly reduced rotavirus replication in cell culture in a dose-dependent manner. The reduction of virus replication correlated positively with activation of the FXR pathway and reduction of cellular triglyceride contents (r(2) = 0.95). Third, oral administration of CDCA significantly reduced fecal virus shedding in mice (P < 0.05). We conclude that bile acids and FXR agonists play important roles in the suppression of rotavirus replication. The inhibition mechanism is proposed to be the downregulation of lipid synthesis induced by rotavirus infection.

  5. NSAIDs Ibuprofen, Indometacin, and Diclofenac do not interact with Farnesoid X Receptor.

    PubMed

    Schmidt, Jurema; Klingler, Franca-Maria; Proschak, Ewgenji; Steinhilber, Dieter; Schubert-Zsilavecz, Manfred; Merk, Daniel

    2015-01-01

    The nuclear farnesoid X receptor (FXR) is a ligand activated transcription factor and acts as cellular sensor for bile acids. In this role, FXR is a highly important liver protector and FXR inhibition by antagonists or knockout has shown several deleterious effects. A recent report characterized non-steroidal anti-rheumatic drugs (NSAIDs) such as ibuprofen or diclofenac as FXR antagonists and linked hepatotoxic effects of these drugs with antagonistic activity on FXR. Since this would guide a way to develop safer anti-inflammatory agents by sparing FXR, we intended to further characterize the reported antagonistic activity and intensively investigated ibuprofen, indometacin and diclofenac. However, we conclude that these agents do not interact with FXR and that the reported reduced FXR signaling induced by CDCA in presence of NSAIDs is merely a consequence than a cause of hepatotoxicity.

  6. NSAIDs Ibuprofen, Indometacin, and Diclofenac do not interact with Farnesoid X Receptor

    PubMed Central

    Schmidt, Jurema; Klingler, Franca-Maria; Proschak, Ewgenji; Steinhilber, Dieter; Schubert-Zsilavecz, Manfred; Merk, Daniel

    2015-01-01

    The nuclear farnesoid X receptor (FXR) is a ligand activated transcription factor and acts as cellular sensor for bile acids. In this role, FXR is a highly important liver protector and FXR inhibition by antagonists or knockout has shown several deleterious effects. A recent report characterized non-steroidal anti-rheumatic drugs (NSAIDs) such as ibuprofen or diclofenac as FXR antagonists and linked hepatotoxic effects of these drugs with antagonistic activity on FXR. Since this would guide a way to develop safer anti-inflammatory agents by sparing FXR, we intended to further characterize the reported antagonistic activity and intensively investigated ibuprofen, indometacin and diclofenac. However, we conclude that these agents do not interact with FXR and that the reported reduced FXR signaling induced by CDCA in presence of NSAIDs is merely a consequence than a cause of hepatotoxicity. PMID:26424593

  7. 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.

  8. Farnesoid X receptor represses hepatic lipase gene expression.

    PubMed

    Sirvent, Audrey; Verhoeven, Adrie J M; Jansen, Hans; Kosykh, Vladimir; Darteil, Raphaël J; Hum, Dean W; Fruchart, Jean-Charles; Staels, Bart

    2004-11-01

    The farnesoid X receptor (FXR) is a nuclear receptor that regulates gene expression in response to bile acids (BAs). FXR plays a central role in BA, cholesterol, and lipoprotein metabolism. Here, we identify HL, an enzyme involved in the metabolism of remnant and high density lipoproteins, as a novel FXR-regulated gene. The natural FXR ligand, chenodeoxycholic acid (CDCA), downregulates HL gene expression in a dose- and time-dependent manner in human hepatoma HepG2 cells. The nonsteroidal synthetic FXR agonist GW4064 also decreases HL mRNA levels in HepG2 cells and in primary human hepatocytes. Moreover, the decrease of HL mRNA levels after treatment with FXR agonists was associated with a significant decrease in secreted enzymatic activity. In addition, FXR-specific gene silencing using small interfering RNAs demonstrated that CDCA- and GW4064-mediated downregulation of HL transcript levels occurs via an FXR-dependent mechanism. Finally, using transient transfection experiments, it is shown that FXR represses transcriptional activity of a reporter driven by the -698/+13 bp human HL promoter. Taken together, these results identify HL as a new FXR-regulated gene in human liver cells. In view of the role of HL in plasma lipoprotein metabolism, our results further emphasize the central role of FXR in lipid homeostasis.

  9. Chronic Activation of FXR in Transgenic Mice Caused Perinatal Toxicity and Sensitized Mice to Cholesterol Toxicity

    PubMed Central

    Cheng, Qiuqiong; Inaba, Yuka; Lu, Peipei; Xu, Meishu; He, Jinhan; Zhao, Yueshui; Guo, Grace L.; Kuruba, Ramalinga; de la Vega, Rona; Evans, Rhobert W.; Li, Song

    2015-01-01

    The nuclear receptor farnesoid X receptor (FXR) (nuclear receptor subfamily 1, group H, member 4, or NR1H4) is highly expressed in the liver and intestine. Previous reports have suggested beneficial functions of FXR in the homeostasis of bile acids, lipids, and glucose, as well as in promoting liver regeneration and inhibiting carcinogenesis. To investigate the effect of chronic FXR activation in vivo, we generated transgenic mice that conditionally and tissue specifically express the activated form of FXR in the liver and intestine. Unexpectedly, the transgenic mice showed several intriguing phenotypes, including partial neonatal lethality, growth retardation, and spontaneous liver toxicity. The transgenic mice also displayed heightened sensitivity to a high-cholesterol diet-induced hepatotoxicity but resistance to the gallstone formation. The phenotypes were transgene specific, because they were abolished upon treatment with doxycycline to silence the transgene expression. The perinatal toxicity, which can be rescued by a maternal vitamin supplement, may have resulted from vitamin deficiency due to low biliary bile acid output as a consequence of inhibition of bile acid formation. Our results also suggested that the fibroblast growth factor-inducible immediate-early response protein 14 (Fn14), a member of the proinflammatory TNF family, is a FXR-responsive gene. However, the contribution of Fn14 induction in the perinatal toxic phenotype of the transgenic mice remains to be defined. Because FXR is being explored as a therapeutic target, our results suggested that a chronic activation of this nuclear receptor may have an unintended side effect especially during the perinatal stage. PMID:25719402

  10. 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.

  11. 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

  12. In vivo imaging of farnesoid X receptor activity reveals the ileum as the primary bile acid signaling tissue.

    PubMed

    Houten, Sander M; Volle, David H; Cummins, Carolyn L; Mangelsdorf, David J; Auwerx, Johan

    2007-06-01

    We generated and characterized a firefly luciferase reporter mouse for the nuclear receptor farnesoid X receptor (FXR). This FXR reporter mouse has basal luciferase expression in the terminal ileum, an organ with well-characterized FXRalpha signaling. In vivo luciferase activity reflected the diurnal activity pattern of the mouse, and is regulated by both natural (bile acids, chenodeoxycholic acid) and synthetic (GW4064) FXRalpha ligands. Moreover, in vivo and in vitro analysis showed luciferase activity after GW4064 administration in the liver, kidney, and adrenal gland, indicating that FXRalpha signaling is functional in these tissues. Hepatic luciferase activity was robustly induced in cholestatic mice, showing that FXRalpha signaling pathways are activated in this disease. In conclusion, we have developed an FXR reporter mouse that is useful to monitor FXRalpha signaling in vivo in health and disease. The use of this animal could facilitate the development of new therapeutic compounds that target FXRalpha in a tissue-specific manner.

  13. Farnesoid X Receptor Critically Determines the Fibrotic Response in Mice but Is Expressed to a Low Extent in Human Hepatic Stellate Cells and Periductal Myofibroblasts

    PubMed Central

    Fickert, Peter; Fuchsbichler, Andrea; Moustafa, Tarek; Wagner, Martin; Zollner, Gernot; Halilbasic, Emina; Stöger, Ulrike; Arrese, Marco; Pizarro, Margarita; Solís, Nancy; Carrasco, Gonzalo; Caligiuri, Alessandra; Sombetzki, Martina; Reisinger, Emil; Tsybrovskyy, Oleksiy; Zatloukal, Kurt; Denk, Helmut; Jaeschke, Hartmut; Pinzani, Massimo; Trauner, Michael

    2009-01-01

    The nuclear bile acid receptor, farnesoid X receptor (FXR), may play a pivotal role in liver fibrosis. We tested the impact of genetic FXR ablation in four different mouse models. Hepatic fibrosis was induced in wild-type and FXR knock-out mice (FXR−/−) by CCl4 intoxication, 3,5-diethoxycarbonyl-1,4-dihydrocollidine feeding, common bile duct ligation, or Schistosoma mansoni (S.m.)-infection. In addition, we determined nuclear receptor expression levels (FXR, pregnane X receptor (PXR), vitamin D receptor, constitutive androstane receptor (CAR), small heterodimer partner (SHP)) in mouse hepatic stellate cells (HSCs), portal myofibroblasts (MFBs), and human HSCs. Cell type-specific FXR protein expression was determined by immunohistochemistry in five mouse models and prototypic human fibrotic liver diseases. Expression of nuclear receptors was much lower in mouse and human HSCs/MFBs compared with total liver expression with the exception of vitamin D receptor. FXR protein was undetectable in mouse and human HSCs and MFBs. FXR loss had no effect in CCl4-intoxicated and S.m.-infected mice, but significantly decreased liver fibrosis of the biliary type (common bile duct ligation, 3,5-diethoxycarbonyl-1,4-dihydrocollidine). These data suggest that FXR loss significantly reduces fibrosis of the biliary type, but has no impact on non-cholestatic liver fibrosis. Since there is no FXR expression in HSCs and MFBs in liver fibrosis, our data indicate that these cells may not represent direct therapeutic targets for FXR ligands. PMID:19910507

  14. 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.

  15. Farnesoid X Receptor Agonists and Other Bile Acid Signaling Strategies for Treatment of Liver Disease.

    PubMed

    Halilbasic, Emina; Fuchs, Claudia; Traussnigg, Stefan; Trauner, Michael

    2016-01-01

    The intracellular nuclear receptor farnesoid X receptor (FXR) and the transmembrane G protein-coupled receptor 5 (TGR5) respond to bile acids (BAs) by activating transcriptional networks and/or signaling cascades. These cascades affect the expression of a great number of target genes relevant for BA, cholesterol, lipid and carbohydrate metabolism, as well as genes involved in inflammation, fibrosis and carcinogenesis. FXR activation in the liver tissue and beyond, such as the gut-liver axis, kidney and adipose tissue, plays a role in metabolic diseases. These BA receptors activators hold promise to become a new class of drugs to be used in the treatment of chronic liver disease, hepatocellular cancer and extrahepatic inflammatory and metabolic diseases. This review discusses the relevant BA receptors, the new drugs that target BA transport and signaling and their possible applications. PMID:27332721

  16. 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.

  17. 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.

  18. Pharmacophore-based discovery of FXR-agonists. Part II: Identification of bioactive triterpenes from Ganoderma lucidum

    PubMed Central

    Grienke, Ulrike; Mihály-Bison, Judit; Schuster, Daniela; Afonyushkin, Taras; Binder, Markus; Guan, Shu-hong; Cheng, Chun-ru; Wolber, Gerhard; Stuppner, Hermann; Guo, De-an; Bochkov, Valery N.; Rollinger, Judith M.

    2011-01-01

    The farnesoid X receptor (FXR) belonging to the metabolic subfamily of nuclear receptors is a ligand-induced transcriptional activator. Its central function is the physiological maintenance of bile acid homeostasis including the regulation of glucose and lipid metabolism. Accessible structural information about its ligand-binding domain renders FXR an attractive target for in silico approaches. Integrated to natural product research these computational tools assist to find novel bioactive compounds showing beneficial effects in prevention and treatment of, for example, the metabolic syndrome, dyslipidemia, atherosclerosis, and type 2 diabetes. Virtual screening experiments of our in-house Chinese Herbal Medicine database with structure-based pharmacophore models, previously generated and validated, revealed mainly lanostane-type triterpenes of the TCM fungus Ganoderma lucidum Karst. as putative FXR ligands. To verify the prediction of the in silico approach, two Ganoderma fruit body extracts and compounds isolated thereof were pharmacologically investigated. Pronounced FXR-inducing effects were observed for the extracts at a concentration of 100 μg/mL. Intriguingly, five lanostanes out of 25 secondary metabolites from G. lucidum, that is, ergosterol peroxide (2), lucidumol A (11), ganoderic acid TR (12), ganodermanontriol (13), and ganoderiol F (14), dose-dependently induced FXR in the low micromolar range in a reporter gene assay. To rationalize the binding interactions, additional pharmacophore profiling and molecular docking studies were performed, which allowed establishing a first structure–activity relationship of the investigated triterpenes. PMID:22014750

  19. FXR induces SOCS3 and suppresses hepatocellular carcinoma

    PubMed Central

    Zhang, Yan; Jiang, Peng; Huang, Gang; Chen, Shan; Lyu, Xilin; Zheng, Ping; Zhao, Xin; Zeng, Yijun; Wang, Shuguang; He, Fengtian

    2015-01-01

    Suppressor of cytokine signaling 3 (SOCS3) is regarded as a vital repressor in the liver carcinogenesis mainly by inhibiting signal transducer and activator of transcription 3 (STAT3) activity. Farnesoid X Receptor (FXR), highly expressed in liver, has an important role in protecting against hepatocellular carcinoma (HCC). However, it is unclear whether the tumor suppressive activity of FXR involves the regulation of SOCS3. In the present study, we found that activation of FXR by its specific agonist GW4064 in HCC cells inhibited cell growth, induced cell cycle arrest at G1 phase, elevated p21 expression and repressed STAT3 activity. The above anti-tumor effects of FXR were dramatically alleviated by knockdown of SOCS3 with siRNA. Reporter assay revealed that FXR activation enhanced the transcriptional activity of SOCS3 promoter. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay displayed that FXR directly bound to IR9 DNA motif within SOCS3 promoter region. The in vivo study in nude mice showed that treatment with FXR ligand GW4064 could decelerate the growth of HCC xenografts, up-regulate SOCS3 and p21 expression and inhibit STAT3 phosphorylation in the xenografts. These results suggest that induction of SOCS3 may be a novel mechanism by which FXR exerts its anti-HCC effects, and the FXR-SOCS3 signaling may serve as a new potential target for the prevention/treatment of HCC. PMID:26416445

  20. FXR induces SOCS3 and suppresses hepatocellular carcinoma.

    PubMed

    Guo, Fei; Xu, Zhizhen; Zhang, Yan; Jiang, Peng; Huang, Gang; Chen, Shan; Lyu, Xilin; Zheng, Ping; Zhao, Xin; Zeng, Yijun; Wang, Shuguang; He, Fengtian

    2015-10-27

    Suppressor of cytokine signaling 3 (SOCS3) is regarded as a vital repressor in the liver carcinogenesis mainly by inhibiting signal transducer and activator of transcription 3 (STAT3) activity. Farnesoid X Receptor (FXR), highly expressed in liver, has an important role in protecting against hepatocellular carcinoma (HCC). However, it is unclear whether the tumor suppressive activity of FXR involves the regulation of SOCS3. In the present study, we found that activation of FXR by its specific agonist GW4064 in HCC cells inhibited cell growth, induced cell cycle arrest at G1 phase, elevated p21 expression and repressed STAT3 activity. The above anti-tumor effects of FXR were dramatically alleviated by knockdown of SOCS3 with siRNA. Reporter assay revealed that FXR activation enhanced the transcriptional activity of SOCS3 promoter. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay displayed that FXR directly bound to IR9 DNA motif within SOCS3 promoter region. The in vivo study in nude mice showed that treatment with FXR ligand GW4064 could decelerate the growth of HCC xenografts, up-regulate SOCS3 and p21 expression and inhibit STAT3 phosphorylation in the xenografts. These results suggest that induction of SOCS3 may be a novel mechanism by which FXR exerts its anti-HCC effects, and the FXR-SOCS3 signaling may serve as a new potential target for the prevention/treatment of HCC.

  1. Tissue-specific function of farnesoid X receptor in liver and intestine.

    PubMed

    Zhu, Yan; Li, Fei; Guo, Grace L

    2011-04-01

    Nuclear receptors (NRs) are ligand-activated transcriptional factors that are involved in various physiological, developmental, and toxicological processes. Farnesoid X receptor (FXR) is a NR that belongs to the NR superfamily. The endogenous ligands of FXR are bile acids. FXR is essential in regulating a network of genes involved in maintaining bile acid and lipid homeostasis. It is clear that FXR is critical for liver and intestinal function. In mice FXR deficiency leads to the development of cholestasis, gallstone disease, nonalcoholic steatohepatitis, liver tumor, and colon tumor. Using mouse models where FXR is deleted either in the whole-body, or selectively in hepatocytes or enterocytes, we start to reveal the importance of tissue-specific FXR function in regulating bile acid and lipid homeostasis. However, a great challenge exists for developing tissue-specific FXR modulators to prevent and treat diseases associated with bile acid or lipid disorders. With further understanding of FXR function in both rodents and humans, this nuclear receptor may emerge as a novel target to prevent and treat liver, gastrointestinal and systemic diseases.

  2. 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.

  3. Lepidozenolide from the liverwort Lepidozia fauriana acts as a farnesoid X receptor agonist.

    PubMed

    Lin, Hsiang-Ru

    2015-01-01

    Lepidozenolide is a sesquiterpenoid isolated from the liverwort Lepidozia fauriana and its possible bioactivity is unclear. The farnesoid X receptor (FXR) is a member of nuclear receptor superfamily that has been widely targeted for developing treatments for chronic liver disease and hyperglycemia. In this study, whether lepidozenolide may act as a FXR agonist was determined. Indeed, in mammalian one-hybrid and transient transfection reporter assays, lepidozenolide transactivated FXR to modulate promoter action including GAL4, CYP7A1, and PLTP promoters in a dose-dependent manner, while it exhibited slightly less agonistic activity than chenodeoxycholic acid, an endogenous FXR agonist. Through the molecular modeling docking studies lepidozenolide was shown to bind to FXR ligand binding pocket fairly well. All these results indicate that lepidozenolide acts as a FXR agonist. PMID:25315435

  4. The farnesoid X receptor induces fetuin-B gene expression in human hepatocytes

    PubMed Central

    Murakami, Takeshi; Walczak, Robert; Caron, Sandrine; Duhem, Christian; Vidal, Vincent; Darteil, Raphaël; Staels, Bart

    2007-01-01

    FXR (farnesoid X receptor), a nuclear receptor activated by BAs (bile acids), is a key factor in the regulation of BA, lipid and carbohydrate metabolism. The recent development of synthetic FXR agonists and knockout mouse models has accelerated the discovery of FXR target genes. In the present study, we identify human fetuin-B as a novel FXR target gene. Treatment with FXR agonists increased fetuin-B expression in human primary hepatocytes and in the human hepatoma HepG2 cell line. In contrast, fetuin-B expression was not responsive to FXR agonist treatment in murine primary hepatocytes. Fetuin-B induction by FXR agonist was abolished upon FXR knockdown by siRNA (small interfering RNA). In addition to the previously described P1 promoter, we show that the human fetuin-B gene is also transcribed from an alternative promoter, termed P2. Transcription via the P2 promoter was induced by FXR agonist treatment, whereas P1 promoter activity was not sensitive to FXR agonist treatment. Two putative FXR-response elements [IR-1 (inverted repeat-1)] were identified in the region –1.6 kb upstream of the predicted P2 transcriptional start site. Both motifs bound FXR–RXR (retinoid X receptor) complexes in vitro and were activated by FXR in transient transfection reporter assays. Mutations in the IR-1 sites abolished FXR–RXR binding and activation. Taken together, these results identify human fetuin-B as a new FXR target gene in human hepatocytes. PMID:17655523

  5. 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

  6. Farnesoid X receptor antagonizes JNK signaling pathway in liver carcinogenesis by activating SOD3.

    PubMed

    Wang, Yan-Dong; Chen, Wei-Dong; Li, Cunbao; Guo, Cong; Li, Yanyan; Qi, Hui; Shen, Hailing; Kong, Jing; Long, Xuecheng; Yuan, Frank; Wang, Xichun; Huang, Wendong

    2015-02-01

    The farnesoid X receptor (FXR) is a key metabolic and homeostatic regulator in the liver. In the present work, we identify a novel role of FXR in antagonizing c-Jun N-terminal kinase (JNK) signaling pathway in liver carcinogenesis by activating superoxide dismutase 3 (SOD3) transcription. Compared with wild-type mouse liver, FXR(-/-) mouse liver showed elevated JNK phosphorylation. JNK1 deletion suppressed the increase of diethylnitrosamine-induced tumor number in FXR(-/-) mice. These results suggest that JNK1 plays a key role in chemical-induced liver carcinogenesis in FXR(-/-) mice. We found that ligand-activated FXR was able to alleviate H₂O₂or tetradecanoylphorbol acetate-induced JNK phosphorylation in human hepatoblastoma (HepG2) cells or mouse primary hepatocytes. FXR ligand decreased H₂O₂-induced reactive oxygen species (ROS) levels in wild-type but not FXR(-/-) mouse hepatocytes. FXR knockdown abolished the inhibition of 3-[2-[2-chloro-4-[[3-(2,6-dichlorophenyl)-5-(1-methylethyl)-4-isoxazolyl]methoxy]phenyl]ethenyl]-Benzoic acid (GW4064) on JNK phosphorylation and ROS production induced by H₂O₂in HepG2 cells. The gene expression of SOD3, an antioxidant defense enzyme, was increased by FXR activation in vitro and in vivo. An FXR-responsive element, inverted repeat separated by 1 nucleotide in SOD3 promoter, was identified by a combination of transcriptional reporter assays, EMSAs, and chromatin immunoprecipitation assays, which indicated that SOD3 could be a direct FXR target gene. SOD3 knockdown abolished the inhibition of GW4064 on JNK phosphorylation induced by H₂O₂in HepG2 cells. In summary, FXR may regulate SOD3 expression to suppress ROS production, resulting in decreasing JNK activity. These results suggest that FXR, as a novel JNK suppressor, may be an attractive therapeutic target for liver cancer treatment.

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

    PubMed

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

    2013-11-29

    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.

  8. FXR is a molecular target for the effects of vertical sleeve gastrectomy

    PubMed Central

    Ryan, Karen K.; Tremaroli, Valentina; Clemmensen, Christoffer; Kovatcheva-Datchary, Petia; Myronovych, Andriy; Karns, Rebekah; Wilson-Pérez, Hilary E.; Sandoval, Darleen A.; Kohli, Rohit; Bäckhed, Fredrik; Seeley, Randy J.

    2014-01-01

    SUMMARY Bariatric surgical procedures, such as vertical sleeve gastrectomy (VSG), are currently the most effective therapy for the treatment of obesity, and are associated with substantial improvements in co-morbidities, including type-2 diabetes mellitus. The underlying molecular mechanisms contributing to these benefits remain largely undetermined, despite offering tremendous potential to reveal new targets for therapeutic intervention. The present study demonstrates that the therapeutic value of VSG does not result from mechanical restriction imposed by a smaller stomach. Rather, we report that VSG is associated with increased circulating bile acids, and associated changes to gut microbial communities. Moreover, in the absence of nuclear bile acid receptor FXR, the ability of VSG to reduce body weight and improve glucose tolerance is substantially reduced. These results point to bile acids and FXR signaling as an important molecular underpinning for the beneficial effects of this weight-loss surgery. PMID:24670636

  9. MiR-22-silenced Cyclin A Expression in Colon and Liver Cancer Cells Is Regulated by Bile Acid Receptor*

    PubMed Central

    Yang, Fan; Hu, Ying; Liu, Hui-Xin; Wan, Yu-Jui Yvonne

    2015-01-01

    Because of the significant tumor-suppressive role of microRNA-22 (miR-22), the current study was designed to understand the regulation of miR-22 and to identify additional downstream miR-22 targets in liver and colon cells. The data showed that miR-22 was transcriptionally regulated by bile acid receptor farnesoid X receptor (FXR) through direct binding to an invert repeat 1 motif located at −1012 to −1025 bp upstream from miR-22. Among the studied primary and secondary bile acids, chenodeoxycholic acid, which has the highest binding affinity to FXR, induced miR-22 level in both Huh7 liver and HCT116 colon cells in a dose- and time-dependent manner. In addition, cyclin A2 (CCNA2) was identified as a miR-22 novel target in liver and colon cancer cells. The sequence of miR-22, which is conserved in mice, rats, humans, and other mammalians, aligns with the sequence of 3′-UTR of CCNA2. Chenodeoxycholic acid treatment and miR-22 mimics reduced CCNA2 protein and increased the number of G0/G1 Huh7 and HCT116 cells. In FXR KO mice, reduction of miR-22 was accompanied by elevated hepatic and ileal CCNA2 protein, as well as an increased number of hepatic and colonic Ki-67-positive cells. In humans, the expression levels of miR-22 and CCNA2 are inversely correlated in liver and colon cancers. Taken together, our data showed that bile acid-activated FXR stimulates miR-22-silenced CCNA2, a novel pathway for FXR to exert its protective effect in the gastrointestinal tract. PMID:25596928

  10. 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.

  11. The farnesoid X receptor modulates adiposity and peripheral insulin sensitivity in mice.

    PubMed

    Cariou, Bertrand; van Harmelen, Kirsten; Duran-Sandoval, Daniel; van Dijk, Theo H; Grefhorst, Aldo; Abdelkarim, Mouaadh; Caron, Sandrine; Torpier, Gérard; Fruchart, Jean-Charles; Gonzalez, Frank J; Kuipers, Folkert; Staels, Bart

    2006-04-21

    The farnesoid X receptor (FXR) is a bile acid (BA)-activated nuclear receptor that plays a major role in the regulation of BA and lipid metabolism. Recently, several studies have suggested a potential role of FXR in the control of hepatic carbohydrate metabolism, but its contribution to the maintenance of peripheral glucose homeostasis remains to be established. FXR-deficient mice display decreased adipose tissue mass, lower serum leptin concentrations, and elevated plasma free fatty acid levels. Glucose and insulin tolerance tests revealed that FXR deficiency is associated with impaired glucose tolerance and insulin resistance. Moreover, whole-body glucose disposal during a hyperinsulinemic euglycemic clamp is decreased in FXR-deficient mice. In parallel, FXR deficiency alters distal insulin signaling, as reflected by decreased insulin-dependent Akt phosphorylation in both white adipose tissue and skeletal muscle. Whereas FXR is not expressed in skeletal muscle, it was detected at a low level in white adipose tissue in vivo and induced during adipocyte differentiation in vitro. Moreover, mouse embryonic fibroblasts derived from FXR-deficient mice displayed impaired adipocyte differentiation, identifying a direct role for FXR in adipocyte function. Treatment of differentiated 3T3-L1 adipocytes with the FXR-specific synthetic agonist GW4064 enhanced insulin signaling and insulin-stimulated glucose uptake. Finally, treatment with GW4064 improved insulin resistance in genetically obese ob/ob mice in vivo. Although the underlying molecular mechanisms remain to be unraveled, these results clearly identify a novel role of FXR in the regulation of peripheral insulin sensitivity and adipocyte function. This unexpected function of FXR opens new perspectives for the treatment of type 2 diabetes.

  12. Human kininogen gene is transactivated by the farnesoid X receptor.

    PubMed

    Zhao, Annie; Lew, Jane-L; Huang, Li; Yu, Jinghua; Zhang, Theresa; Hrywna, Yaroslav; Thompson, John R; de Pedro, Nuria; Blevins, Richard A; Peláez, Fernando; Wright, Samuel D; Cui, Jisong

    2003-08-01

    Human kininogen belongs to the plasma kallikreinkinin system. High molecular weight kininogen is the precursor for two-chain kinin-free kininogen and bradykinin. It has been shown that the two-chain kinin-free kininogen has the properties of anti-adhesion, anti-platelet aggregation, and anti-thrombosis, whereas bradykinin is a potent vasodilator and mediator of inflammation. In this study we show that the human kininogen gene is strongly up-regulated by agonists of the farnesoid X receptor (FXR), a nuclear receptor for bile acids. In primary human hepatocytes, both the endogenous FXR agonist chenodeoxycholate and synthetic FXR agonist GW4064 increased kininogen mRNA with a maximum induction of 8-10-fold. A more robust induction of kininogen expression was observed in HepG2 cells, where kininogen mRNA was increased by chenodeoxycholate or GW4064 up to 130-140-fold as shown by real time PCR. Northern blot analysis confirmed the up-regulation of kininogen expression by FXR agonists. To determine whether kininogen is a direct target of FXR, we examined the sequence of the kininogen promoter and identified a highly conserved FXR response element (inverted repeat, IR-1) in the proximity of the kininogen promoter (-66/-54). FXR/RXRalpha heterodimers specifically bind to this IR-1. A construct of a minimal promoter with the luciferase reporter containing this IR-1 was transactivated by FXR. Deletion or mutation of this IR-1 abolished FXR-mediated promoter activation, indicating that this IR-1 element is responsible for the promoter transactivation by FXR. We conclude that kininogen is a novel and direct target of FXR, and bile acids may play a role in the vasodilation and anti-coagulation processes.

  13. Regulation of human class I alcohol dehydrogenases by bile acids

    PubMed Central

    Langhi, Cédric; Pedraz-Cuesta, Elena; Haro, Diego; Marrero, Pedro F.; Rodríguez, Joan C.

    2013-01-01

    Class I alcohol dehydrogenases (ADH1s) are the rate-limiting enzymes for ethanol and vitamin A (retinol) metabolism in the liver. Because previous studies have shown that human ADH1 enzymes may participate in bile acid metabolism, we investigated whether the bile acid-activated nuclear receptor farnesoid X receptor (FXR) regulates ADH1 genes. In human hepatocytes, both the endogenous FXR ligand chenodeoxycholic acid and synthetic FXR-specific agonist GW4064 increased ADH1 mRNA, protein, and activity. Moreover, overexpression of a constitutively active form of FXR induced ADH1A and ADH1B expression, whereas silencing of FXR abolished the effects of FXR agonists on ADH1 expression and activity. Transient transfection studies and electrophoretic mobility shift assays revealed functional FXR response elements in the ADH1A and ADH1B proximal promoters, thus indicating that both genes are direct targets of FXR. These findings provide the first evidence for direct connection of bile acid signaling and alcohol metabolism. PMID:23772048

  14. Regulation of human class I alcohol dehydrogenases by bile acids.

    PubMed

    Langhi, Cédric; Pedraz-Cuesta, Elena; Haro, Diego; Marrero, Pedro F; Rodríguez, Joan C

    2013-09-01

    Class I alcohol dehydrogenases (ADH1s) are the rate-limiting enzymes for ethanol and vitamin A (retinol) metabolism in the liver. Because previous studies have shown that human ADH1 enzymes may participate in bile acid metabolism, we investigated whether the bile acid-activated nuclear receptor farnesoid X receptor (FXR) regulates ADH1 genes. In human hepatocytes, both the endogenous FXR ligand chenodeoxycholic acid and synthetic FXR-specific agonist GW4064 increased ADH1 mRNA, protein, and activity. Moreover, overexpression of a constitutively active form of FXR induced ADH1A and ADH1B expression, whereas silencing of FXR abolished the effects of FXR agonists on ADH1 expression and activity. Transient transfection studies and electrophoretic mobility shift assays revealed functional FXR response elements in the ADH1A and ADH1B proximal promoters, thus indicating that both genes are direct targets of FXR. These findings provide the first evidence for direct connection of bile acid signaling and alcohol metabolism.

  15. Regulation of carbohydrate metabolism by the farnesoid X receptor.

    PubMed

    Stayrook, Keith R; Bramlett, Kelli S; Savkur, Rajesh S; Ficorilli, James; Cook, Todd; Christe, Michael E; Michael, Laura F; Burris, Thomas P

    2005-03-01

    The farnesoid X receptor (FXR; NR1H4) is a nuclear hormone receptor that functions as the bile acid receptor. In addition to the critical role FXR plays in bile acid metabolism and transport, it regulates a variety of genes important in lipoprotein metabolism. We demonstrate that FXR also plays a role in carbohydrate metabolism via regulation of phosphoenolpyruvate carboxykinase (PEPCK) gene expression. Treatment of either H4IIE or MH1C1 rat hepatoma cell lines as well as primary rat or human hepatocytes with FXR agonists led to stimulation of PEPCK mRNA expression to levels comparable to those obtained with glucocorticoid receptor agonists. We examined the physiological significance of FXR agonist-induced enhancement of PEPCK expression in primary rat hepatocytes. In addition to inducing PEPCK expression in primary hepatocytes, FXR agonists stimulated glucose output to levels comparable to those observed with a glucocorticoid receptor agonist. Consistent with these observations, treatment of C57BL6 mice with GW4064 significantly increased hepatic PEPCK expression. Activation of FXR initiated a cascade involving induction of peroxisome proliferator-activated receptor alpha and TRB3 expression that is consistent with stimulation of PEPCK gene expression via interference with a pathway that may involve Akt-dependent phosphorylation of Forkhead/winged helix transcription factor (FOXO1). The FXR-peroxisome proliferator-activated receptor alpha-TRB3 pathway was conserved in rat hepatoma cell lines, mice, as well as primary human hepatocytes. Thus, in addition to its role in the regulation of lipid metabolism, FXR regulates carbohydrate metabolism.

  16. The glucocorticoid mometasone furoate is a novel FXR ligand that decreases inflammatory but not metabolic gene expression

    PubMed Central

    Bijsmans, Ingrid T. G. W.; Guercini, Chiara; Ramos Pittol, José M.; Omta, Wienand; Milona, Alexandra; Lelieveld, Daphne; Egan, David A.; Pellicciari, Roberto; Gioiello, Antimo; van Mil, Saskia W. C.

    2015-01-01

    The Farnesoid X receptor (FXR) regulates bile salt, glucose and cholesterol homeostasis by binding to DNA response elements, thereby activating gene expression (direct transactivation). FXR also inhibits the immune response via tethering to NF-κB (tethering transrepression). FXR activation therefore has therapeutic potential for liver and intestinal inflammatory diseases. We aim to identify and develop gene-selective FXR modulators, which repress inflammation, but do not interfere with its metabolic capacity. In a high-throughput reporter-based screen, mometasone furoate (MF) was identified as a compound that reduced NF-κB reporter activity in an FXR-dependent manner. MF reduced mRNA expression of pro-inflammatory cytokines, and induction of direct FXR target genes in HepG2-GFP-FXR cells and intestinal organoids was minor. Computational studies disclosed three putative binding modes of the compound within the ligand binding domain of the receptor. Interestingly, mutation of W469A residue within the FXR ligand binding domain abrogated the decrease in NF-κB activity. Finally, we show that MF-bound FXR inhibits NF-κB subunit p65 recruitment to the DNA of pro-inflammatory genes CXCL2 and IL8. Although MF is not suitable as selective anti-inflammatory FXR ligand due to nanomolar affinity for the glucocorticoid receptor, we show that separation between metabolic and anti-inflammatory functions of FXR can be achieved. PMID:26369990

  17. 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.

  18. 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.

  19. Coordinated regulation of dimethylarginine dimethylaminohydrolase-1 and cationic amino acid transporter-1 by farnesoid X receptor in mouse liver and kidney and its implication in the control of blood levels of asymmetric dimethylarginine.

    PubMed

    Li, Jiang; Wilson, Annette; Gao, Xiang; Kuruba, Ramalinga; Liu, Youhua; Poloyac, Samuel; Pitt, Bruce; Xie, Wen; Li, Song

    2009-10-01

    Asymmetric dimethylarginine (ADMA) is a potent endogenous inhibitor of endothelial nitric-oxide synthase (eNOS), and increased plasma concentrations of ADMA have been regarded as a risk factor for a number of cardiovascular diseases. Circulating ADMA is largely taken up by liver and kidney via system y(+) carriers of the cationic amino acid (CAT) family and subsequently metabolized by dimethylarginine dimethylaminohydrolases (DDAHs). As such, agents targeted at enhancing ADMA metabolism may prove to be useful in the prevention and/or treatment of various types of cardiovascular disease. Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily and plays an important role in the maintenance of cholesterol and bile acid homeostasis. We report here that treatment of mice with an FXR agonist 3-(2,6-dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl)oxymethyl-5-isopropylisoxazole; GW4064) led to increased expression of DDAH-1 and CAT-1 in both liver and kidney. In cultured human hepatocytes and kidney proximal tubular epithelial cells, GW4064 increased CAT-1 expression, and this was associated with a significant increase in the cellular uptake of ADMA. Promoter analyses suggest that CAT-1 is a likely target of FXR, and a functional FXR response element was found in the promoter region of CAT-1 gene. These data suggest that FXR may play an important role in regulating blood levels of ADMA via coordinated regulation of DDAH-1 and CAT-1 in liver and kidney.

  20. Diacylglycerol kinase θ couples farnesoid X receptor-dependent bile acid signalling to Akt activation and glucose homoeostasis in hepatocytes.

    PubMed

    Cai, Kai; Sewer, Marion B

    2013-09-01

    DGKs (diacylglycerol kinases) catalyse the conversion of diacylglycerol into PA (phosphatidic acid), a positive modulator of mTOR (mammalian target of rapamycin). We have found that chenodeoxycholic acid and the synthetic FXR (farnesoid X receptor) ligand GW4064 induce the mRNA and protein expression of DGKθ in the HepG2 cell line and in primary human hepatocytes. Reporter gene studies using 1.5 kB of the DGKθ promoter fused to the luciferase gene revealed that bile acids increase DGKθ transcriptional activity. Mutation of putative FXR-binding sites attenuated the ability of GW4046 to increase DGKθ luciferase activity. Consistent with this finding, ChIP (chromatin immunoprecipitation) assays demonstrated that bile acid signalling increased the recruitment of FXR to the DGKθ promoter. Furthermore, GW4064 evoked a time-dependent increase in the cellular concentration of PA. We also found that GW4064 and PA promote the phosphorylation of mTOR, Akt and FoxO1 (forkhead box O1), and that silencing DGKθ expression significantly abrogated the ability of GW4046 to promote the phosphorylation of these PA-regulated targets. DGKθ was also required for bile-acid-dependent decreased glucose production. Taken together, our results establish DGKθ as a key mediator of bile-acid-stimulated modulation of mTORC2 (mTOR complex 2), the Akt pathway and glucose homoeostasis.

  1. Farnesoid X receptor protects liver cells from apoptosis induced by serum deprivation in vitro and fasting in vivo.

    PubMed

    Wang, Yan-Dong; Yang, Fan; Chen, Wei-Dong; Huang, Xiongfei; Lai, Lily; Forman, Barry M; Huang, Wendong

    2008-07-01

    The farnesoid X receptor (FXR) is a key metabolic regulator in the liver by maintaining the homeostasis of liver metabolites. Recent findings suggest that FXR may have a much broader function in liver physiology and pathology. In the present work, we identify a novel role of FXR in protecting liver cell from apoptosis induced by nutritional withdrawal including serum deprivation in vitro or starvation in vivo. Two FXR ligands, chenodeoxycholic acid (CDCA) and GW4064, rescued HepG2 cells from serum deprivation-induced apoptosis in a dose-dependent manner. This effect of FXR on apoptotic suppression was compromised when FXR was knocked down by short interfering RNA. Similarly, the effects of both CDCA and GW4064 were abolished after inhibition of the MAPK pathway by a specific inhibitor of MAPK kinase 1/2. Immunoblotting results indicated that FXR activation by CDCA and GW4064 induced ERK1/2 phosphorylation, which was attenuated by serum deprivation. In vivo, FXR(-/-) mice exhibited an exacerbated liver apoptosis and lower levels of phosphorylated-ERK1/2 compared to wild-type mice after starvation. In conclusion, our results suggest a novel role of FXR in modulating liver cell apoptosis.

  2. Ursodeoxycholic acid exerts farnesoid X receptor-antagonistic effects on bile acid and lipid metabolism in morbid obesity

    PubMed Central

    Mueller, Michaela; Thorell, Anders; Claudel, Thierry; Jha, Pooja; Koefeler, Harald; Lackner, Carolin; Hoesel, Bastian; Fauler, Guenter; Stojakovic, Tatjana; Einarsson, Curt; Marschall, Hanns-Ulrich; Trauner, Michael

    2015-01-01

    Background & Aims Bile acids (BAs) are major regulators of hepatic BA and lipid metabolism but their mechanisms of action in non-alcoholic fatty liver disease (NAFLD) are still poorly understood. Here we aimed to explore the molecular and biochemical mechanisms of ursodeoxycholic acid (UDCA) in modulating the cross-talk between liver and visceral white adipose tissue (vWAT) regarding BA and cholesterol metabolism and fatty acid/lipid partitioning in morbidly obese NAFLD patients. Methods In this randomized controlled pharmacodynamic study, we analyzed serum, liver and vWAT samples from 40 well-matched morbidly obese patients receiving UDCA (20 mg/kg/day) or no treatment three weeks prior to bariatric surgery. Results Short term UDCA administration stimulated BA synthesis by reducing circulating fibroblast growth factor 19 and farnesoid X receptor (FXR) activation, resulting in cholesterol 7α-hydroxylase induction mirrored by elevated C4 and 7α-hydroxycholesterol. Enhanced BA formation depleted hepatic and LDL-cholesterol with subsequent activation of the key enzyme of cholesterol synthesis 3-hydroxy-3-methylglutaryl-CoA reductase. Blunted FXR anti-lipogenic effects induced lipogenic stearoyl-CoA desaturase (SCD) in the liver, thereby increasing hepatic triglyceride content. In addition, induced SCD activity in vWAT shifted vWAT lipid metabolism towards generation of less toxic and more lipogenic monounsaturated fatty acids such as oleic acid. Conclusion These data demonstrate that by exerting FXR-antagonistic effects, UDCA treatment in NAFLD patients strongly impacts on cholesterol and BA synthesis and induces neutral lipid accumulation in both liver and vWAT. PMID:25617503

  3. Development of time resolved fluorescence resonance energy transfer-based assay for FXR antagonist discovery.

    PubMed

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

    2013-07-15

    FXR (farnesoid X receptor, NRIH4), a nuclear receptor, plays a major role in the control of cholesterol metabolism. FXR ligands have been investigated in preclinical studies for targeted therapy against metabolic diseases, but have shown limitations. Therefore, there is a need for new agonist or antagonist ligands of FXR, both for potential clinical applications, as well as to further elucidate its biological functions. Here we describe the use of the X-ray crystal structure of FXR complexed with the potent small molecule agonist GW4064 to design and synthesize a novel fluorescent, high-affinity probe (DY246) for time resolved fluorescence resonance energy transfer (TR-FRET) assays. We then used the TR-FRET assay for high throughput screening of a library of over 5000 bioactive compounds. From this library, we identified 13 compounds that act as putative FXR transcriptional antagonists.

  4. Pregnane X receptor is a target of farnesoid X receptor.

    PubMed

    Jung, Diana; Mangelsdorf, David J; Meyer, Urs A

    2006-07-14

    The pregnane X receptor (PXR) is an essential component of the body's detoxification system. PXR is activated by a broad spectrum of xenobiotics and endobiotics, including bile acids and their precursors. Bile acids in high concentrations are toxic; therefore, their synthesis is tightly regulated by the farnesoid X receptor, and their catabolism involves several enzymes regulated by PXR. Here we demonstrate that the expression of PXR is regulated by farnesoid X receptor. Feeding mice with cholic acid or the synthetic farnesoid X receptor (FXR) agonist GW4064 resulted in a robust PXR induction. This effect was abolished in FXR knock-out mice. Long time bile acid treatment resulted in an increase of PXR target genes in wild type mice. A region containing four FXR binding sites (IR1) was identified in the mouse Pxr gene. This region was able to trigger an 8-fold induction after GW4064 treatment in transactivation studies. Deletion or mutation of single IR1 sites caused a weakened response. The importance of each individual IR1 element was assessed by cloning a triple or a single copy and was tested in transactivation studies. Two elements were able to trigger a strong response, one a moderate response, and one no response to GW4064 treatment. Mobility shift assays demonstrated that the two stronger responding elements were able to bind FXR protein. This result was confirmed by chromatin immunoprecipitation. These results strongly suggest that PXR is regulated by FXR. Bile acids activate FXR, which blocks synthesis of bile acids and also leads to the transcriptional activation of PXR, promoting breakdown of bile acids. The combination of the two mechanisms leads to an efficient protection of the liver against bile acid induced toxicity.

  5. O-GlcNAcylation Links ChREBP and FXR to Glucose-Sensing

    PubMed Central

    Benhamed, Fadila; Filhoulaud, Gaelle; Caron, Sandrine; Lefebvre, Philippe; Staels, Bart; Postic, Catherine

    2015-01-01

    Accumulating evidence suggests that O-GlcNAc transferase, an enzyme responsible for O-GlcNAc post-translational modification acts as a nutrient sensor that links glucose and the hexosamine biosynthetic pathway to the regulation of transcriptional factors involved in energy homeostasis. In liver, glucose signaling is mediated by carbohydrate response element-binding protein (ChREBP), which stimulates glycolytic and lipogenic gene expression through its binding on a specific ChoRE DNA sequence. Modulation of ChREBP by O-GlcNAcylation increases its DNA binding affinity and its activity. ChREBP transcriptional activity also depends on the presence of several other co-factors and transcriptional factors. Among them, the nuclear Farnesoid X Receptor (FXR), a key transcription factor of bile acid metabolism involved in the gut–liver axis homeostasis was recently shown to directly interact with ChREBP, acting as a repressor on the ChoRE of glycolytic genes. Interestingly, similarly to ChREBP, FXR is O-GlcNAcylated in response to glucose. This review discusses the importance of ChREBP and FXR modifications through O-GlcNAcylation in liver and how glucose can modify their mutual affinity and transcriptional activity. PMID:25628602

  6. Bridging cell surface receptor with nuclear receptors in control of bile acid homeostasis

    PubMed Central

    Li, Shuangwei; Ni, Andrew; Feng, Gen-sheng

    2015-01-01

    Bile acids (BAs) are traditionally considered as “physiological detergents” for emulsifying hydrophobic lipids and vitamins due to their amphipathic nature. But accumulating clinical and experimental evidence shows an association between disrupted BA homeostasis and various liver disease conditions including hepatitis infection, diabetes and cancer. Consequently, BA homeostasis regulation has become a field of heavy interest and investigation. After identification of the Farnesoid X Receptor (FXR) as an endogenous receptor for BAs, several nuclear receptors (SHP, HNF4α, and LRH-1) were also found to be important in regulation of BA homeostasis. Some post-translational modifications of these nuclear receptors have been demonstrated, but their physiological significance is still elusive. Gut secrets FGF15/19 that can activate hepatic FGFR4 and its downstream signaling cascade, leading to repressed hepatic BA biosynthesis. However, the link between the activated kinases and these nuclear receptors is not fully elucidated. Here, we review the recent literature on signal crosstalk in BA homeostasis. PMID:25500873

  7. Farnesoid X receptor activation promotes cell proliferation via PDK4-controlled metabolic reprogramming

    PubMed Central

    Xie, Yang; Wang, Hong; Cheng, Xuefang; Wu, Yuzheng; Cao, Lijuan; Wu, Mengqiu; Xie, Wen; Wang, Guangji; Hao, Haiping

    2016-01-01

    Farnesoid X receptor (FXR) plays a pivotal role in the regulation of various metabolic pathways as well as liver regeneration. However, the casual link between cell proliferative effects during liver regeneration and metabolic regulation of FXR was elusive. In this study, we found that FXR activation significantly promotes HepG2 cell proliferation accompanied with metabolic switch towards the excessive accumulation of aerobic glycolytic intermediates including lactic acid, pyruvate and the subsequently increased biosynthesis of glycine. This FXR-induced metabolic switch was found dependent on an up-regulation of pyruvate dehydrogenate kinase 4 (PDK4), a FXR target gene. FXR agonists were found to promote liver regeneration in the murine model of APAP induced liver injury, which was associated with a metabolic switch favoring the accumulation of glycolytic intermediates as precursors for generation of biomass. However, FXR activation has little effect on the glycolytic metabolism in healthy primary hepatocytes in vitro and the liver of healthy mice in vivo. Therefore, we conclude that FXR may promote the proliferation of tumor cells and the hepatocytes in the process of liver regeneration by activating the PDK4-mediated metabolic reprogramming to generate glycolytic intermediates essential for rapid biomass generation, establishing a mechanistic link between cell proliferation and metabolic switch. PMID:26728993

  8. A dysregulated acetyl/SUMO switch of FXR promotes hepatic inflammation in obesity

    PubMed Central

    Kim, Dong-Hyun; Xiao, Zhen; Kwon, Sanghoon; Sun, Xiaoxiao; Ryerson, Daniel; Tkac, David; Ma, Ping; Wu, Shwu-Yuan; Chiang, Cheng-Ming; Zhou, Edward; Xu, H Eric; Palvimo, Jorma J; Chen, Lin-Feng; Kemper, Byron; Kemper, Jongsook Kim

    2015-01-01

    Acetylation of transcriptional regulators is normally dynamically regulated by nutrient status but is often persistently elevated in nutrient-excessive obesity conditions. We investigated the functional consequences of such aberrantly elevated acetylation of the nuclear receptor FXR as a model. Proteomic studies identified K217 as the FXR acetylation site in diet-induced obese mice. In vivo studies utilizing acetylation-mimic and acetylation-defective K217 mutants and gene expression profiling revealed that FXR acetylation increased proinflammatory gene expression, macrophage infiltration, and liver cytokine and triglyceride levels, impaired insulin signaling, and increased glucose intolerance. Mechanistically, acetylation of FXR blocked its interaction with the SUMO ligase PIASy and inhibited SUMO2 modification at K277, resulting in activation of inflammatory genes. SUMOylation of agonist-activated FXR increased its interaction with NF-κB but blocked that with RXRα, so that SUMO2-modified FXR was selectively recruited to and trans-repressed inflammatory genes without affecting FXR/RXRα target genes. A dysregulated acetyl/SUMO switch of FXR in obesity may serve as a general mechanism for diminished anti-inflammatory response of other transcriptional regulators and provide potential therapeutic and diagnostic targets for obesity-related metabolic disorders. PMID:25425577

  9. 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

    2015-01-01

    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. PMID:26134028

  10. Farnesoid X receptor agonist reduces serum asymmetric dimethylarginine levels through hepatic dimethylarginine dimethylaminohydrolase-1 gene regulation.

    PubMed

    Hu, Tonghuan; Chouinard, Michael; Cox, Amy L; Sipes, Philip; Marcelo, Marialuisa; Ficorilli, James; Li, Shuyu; Gao, Hong; Ryan, Timothy P; Michael, M Dodson; Michael, Laura F

    2006-12-29

    The farnesoid X receptor (FXR, NR1H4) is a bile acid-responsive nuclear receptor that plays critical roles in the transcriptional regulation genes involved in cholesterol, bile acid, triglyceride, and carbohydrate metabolism. By microarray analysis of hepatic genes from female Zucker diabetic fatty (ZDF) rats treated with the FXR agonist GW4064, we have identified dimethylarginine dimethylaminohydrolase-1 (DDAH1) as an FXR target gene. DDAH1 is a key catabolic enzyme of asymmetric dimethylarginine (ADMA), a major endogenous nitric-oxide synthase inhibitor. Sequence analysis of the DDAH1 gene reveals the presence of an FXR response element (FXRE) located 90 kb downstream of the transcription initiation site and within the first intron. Functional analysis of the putative FXRE demonstrated GW4064 dose-dependent transcriptional activation from the element, and we have demonstrated that the FXRE sequence binds the FXR-RXR heterodimer. In vivo administration of GW4064 to female ZDF rats promoted a dose-dependent and >6-fold increase in hepatic DDAH1 gene expression. The level of serum ADMA was reduced concomitantly. These findings provide a mechanism by which FXR may increase endothelium-derived nitric oxide levels through modulation of serum ADMA levels via direct regulation of hepatic DDAH1 gene expression. Thus, beneficial clinical outcomes of FXR agonist therapy may include prevention of atherosclerosis and improvement of the metabolic syndrome.

  11. Farnesoid X receptor, through the binding with steroidogenic factor 1-responsive element, inhibits aromatase expression in tumor Leydig cells.

    PubMed

    Catalano, Stefania; Malivindi, Rocco; Giordano, Cinzia; Gu, Guowei; Panza, Salvatore; Bonofiglio, Daniela; Lanzino, Marilena; Sisci, Diego; Panno, Maria Luisa; Andò, Sebastiano

    2010-02-19

    The farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily that regulates bile acid homeostasis. It is expressed in the liver and the gastrointestinal tract, but also in several non-enterohepatic tissues including testis. Recently, FXR was identified as a negative modulator of the androgen-estrogen-converting aromatase enzyme in human breast cancer cells. In the present study we detected the expression of FXR in Leydig normal and tumor cell lines and in rat testes tissue. We found, in rat Leydig tumor cells, R2C, that FXR activation by the primary bile acid chenodeoxycholic acid (CDCA) or a synthetic agonist GW4064, through a SHP-independent mechanism, down-regulates aromatase expression in terms of mRNA, protein levels, and its enzymatic activity. Transient transfection experiments, using vector containing rat aromatase promoter PII, evidenced that CDCA reduces basal aromatase promoter activity. Mutagenesis studies, electrophoretic mobility shift, and chromatin immunoprecipitation analysis reveal that FXR is able to compete with steroidogenic factor 1 in binding to a common sequence present in the aromatase promoter region interfering negatively with its activity. Finally, the FXR-mediated anti-proliferative effects exerted by CDCA on tumor Leydig cells are at least in part due to an inhibition of estrogen-dependent cell growth. In conclusion our findings identify for the first time the activators of FXR as negative modulators of the aromatase enzyme in Leydig tumor cell lines.

  12. Farnesoid X receptor activation by chenodeoxycholic acid induces detoxifying enzymes through AMP-activated protein kinase and extracellular signal-regulated kinase 1/2-mediated phosphorylation of CCAAT/enhancer binding protein β.

    PubMed

    Noh, Kyoung; Kim, Young Mi; Kim, Young Woo; Kim, Sang Geon

    2011-08-01

    Farnesoid X receptor (FXR) regulates redox homeostasis and elicits a cytoprotective effect. CCAAT/enhancer binding protein-β (C/EBPβ) plays a role in regulating the expression of hepatocyte-specific genes and contributes to hepatocyte protection and liver regeneration. In view of the role of FXR in xenobiotic metabolism and hepatocyte survival, this study investigated the potential of FXR to activate C/EBPβ for the induction of detoxifying enzymes and the responsible regulatory pathway. Chenodeoxycholic acid (CDCA), a major component in bile acids, activates FXR. In HepG2 cells, CDCA treatment activated C/EBPβ, as shown by increases in its phosphorylation, nuclear accumulation, and expression. 3-(2,6-Dichlorophenyl)-4-(3'-carboxy-2-chlorostilben-4-yl-)oxymethyl-5-isopropyl-isoxazole (GW4064), a synthetic FXR ligand, had similar effects. In addition, CDCA enhanced luciferase gene transcription from the construct containing -1.65-kb GSTA2 promoter, which contained C/EBP response element (pGL-1651). Moreover, CDCA treatment activated AMP-activated protein kinase (AMPK), which led to extracellular signal-regulated kinase 1/2 (ERK1/2) activation, as evidenced by the results of experiments using a dominant-negative mutant of AMPKα and chemical inhibitor. The activation of ERK1/2 was responsible for the activating phosphorylation of C/EBPβ. FXR knockdown attenuated the ability of CDCA to activate AMPK and ERK1/2 and phosphorylate C/EBPβ. Consistently, enforced expression of FXR promoted the phosphorylation of AMPKα, ERK1/2, and C/EBPβ, verifying that C/EBPβ phosphorylation elicited by CDCA results from the activation of AMPK and ERK1/2 by FXR. In mice, CDCA treatment activated C/EBPβ with the induction of detoxifying enzymes in the liver. Our results demonstrate that CDCA induces antioxidant and xenobiotic-metabolizing enzymes by activating C/EBPβ through AMPK-dependent ERK1/2 pathway downstream of FXR.

  13. Suppression of interleukin-6-induced C-reactive protein expression by FXR agonists

    SciTech Connect

    Zhang Songwen Liu Qiangyuan; Wang Juan; Harnish, Douglas C.

    2009-02-06

    C-reactive protein (CRP), a human acute-phase protein, is a risk factor for future cardiovascular events and exerts direct pro-inflammatory and pro-atherogenic properties. The farnesoid X receptor (FXR), a member of the nuclear hormone receptor superfamily, plays an essential role in the regulation of enterohepatic circulation and lipid homeostasis. In this study, we report that two synthetic FXR agonists, WAY-362450 and GW4064, suppressed interleukin-6-induced CRP expression in human Hep3B hepatoma cells. Knockdown of FXR by short interfering RNA attenuated the inhibitory effect of the FXR agonists and also increased the ability of interleukin-6 to induce CRP production. Furthermore, treatment of wild type C57BL/6 mice with the FXR agonist, WAY-362450, attenuated lipopolysaccharide-induced serum amyloid P component and serum amyloid A3 mRNA levels in the liver, whereas no effect was observed in FXR knockout mice. These data provide new evidence for direct anti-inflammatory properties of FXR.

  14. FXR agonists enhance the sensitivity of biliary tract cancer cells to cisplatin via SHP dependent inhibition of Bcl-xL expression

    PubMed Central

    Wang, Wei; Zhan, Ming; Li, Qi; Chen, Wei; Chu, Huiling; Huang, Qihong; Hou, Zhaoyuan; Man, Mohan; Wang, Jian

    2016-01-01

    Chemoresistance is common in patients with biliary tract cancer (BTC) including gallbladder cancer (GBC) and cholangiocarcinoma (CC). Therefore, it is necessary to identify effective chemotherapeutic agents for BTC. In the present study, we for the first time tested the effect of farnesoid X receptor (FXR) agonists GW4064 and CDCA (chenodeoxycholic acid) in combination with cisplatin (CDDP) on increasing the chemosensitivity in BTC. Our results show that co-treatment of CDDP with FXR agonists remarkably enhance chemosensitivity of BTC cells. Mechanistically, we found that activation of FXR induced expression of small heterodimer partner (SHP), which in turn inhibited signal transducer and activator of transcription 3 (STAT3) phosphorylation and resulted in down-regulation of Bcl-xL expression in BTC cells, leading to increased susceptibility to CDDP. Moreover, the experiments on tumor-bearing mice showed that GW4064/CDDP co-treatment inhibited the tumor growth in vivo by up-regulating SHP expression and down-regulating STAT3 phosphorylation. These results suggest CDDP in combination with FXR agonists could be a potential new therapeutic strategy for BTC. PMID:27127878

  15. Transactivation and Coactivator Recruitment Assays for Measuring Farnesoid X Receptor Activity.

    PubMed

    Hsu, Chia-Wen Amy; Zhao, Jinghua; Xia, Menghang

    2016-01-01

    The farnesoid X receptor (FXR) is a nuclear receptor responsible for homeostasis of bile acids, lipids, and glucose. Compounds that alter endogenous FXR signaling can be used as therapeutic candidates or identified as potentially hazardous compounds depending on exposure doses and health states. Therefore, there is an increasing need for high-throughput screening assays of FXR activity to profile large numbers of environmental chemicals and drugs. This chapter describes a workflow of FXR modulator identification and characterization. To identify compounds that modulate FXR transactivation at the cellular level, we first screen compounds from the Tox21 10 K compound library in an FXR-driven beta-lactamase reporter gene assay multiplexed with a cell viability assay in the same well of the 1536-well plates. The selected compounds are then tested biochemically for their ability to modulate FXR-coactivator binding interactions using a time-resolved fluorescence resonance energy transfer (TR-FRET) coactivator assay. The assay results from the workflow can be used to prioritize compounds for more extensive investigations. PMID:27518622

  16. 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

  17. Transactivation and Coactivator Recruitment Assays for Measuring Farnesoid X Receptor Activity.

    PubMed

    Hsu, Chia-Wen Amy; Zhao, Jinghua; Xia, Menghang

    2016-01-01

    The farnesoid X receptor (FXR) is a nuclear receptor responsible for homeostasis of bile acids, lipids, and glucose. Compounds that alter endogenous FXR signaling can be used as therapeutic candidates or identified as potentially hazardous compounds depending on exposure doses and health states. Therefore, there is an increasing need for high-throughput screening assays of FXR activity to profile large numbers of environmental chemicals and drugs. This chapter describes a workflow of FXR modulator identification and characterization. To identify compounds that modulate FXR transactivation at the cellular level, we first screen compounds from the Tox21 10 K compound library in an FXR-driven beta-lactamase reporter gene assay multiplexed with a cell viability assay in the same well of the 1536-well plates. The selected compounds are then tested biochemically for their ability to modulate FXR-coactivator binding interactions using a time-resolved fluorescence resonance energy transfer (TR-FRET) coactivator assay. The assay results from the workflow can be used to prioritize compounds for more extensive investigations.

  18. 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

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

    PubMed

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

    2016-08-14

    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. Src-mediated cross-talk between farnesoid X and epidermal growth factor receptors inhibits human intestinal cell proliferation and tumorigenesis.

    PubMed

    Peng, Zhongsheng; Raufman, Jean-Pierre; Xie, Guofeng

    2012-01-01

    Besides its essential role in controlling bile acid and lipid metabolism, the farnesoid X receptor (FXR) protects against intestinal tumorigenesis by promoting apoptosis and inhibiting cell proliferation. However, the mechanisms underlying these anti-proliferative actions of FXR remain to be elucidated. In the present study, we examined the effects of FXR activation (FXR overexpression and treatment with an FXR agonist GW4064) and inactivation (treatment with FXR siRNA and an FXR antagonist guggulsterone) on colon cancer cell proliferation in vitro using human colon cancer cell lines (H508, SNU-C4 and HT-29) and in vivo using xenografts in nude mice. Blocking FXR activity with guggulsterone stimulated time- and dose-dependent EGFR (Tyr845) phosphorylation and ERK activation. In contrast, FXR overexpression and activation with GW4064 attenuated cell proliferation by down-regulating EGFR (Tyr845) phosphorylation and ERK activation. Treatment with guggulsterone and GW4064 also caused dose-dependent changes in Src (Tyr416) phosphorylation. In stably-transfected human colon cancer cells, overexpression of FXR reduced EGFR, ERK, Src phosphorylation and cell proliferation, and in nude mice attenuated the growth of human colon cancer xenografts (64% reduction in tumor volume; 47% reduction in tumor weight; both P<0.01). Moreover, guggulsterone-induced EGFR and ERK phosphorylation and cell proliferation were abolished by inhibiting activation of Src, EGFR and MEK. Collectively these data support the novel conclusion that in human colon cancer cells Src-mediated cross-talk between FXR and EGFR modulates ERK phosphorylation, thereby regulating intestinal cell proliferation and tumorigenesis.

  1. Farnesoid X Receptor Antagonizes JNK Signaling Pathway in Liver Carcinogenesis by Activating SOD3

    PubMed Central

    Li, Cunbao; Guo, Cong; Li, Yanyan; Qi, Hui; Shen, Hailing; Kong, Jing; Long, Xuecheng; Yuan, Frank; Wang, Xichun

    2015-01-01

    The farnesoid X receptor (FXR) is a key metabolic and homeostatic regulator in the liver. In the present work, we identify a novel role of FXR in antagonizing c-Jun N-terminal kinase (JNK) signaling pathway in liver carcinogenesis by activating superoxide dismutase 3 (SOD3) transcription. Compared with wild-type mouse liver, FXR−/− mouse liver showed elevated JNK phosphorylation. JNK1 deletion suppressed the increase of diethylnitrosamine-induced tumor number in FXR−/− mice. These results suggest that JNK1 plays a key role in chemical-induced liver carcinogenesis in FXR−/− mice. We found that ligand-activated FXR was able to alleviate H2O2 or tetradecanoylphorbol acetate-induced JNK phosphorylation in human hepatoblastoma (HepG2) cells or mouse primary hepatocytes. FXR ligand decreased H2O2-induced reactive oxygen species (ROS) levels in wild-type but not FXR−/− mouse hepatocytes. FXR knockdown abolished the inhibition of 3-[2-[2-chloro-4-[[3-(2,6-dichlorophenyl)-5-(1-methylethyl)-4-isoxazolyl]methoxy]phenyl]ethenyl]-Benzoic acid (GW4064) on JNK phosphorylation and ROS production induced by H2O2 in HepG2 cells. The gene expression of SOD3, an antioxidant defense enzyme, was increased by FXR activation in vitro and in vivo. An FXR-responsive element, inverted repeat separated by 1 nucleotide in SOD3 promoter, was identified by a combination of transcriptional reporter assays, EMSAs, and chromatin immunoprecipitation assays, which indicated that SOD3 could be a direct FXR target gene. SOD3 knockdown abolished the inhibition of GW4064 on JNK phosphorylation induced by H2O2 in HepG2 cells. In summary, FXR may regulate SOD3 expression to suppress ROS production, resulting in decreasing JNK activity. These results suggest that FXR, as a novel JNK suppressor, may be an attractive therapeutic target for liver cancer treatment. PMID:25496033

  2. Regulation of pyruvate dehydrogenase kinase expression by the farnesoid X receptor

    SciTech Connect

    Savkur, Rajesh S.; Bramlett, Kelli S.; Michael, Laura F.; Burris, Thomas P. . E-mail: burris_thomas_p@lilly.com

    2005-04-01

    The pyruvate dehydrogenase complex (PDC) functions as an important junction in intermediary metabolism by influencing the utilization of fat versus carbohydrate as a source of fuel. Activation of PDC is achieved by phosphatases, whereas, inactivation is catalyzed by pyruvate dehydrogenase kinases (PDKs). The expression of PDK4 is highly regulated by the glucocorticoid and peroxisome proliferator-activated receptors. We demonstrate that the farnesoid X receptor (FXR; NR1H4), which regulates a variety of genes involved in lipoprotein metabolism, also regulates the expression of PDK4. Treatment of rat hepatoma cells as well as human primary hepatocytes with FXR agonists stimulates the expression of PDK4 to levels comparable to those obtained with glucocorticoids. In addition, treatment of mice with an FXR agonist significantly increased hepatic PDK4 expression, while concomitantly decreasing plasma triglyceride levels. Thus, activation of FXR may suppress glycolysis and enhance oxidation of fatty acids via inactivation of the PDC by increasing PDK4 expression.

  3. Hepatoprotection by the farnesoid X receptor agonist GW4064 in rat models of intra- and extrahepatic cholestasis.

    PubMed

    Liu, Yaping; Binz, Jane; Numerick, Mary Jo; Dennis, Steve; Luo, Guizhen; Desai, Bhasha; MacKenzie, Kathleen I; Mansfield, Traci A; Kliewer, Steven A; Goodwin, Bryan; Jones, Stacey A

    2003-12-01

    Farnesoid X receptor (FXR) is a bile acid-activated transcription factor that is a member of the nuclear hormone receptor superfamily. Fxr-null mice exhibit a phenotype similar to Byler disease, an inherited cholestatic liver disorder. In the liver, activation of FXR induces transcription of transporter genes involved in promoting bile acid clearance and represses genes involved in bile acid biosynthesis. We investigated whether the synthetic FXR agonist GW4064 could protect against cholestatic liver damage in rat models of extrahepatic and intrahepatic cholestasis. In the bile duct-ligation and alpha-naphthylisothiocyanate models of cholestasis, GW4064 treatment resulted in significant reductions in serum alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase, as well as other markers of liver damage. Rats that received GW4064 treatment also had decreased incidence and extent of necrosis, decreased inflammatory cell infiltration, and decreased bile duct proliferation. Analysis of gene expression in livers from GW4064-treated cholestatic rats revealed decreased expression of bile acid biosynthetic genes and increased expression of genes involved in bile acid transport, including the phospholipid flippase MDR2. The hepatoprotection seen in these animal models by the synthetic FXR agonist suggests FXR agonists may be useful in the treatment of cholestatic liver disease.

  4. Upregulation of decorin by FXR in vascular smooth muscle cells

    SciTech Connect

    He Fengtian; Zhang Qiuhong; Kuruba, Ramalinga; Gao Xiang; Li Jiang; Li Yong; Gong Wei; Jiang, Yu; Xie Wen; Li Song

    2008-08-08

    Decorin is a member of the family of small leucine-rich proteoglycans that are present in blood vessels and synthesized by vascular smooth muscle cells (VSMCs). Decorin plays complex roles in both normal vascular physiology and the pathogenesis of various types of vascular disorders. However, the mechanisms of regulation of decorin expression in vasculature are not clearly understood. Particularly little information is available about a role of nuclear receptors in the regulation of decorin expression. In the present study, we report that activation of vascular FXR by a specific ligand resulted in upregulation of decorin at the levels of both mRNA and protein. FXR appears to induce decorin expression at a transcriptional level because (1) upregulation of decorin mRNA expression was abolished by the treatment of a transcription inhibitor, actinomycin D; and (2) decorin promoter activity was significantly increased by activation of FXR. Functional analysis of human decorin promoter identified an imperfect inverted repeat DNA motif, IR8 (-2313TGGTCAtagtgtcaTGACCT-2294), as a likely FXR-responsive element that is involved in decorin regulation.

  5. Bile acid signaling and liver regeneration.

    PubMed

    Fan, Mingjie; Wang, Xichun; Xu, Ganyu; Yan, Qingfeng; Huang, Wendong

    2015-02-01

    The liver is able to regenerate itself in response to partial hepatectomy or liver injury. This is accomplished by a complex network of different cell types and signals both inside and outside the liver. Bile acids (BAs) are recently identified as liver-specific metabolic signals and promote liver regeneration by activating their receptors: Farnesoid X Receptor (FXR) and G-protein-coupled BA receptor 1 (GPBAR1, or TGR5). FXR is a member of the nuclear hormone receptor superfamily of ligand-activated transcription factors. FXR promotes liver regeneration after 70% partial hepatectomy (PHx) or liver injury. Moreover, activation of FXR is able to alleviate age-related liver regeneration defects. Both liver- and intestine-FXR are activated by BAs after liver resection or injury and promote liver regeneration through distinct mechanism. TGR5 is a membrane-bound BA receptor and it is also activated during liver regeneration. TGR5 regulates BA hydrophobicity and stimulates BA excretion in urine during liver regeneration. BA signaling thus represents a novel metabolic pathway during liver regeneration. This article is part of a Special Issue entitled: Nuclear receptors in animal development.

  6. 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.

  7. 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

  8. Recent Progress on Bile Acid Receptor Modulators for Treatment of Metabolic Diseases.

    PubMed

    Xu, Yanping

    2016-07-28

    Bile acids are steroid-derived molecules synthesized in the liver, secreted from hepatocytes into the bile canaliculi, and subsequently stored in the gall bladder. During the feeding, bile flows into the duodenum, where it contributes to the solubilization and digestion of lipid-soluble nutrients. After a meal, bile-acid levels increase in the intestine, liver, and also in the systemic circulation. Therefore, serum bile-acid levels serve as an important sensing mechanism for nutrient and energy. Recent studies have described bile acids as versatile signaling molecules endowed with systemic endocrine functions. Bile acids are ligands for G-protein coupled receptors (GPCRs) such as TGR5 (also known as GPBAR1, M-BAR, and BG37) and nuclear hormone receptors including farnesoid X receptor (FXR; also known as NR1H4). Acting through these diverse signaling pathways, bile acids regulate triglyceride, cholesterol, glucose homeostasis, and energy expenditure. These bile-acid-controlled signaling pathways have become the source of promising novel drug targets to treat common metabolic and hepatic diseases. PMID:26878262

  9. Identification of chemerin as a novel FXR target gene down-regulated in the progression of nonalcoholic steatohepatitis.

    PubMed

    Deng, Yujie; Wang, Hui; Lu, Yan; Liu, Shuang; Zhang, Qiang; Huang, Jian; Zhu, Rongfeng; Yang, Jian; Zhang, Rong; Zhang, Di; Shen, Weili; Ning, Guang; Yang, Ying

    2013-05-01

    Chemerin is an adipokine involved in obesity, inflammation, and innate immune system that is highly expressed in the liver. In the present study, we find that chemerin mRNA expression is decreased in the livers of rodents with nonalcoholic fatty liver disease as well as in HepG2 cells after lipid overloading. Moreover, we report that chemerin expression and secretion are induced in HepG2 cells and primary hepatocytes from wild-type mice, but not farnesoid X receptor (FXR)-/- mice, in response to the synthetic FXR ligand GW4064. Hepatic chemerin expression is decreased in FXR-/- mice but up-regulated by GW4064 administration in wild-type mice. Dual-luciferase reporter assay and chromatin immunoprecipitation analyses further identified a functional FXR response element located in the -258-bp /+121-bp region of the chemerin gene. These data demonstrate that chemerin, a novel target gene of FXR, is related to nonalcoholic steatohepatitis.

  10. Inhibition of endothelin-1-mediated contraction of hepatic stellate cells by FXR ligand.

    PubMed

    Li, Jiang; Kuruba, Ramalinga; Wilson, Annette; Gao, Xiang; Zhang, Yifei; Li, Song

    2010-11-11

    Activation of hepatic stellate cells (HSCs) plays an important role in the development of cirrhosis through the increased production of collagen and the enhanced contractile response to vasoactive mediators such as endothelin-1 (ET-1). The farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily that is highly expressed in liver, kidneys, adrenals, and intestine. FXR is also expressed in HSCs and activation of FXR in HSCs is associated with significant decreases in collagen production. However, little is known about the roles of FXR in the regulation of contraction of HSCs. We report in this study that treatment of quiescent HSCs with GW4064, a synthetic FXR agonist, significantly inhibited the HSC transdifferentiation, which was associated with an inhibition of the upregulation of ET-1 expression. These GW4064-treated cells also showed reduced contractile response to ET-1 in comparison to HSCs without GW4064 treatment. We have further shown that GW4064 treatment inhibited the ET-1-mediated contraction in fully activated HSCs. To elucidate the potential mechanism we showed that GW4064 inhibited ET-1-mediated activation of Rho/ROCK pathway in activated HSCs. Our studies unveiled a new mechanism that might contribute to the anti-cirrhotic effects of FXR ligands.

  11. Bile acids are "homeotrophic" sensors of the functional hepatic capacity and regulate adaptive growth during liver regeneration.

    PubMed

    Geier, Andreas; Trautwein, Christian

    2007-01-01

    Liver mass depends on one or more unidentified humoral signals that drive regeneration when liver functional capacity is diminished. Bile acids are important liver products, and their levels are tightly regulated. Here, we identify a role for nuclear receptor-dependent bile acid signaling in normal liver regeneration. Elevated bile acid levels accelerate regeneration, and decreased levels inhibit liver regrowth, as does the absence of the primary nuclear bile acid receptor FXR. We propose that FXR activation by increased bile acid flux is a signal of decreased functional capacity of the liver. FXR, and possibly other nuclear receptors, may promote homeostasis not only by regulating expression of appropriate metabolic target genes but also by driving homeotrophic liver growth.

  12. 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.

  13. 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

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

    PubMed

    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

    2015-12-15

    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.

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

    PubMed

    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

  16. Unbinding pathways of GW4064 from human farnesoid X receptor as revealed by molecular dynamics simulations.

    PubMed

    Li, Weihua; Fu, Jing; Cheng, Feixiong; Zheng, Mingyue; Zhang, Jian; Liu, Guixia; Tang, Yun

    2012-11-26

    Farnesoid X receptor (FXR, NR1H4) is a member of a nuclear receptor superfamily, which plays important roles in bile acid homeostasis, lipoprotein and glucose metabolism, and hepatic regeneration. GW4064 is a potent and selective FXR agonist and has become a tool compound to probe the physiological functions of FXR. Until now, the mechanism of GW4064 entering and leaving the FXR pocket is still poorly understood. Here, we report a computational study of GW4064 unbinding pathways from FXR by using several molecular dynamics (MD) simulation techniques. Based on the crystal structure of FXR in complex with GW4064, conventional MD was first used to refine the binding and check the stability of GW4064 in the FXR pocket. Random acceleration MD simulations were then performed to explore the possible unbinding pathways of GW4064 from FXR. Four main pathway clusters were found, among which three subpathways, namely Paths 2A, 2B, and 1B, were observed most frequently. Multiple steered MD simulations were further employed to estimate the maximum rupture force and the sum of the forces and to characterize the intermediate states of the ligand unbinding process. By comparing the average force profiles and structural changes, Paths 2A and 2B were identified to be the most favorable unbinding pathways. The former is located between the H1-H2 loop and the H5-H6 loop, and the latter is located in the cleft formed by the H5-H6 loop, H6, and H7. Moreover, the residues lining the pathways were analyzed for their roles in ligand unbinding. Based on our results, the possible structural modification strategies on GW4064 were also proposed.

  17. FXR agonist GW4064 increases plasma glucocorticoid levels in C57BL/6 mice.

    PubMed

    Hoekstra, Menno; van der Sluis, Ronald J; Li, Zhaosha; Oosterveer, Maaike H; Groen, Albert K; Van Berkel, Theo J C

    2012-10-15

    Since high expression of farnesoid X receptor (FXR) has been detected in glucocorticoid-producing adrenocortical cells, we evaluated the potential role of FXR in adrenal glucocorticoid production. FXR agonist GW4064 increased fasting plasma corticosterone levels (+45%; P<0.01) in C57BL/6 mice, indicative of enhanced adrenal steroidogenesis. GW4064 treatment did not affect plasma ACTH levels, adrenal weight, or adrenal expression of steroidogenic genes. Scavenger receptor BI (SR-BI) mRNA and protein expression, respectively, increased 1.9-fold (P<0.01) and 1.5-fold, which suggests a stimulated lipoprotein-associated cholesterol uptake into the adrenals upon GW4064 treatment. In line with an enhanced flux of cellular cholesterol into the steroidogenic pathway, adrenal unesterified and esterified cholesterol stores were 21-41% decreased (P<0.01) upon GW4064 treatment. In conclusion, we have shown that the FXR agonist GW4064 stimulates plasma corticosterone levels in C57BL/6 mice. Our findings suggest a novel role for FXR in the modulation of adrenal cholesterol metabolism and glucocorticoid synthesis in mice.

  18. Farnesoid X receptor: a master regulator of hepatic triglyceride and glucose homeostasis

    PubMed Central

    Jiao, Yang; Lu, Yan; Li, Xiao-ying

    2015-01-01

    Non-alcoholic fatty liver disease (NAFLD) is characterized by the aberrant accumulation of triglycerides in hepatocytes in the absence of significant alcohol consumption, viral infection or other specific causes of liver disease. NAFLD has become a burgeoning health problem both worldwide and in China, but its pathogenesis remains poorly understood. Farnesoid X receptor (FXR), a member of the nuclear receptor (NR) superfamily, has been demonstrated to be the primary sensor for endogenous bile acids, and play a crucial role in hepatic triglyceride homeostasis. Deciphering the synergistic contributions of FXR to triglyceride metabolism is critical for discovering therapeutic agents in the treatment of NAFLD and hypertriglyceridemia. PMID:25500875

  19. SIRT1 controls liver regeneration by regulating BA metabolism through FXR and mTOR signaling

    PubMed Central

    García-Rodríguez, Juan L.; Barbier-Torres, Lucía; Fernández-Álvarez, Sara; Juan, Virginia Gutiérrez-de; Monte, María J.; Halilbasic, Emina; Herranz, Daniel; Álvarez, Luis; Aspichueta, Patricia; Marín, Jose J. G.; Trauner, Michael; Mato, Jose M.; Serrano, Manuel; Beraza, Naiara; Martínez-Chantar, María Luz

    2014-01-01

    Sirtuin1 (SIRT1) regulates central metabolic functions such as lipogenesis, protein synthesis, gluconeogenesis and bile acid (BA) homeostasis through deacetylation. Here, we describe that SIRT1 tightly controls the regenerative response of the liver. We performed partial hepatectomy (PH) to transgenic mice that overexpress SIRT1 (SIRT). SIRT mice showed increased mortality, impaired hepatocyte proliferation, BA accumulation and profuse liver injury after surgery. The damaging phenotype in SIRT mice correlated with impaired FXR activity due to persistent deacetylation and lower protein expression that led to decreased FXR-target gene expression; SHP, BSEP and increased Cyp7A1. Next, we convincingly show that 24-norUrsodeoxycholic acid (NorUDCA) attenuates SIRT protein expression, increases the acetylation of FXR and neighboring histones, restores trimethylation of H3K4 and H3K9 and increases miR34a expression, thus re-establishing BA homeostasis. Consequently, NorUDCA restored liver regeneration in SIRT mice, which showed increased survival and hepatocyte proliferation. Furthermore, a Leucine-enriched diet restored mTOR activation, acetylation of FXR and histones, leading to an overall lower BA production through SHP-inhibition of Cyp7A1 and higher transport (BSEP) and detoxification (Sult2a1) leading to an improved liver regeneration. Finally, we found that human HCC samples have increased presence of SIRT1, which correlated with absence of FXR suggesting its oncogenic potential. Conclusions Overall, we define SIRT1 as a key regulator of the regenerative response in the liver through post-transcriptional modifications that regulate the activity of FXR, histones and mTOR. Moreover, our data suggest that SIRT1 contributes to liver tumorigenesis through dysregulation of BA homeostasis by persistent FXR deacetylation. PMID:24338587

  20. The Farnesoid X Receptor Regulates Adipocyte Differentiation and Function by Promoting Peroxisome Proliferator-activated Receptor-γ and Interfering with the Wnt/β-Catenin Pathways*

    PubMed Central

    Abdelkarim, Mouaadh; Caron, Sandrine; Duhem, Christian; Prawitt, Janne; Dumont, Julie; Lucas, Anthony; Bouchaert, Emmanuel; Briand, Olivier; Brozek, John; Kuipers, Folkert; Fievet, Catherine; Cariou, Bertrand; Staels, Bart

    2010-01-01

    The bile acid receptor farnesoid X receptor (FXR) is expressed in adipose tissue, but its function remains poorly defined. Peroxisome proliferator-activated receptor-γ (PPARγ) is a master regulator of adipocyte differentiation and function. The aim of this study was to analyze the role of FXR in adipocyte function and to assess whether it modulates PPARγ action. Therefore, we tested the responsiveness of FXR-deficient mice (FXR−/−) and cells to the PPARγ activator rosiglitazone. Our results show that genetically obese FXR−/−/ob/ob mice displayed a resistance to rosiglitazone treatment. In vitro, rosiglitazone treatment did not induce normal adipocyte differentiation and lipid droplet formation in FXR−/− mouse embryonic fibroblasts (MEFs) and preadipocytes. Moreover, FXR−/− MEFs displayed both an increased lipolysis and a decreased de novo lipogenesis, resulting in reduced intracellular triglyceride content, even upon PPARγ activation. Retroviral-mediated FXR re-expression in FXR−/− MEFs restored the induction of adipogenic marker genes during rosiglitazone-forced adipocyte differentiation. The expression of Wnt/β-catenin pathway and target genes was increased in FXR−/− adipose tissue and MEFs. Moreover, the expression of several endogenous inhibitors of this pathway was decreased early during the adipocyte differentiation of FXR−/− MEFs. These findings demonstrate that FXR regulates adipocyte differentiation and function by regulating two counteracting pathways of adipocyte differentiation, the PPARγ and Wnt/β-catenin pathways. PMID:20851881

  1. Definition of a novel growth factor-dependent signal cascade for the suppression of bile acid biosynthesis.

    PubMed

    Holt, Jason A; Luo, Guizhen; Billin, Andrew N; Bisi, John; McNeill, Y Yvette; Kozarsky, Karen F; Donahee, Mary; Wang, Da Yuan; Mansfield, Traci A; Kliewer, Steven A; Goodwin, Bryan; Jones, Stacey A

    2003-07-01

    The nuclear bile acid receptor FXR has been proposed to play a central role in the feedback repression of the gene encoding cholesterol 7 alpha-hydroxylase (CYP7A1), the first and rate-limiting step in the biosynthesis of bile acids. We demonstrate that FXR directly regulates expression of fibroblast growth factor-19 (FGF-19), a secreted growth factor that signals through the FGFR4 cell-surface receptor tyrosine kinase. In turn, FGF-19 strongly suppresses expression of CYP7A1 in primary cultures of human hepatocytes and mouse liver through a c-Jun N-terminal kinase (JNK)-dependent pathway. This signaling cascade defines a novel mechanism for feedback repression of bile acid biosynthesis and underscores the vital role of FXR in the regulation of multiple pathways of cholesterol catabolism in the liver.

  2. 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.

  3. Reconstruction of FXR Beam Conditions

    SciTech Connect

    Nexen, W E; Scarpetti, R D; Zentler, J

    2001-05-31

    Beam-envelope radius, envelope angle, and beam emittance can be derived from measurements of beam radius for at least three different transport conditions. We have used this technique to reconstruct exit parameters from the FXR injector and accelerator. We use a diamagnetic loop (DML) to measure the magnetic moment of the high current beam. With no assumptions about radial profile, we can derive the beam mean squire radius from the moment under certain easily met conditions. Since it is this parameter which is required for the reconstruction, it is evident that the DML is the ideal diagnostic for this technique. The simplest application of this technique requires at least three shots for a reconstruction but in reality requires averaging over many more shots because of shot to shot variation. Since DML measurements do not interfere with the beam, single shot time resolved measurements of the beam parameters appear feasible if one uses an array of at least three DMLs separated by known transport conditions.

  4. Activation of farnesoid X receptor increases the expression of cytokine inducible SH2-containing protein in HepG2 cells.

    PubMed

    Xu, Zhizhen; Huang, Gang; Gong, Wei; Zhao, Yuanyin; Zhou, Peng; Zeng, Yijun; He, Fengtian

    2012-11-01

    Cytokine inducible SH2-containing protein (CISH), which negatively regulates cytokine signaling by inhibiting JAK2/STAT5 activity, is regarded as a therapeutic target for inflammatory diseases. Farnesoid X receptor (FXR), a ligand-activated transcription factor, has been proposed to play a protective function in the inflammatory responses. However, the role of FXR in modulation of CISH expression is unknown. In the present study, we for the first time identified that in human hepatoma cell line HepG2 the activation of FXR by the natural agonist chenodeoxycholic acid (CDCA) and the synthetic specific agonist GW4064 upregulated CISH at both transcriptional and translational levels, and inhibited interleukin (IL)6-induced STAT5 activation. Moreover, the in vivo experiment demonstrated that gavaging mice with CDCA increased CISH expression and reduced basal STAT5 phosphorylation in liver tissues. Reporter assay showed that FXR agonists enhanced the transcriptional activity of CISH promoter. These data suggest that FXR may serve as a novel molecular target for manipulating CISH expression in hepatocytes. FXR-mediated upregulation of CISH may play an important role in the homeostasis of cytokine signal networks and be beneficial to control cytokine-associated inflammatory diseases.

  5. 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

  6. A pathway involving farnesoid X receptor and small heterodimer partner positively regulates hepatic sirtuin 1 levels via microRNA-34a inhibition.

    PubMed

    Lee, Jiyoung; Padhye, Amruta; Sharma, Abhilasha; Song, Guisheng; Miao, Ji; Mo, Yin-Yuan; Wang, Li; Kemper, Jongsook Kim

    2010-04-23

    Sirtuin 1 (SIRT1) is a NAD-dependent deacetylase that is critically involved in diverse cellular processes including metabolic disease, cancer, and possibly aging. Despite extensive studies on SIRT1 function, how SIRT1 levels are regulated remains relatively unknown. Here, we report that the nuclear bile acid receptor farnesoid X receptor (FXR) inhibits microRNA-34a (miR-34a) in the liver, which results in a positive regulation of SIRT1 levels. Activation of FXR by the synthetic agonist GW4064 decreases hepatic miR-34a levels in normal mice, and consistently, hepatic miR-34a levels are elevated in FXR-null mice. FXR induces expression of small heterodimer partner (SHP), an orphan nuclear receptor and transcriptional corepressor, which in turn results in repression of p53, a key activator of the miR-34a gene, by inhibiting p53 occupancy at the promoter. MiR-34a decreased SIRT1 levels by binding to the 3'-untranslated region of SIRT1 mRNA, and adenovirus-mediated overexpression of miR-34a substantially decreased SIRT1 protein levels in mouse liver. Remarkably, miR-34a levels were elevated, and SIRT1 protein levels were reduced in diet-induced obese mice, and FXR activation in these mice reversed the miR-34a and SIRT1 levels, indicating an intriguing link among FXR activation, decreased miR-34a, and subsequently, increased SIRT1 levels. Our study demonstrates an unexpected role of the FXR/SHP pathway in controlling SIRT1 levels via miR-34a inhibition and that elevated miR-34a levels in obese mice contribute to decreased SIRT1 levels. Manipulation of this regulatory network may be useful for treating diseases of aging, such as metabolic disease and cancer.

  7. Dual Farnesoid X Receptor/TGR5 Agonist INT-767 Reduces Liver Injury in the Mdr2−/− (Abcb4−/−) Mouse Cholangiopathy Model by Promoting Biliary HCO3− Output

    PubMed Central

    Baghdasaryan, Anna; Claudel, Thierry; Gumhold, Judith; Silbert, Dagmar; Adorini, Luciano; Roda, Aldo; Vecchiotti, Stefania; Gonzalez, Frank J.; Schoonjans, Kristina; Strazzabosco, Mario; Fickert, Peter; Trauner, Michael

    2013-01-01

    Chronic cholangiopathies have limited therapeutic options and represent an important indication for liver transplantation. The nuclear farnesoid X receptor (FXR) and the membrane G protein-coupled receptor, TGR5, regulate bile acid (BA) homeostasis and inflammation. Therefore, we hypothesized that activation of FXR and/or TGR5 could ameliorate liver injury in Mdr2−/− (Abcb4−/−) mice, a model of chronic cholangiopathy. Hepatic inflammation, fibrosis, as well as bile secretion and key genes of BA homeostasis were addressed in Mdr2−/− mice fed either a chow diet or a diet supplemented with the FXR agonist, INT-747, the TGR5 agonist, INT-777, or the dual FXR/TGR5 agonist, INT-767 (0.03% w/w). Only the dual FXR/TGR5 agonist, INT-767, significantly improved serum liver enzymes, hepatic inflammation, and biliary fibrosis in Mdr2−/− mice, whereas INT-747 and INT-777 had no hepatoprotective effects. In line with this, INT-767 significantly induced bile flow and biliary HCO3− output, as well as gene expression of carbonic anhy-drase 14, an important enzyme able to enhance HCO3− transport, in an Fxr-dependent manner. In addition, INT-767 dramatically reduced bile acid synthesis via the induction of ileal Fgf15 and hepatic Shp gene expression, thus resulting in significantly reduced biliary bile acid output in Mdr2−/− mice. Conclusion This study shows that FXR activation improves liver injury in a mouse model of chronic cholangiopathy by reduction of biliary BA output and promotion of HCO3−-rich bile secretion. PMID:22006858

  8. Intestinal farnesoid X receptor signaling promotes nonalcoholic fatty liver disease

    PubMed Central

    Jiang, Changtao; Xie, Cen; Li, Fei; Zhang, Limin; Nichols, Robert G.; Krausz, Kristopher W.; Cai, Jingwei; Qi, Yunpeng; Fang, Zhong-Ze; Takahashi, Shogo; Tanaka, Naoki; Desai, Dhimant; Amin, Shantu G.; Albert, Istvan; Patterson, Andrew D.; Gonzalez, Frank J.

    2014-01-01

    Nonalcoholic fatty liver disease (NAFLD) is a major worldwide health problem. Recent studies suggest that the gut microbiota influences NAFLD pathogenesis. Here, a murine model of high-fat diet–induced (HFD-induced) NAFLD was used, and the effects of alterations in the gut microbiota on NAFLD were determined. Mice treated with antibiotics or tempol exhibited altered bile acid composition, with a notable increase in conjugated bile acid metabolites that inhibited intestinal farnesoid X receptor (FXR) signaling. Compared with control mice, animals with intestine-specific Fxr disruption had reduced hepatic triglyceride accumulation in response to a HFD. The decrease in hepatic triglyceride accumulation was mainly due to fewer circulating ceramides, which was in part the result of lower expression of ceramide synthesis genes. The reduction of ceramide levels in the ileum and serum in tempol- or antibiotic-treated mice fed a HFD resulted in downregulation of hepatic SREBP1C and decreased de novo lipogenesis. Administration of C16:0 ceramide to antibiotic-treated mice fed a HFD reversed hepatic steatosis. These studies demonstrate that inhibition of an intestinal FXR/ceramide axis mediates gut microbiota–associated NAFLD development, linking the microbiome, nuclear receptor signaling, and NAFLD. This work suggests that inhibition of intestinal FXR is a potential therapeutic target for NAFLD treatment. PMID:25500885

  9. Individual bile acids have differential effects on bile acid signaling in mice.

    PubMed

    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

  10. MAFG is a transcriptional repressor of bile acid synthesis and metabolism.

    PubMed

    de Aguiar Vallim, Thomas Q; Tarling, Elizabeth J; Ahn, Hannah; Hagey, Lee R; Romanoski, Casey E; Lee, Richard G; Graham, Mark J; Motohashi, Hozumi; Yamamoto, Masayuki; Edwards, Peter A

    2015-02-01

    Specific bile acids are potent signaling molecules that modulate metabolic pathways affecting lipid, glucose and bile acid homeostasis, and the microbiota. Bile acids are synthesized from cholesterol in the liver, and the key enzymes involved in bile acid synthesis (Cyp7a1, Cyp8b1) are regulated transcriptionally by the nuclear receptor FXR. We have identified an FXR-regulated pathway upstream of a transcriptional repressor that controls multiple bile acid metabolism genes. We identify MafG as an FXR target gene and show that hepatic MAFG overexpression represses genes of the bile acid synthetic pathway and modifies the biliary bile acid composition. In contrast, loss-of-function studies using MafG(+/-) mice causes de-repression of the same genes with concordant changes in biliary bile acid levels. Finally, we identify functional MafG response elements in bile acid metabolism genes using ChIP-seq analysis. Our studies identify a molecular mechanism for the complex feedback regulation of bile acid synthesis controlled by FXR.

  11. MAFG Is a Transcriptional Repressor of Bile Acid Synthesis and Metabolism

    PubMed Central

    de Aguiar Vallim, Thomas Q.; Tarling, Elizabeth J.; Ahn, Hannah; Hagey, Lee R.; Romanoski, Casey E.; Lee, Richard G.; Graham, Mark J.; Motohashi, Hozumi; Yamamoto, Masayuki; Edwards, Peter A.

    2015-01-01

    Summary Specific bile acids are potent signaling molecules that modulate metabolic pathways affecting lipid, glucose and bile acid homeostasis and the microbiota. Bile acids are synthesized from cholesterol in the liver, and the key enzymes involved in bile acid synthesis (Cyp7a1, Cyp8b1) are regulated transcriptionally by the nuclear receptor FXR. We have identified an FXR-regulated pathway upstream of a transcriptional repressor that controls multiple bile acid metabolism genes. We identify MafG as an FXR target gene and show that hepatic MAFG overexpression represses genes of the bile acid synthetic pathway, and modifies the biliary bile acid composition. In contrast, loss-of-function studies using MafG+/− mice causes de-repression of the same genes with concordant changes in biliary bile acid levels. Finally, we identify functional MafG response elements in bile acid metabolism genes using ChIP-Seq analysis. Our studies identify a molecular mechanism for the complex feedback regulation of bile acid synthesis controlled by FXR. PMID:25651182

  12. Effects of essential fatty acid deficiency on enterohepatic circulation of bile salts in mice.

    PubMed

    Lukovac, S; Los, E L; Stellaard, F; Rings, E H H M; Verkade, H J

    2009-09-01

    Essential fatty acid (EFA) deficiency in mice has been associated with increased bile production, which is mainly determined by the enterohepatic circulation (EHC) of bile salts. To establish the mechanism underlying the increased bile production, we characterized in detail the EHC of bile salts in EFA-deficient mice using stable isotope technique, without interrupting the normal EHC. Farnesoid X receptor (FXR) has been proposed as an important regulator of bile salt synthesis and homeostasis. In Fxr(-/-) mice we additionally investigated to what extent alterations in bile production during EFA deficiency were FXR dependent. Furthermore, we tested in differentiating Caco-2 cells the effects of EFA deficiency on expression of FXR-target genes relevant for feedback regulation of bile salt synthesis. EFA deficiency-enhanced bile flow and biliary bile salt secretion were associated with elevated bile salt pool size and synthesis rate (+146 and +42%, respectively, P < 0.05), despite increased ileal bile salt reabsorption (+228%, P < 0.05). Cyp7a1 mRNA expression was unaffected in EFA-deficient mice. However, ileal mRNA expression of Fgf15 (inhibitor of bile salt synthesis) was significantly reduced, in agreement with absent inhibition of the hepatic bile salt synthesis. Bile flow and biliary secretion were enhanced to the same extent in EFA-deficient wild-type and Fxr(-/-) mice, indicating contribution of other factors besides FXR in regulation of EHC during EFA deficiency. In vitro experiments show reduced induction of mRNA expression of relevant genes upon chenodeoxycholic acid and a selective FXR agonist GW4064 stimulation in EFA-deficient Caco-2 cells. In conclusion, our data indicate that EFA deficiency is associated with interrupted negative feedback of bile salt synthesis, possibly because of reduced ileal Fgf15 expression.

  13. Estrogen and Estrogen Receptor-α-Mediated Transrepression of Bile Salt Export Pump

    PubMed Central

    Chen, Yuan; Vasilenko, Alex; Song, Xiulong; Valanejad, Leila; Verma, Ruchi; You, Sangmin; Yan, Bingfang; Shiffka, Stephanie; Hargreaves, Leeza; Nadolny, Christina

    2015-01-01

    Among diseases unique to pregnancy, intrahepatic cholestasis of pregnancy is the most prevalent disorder with elevated serum bile acid levels. We have previously shown that estrogen 17β-estradiol (E2) transrepresses bile salt export pump (BSEP) through an interaction between estrogen receptor (ER)-α and farnesoid X receptor (FXR) and transrepression of BSEP by E2/ERα is an etiological contributing factor to intrahepatic cholestasis of pregnancy. Currently the mechanistic insights into such transrepression are not fully understood. In this study, the dynamics of coregulator recruitment to BSEP promoter after FXR activation and E2 treatment were established with quantitative chromatin immunoprecipitation assays. Coactivator peroxisome proliferator-activated receptor-γ coactivator-1 was predominantly recruited to the BSEP promoter upon FXR activation, and its recruitment was decreased by E2 treatment. Meanwhile, recruitment of nuclear receptor corepressor was markedly increased upon E2 treatment. Functional evaluation of ERα and ERβ chimeras revealed that domains AC of ERα are the determinants for ERα-specific transrepression on BSEP. Further studies with various truncated ERα proteins identified the domains in ERα responsible for ligand-dependent and ligand-independent transrepression. Truncated ERα-AD exhibited potent ligand-independent transrepressive activity, whereas ERα-CF was fully capable of transrepressing BSEP ligand dependently in vitro in Huh 7 cells and in vivo in mice. Both ERα-AD and ERα-CF proteins were associated with FXR in the coimmunoprecipitation assays. In conclusion, E2 repressed BSEP expression through diminishing peroxisome proliferator-activated receptor-γ coactivator-1 recruitment with a concurrent increase in nuclear receptor corepressor recruitment to the BSEP promoter. Domains AD and CF in ERα mediated ligand-independent and ligand-dependent transrepression on BSEP, respectively, through interacting with FXR. PMID:25675114

  14. 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

  15. GW4064, an agonist of farnesoid X receptor, represses CYP3A4 expression in human hepatocytes by inducing small heterodimer partner expression.

    PubMed

    Zhang, Shu; Pan, Xian; Jeong, Hyunyoung

    2015-05-01

    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.

  16. Effects of alfalfa saponin extract on mRNA expression of Ldlr, LXRα, and FXR in BRL cells*

    PubMed Central

    Liang, Xin-ping; Zhang, Dong-qiang; Chen, Yan-yan; Guo, Rui; Wang, Jie; Wang, Cheng-zhang; Shi, Ying-hua

    2015-01-01

    We studied the effects of alfalfa saponin extract (ASE) on low density lipoprotein receptor (Ldlr), liver X receptor α (LXRα), and farnesoid X receptor (FXR) in normal and hyperlipidemic Buffalo rat liver (BRL) cells. Normal and hyperlipidemic BRL cells were divided into eight groups: normal, or normal cells treated with 50, 100, and 150 mg/L ASE, hyperlipidemic, or hyperlipidemic cells treated with 50, 100, and 150 mg/L ASE. After treatment for 24 h, Ldlr, LXRα, and FXR mRNA expression levels were measured by quantitative real-time polymerase chain reaction (qRT-PCR). Data showed that mRNA expression of Ldlr in normal BRL cells was significantly up-regulated by ASE treatment and mRNA expressions of LXRα and FXR were significantly down-regulated both in normal and hyperlipidemic BRL cells after ASE treatment. Thus, ASE might ameliorate hepatic steatosis by regulating genes involved in cholesterol metabolism, including up-regulation of Ldlr as well as down-regulation of LXRα and FXR. PMID:26055909

  17. Interactions of methoxyacetic acid with androgen receptor

    SciTech Connect

    Bagchi, Gargi; Hurst, Christopher H.; Waxman, David J.

    2009-07-15

    Endocrine disruptive compounds (EDC) alter hormone-stimulated, nuclear receptor-dependent physiological and developmental processes by a variety of mechanisms. One recently identified mode of endocrine disruption is through hormone sensitization, where the EDC modulates intracellular signaling pathways that control nuclear receptor function, thereby regulating receptor transcriptional activity indirectly. Methoxyacetic acid (MAA), the primary, active metabolite of the industrial solvent ethylene glycol monomethyl ether and a testicular toxicant, belongs to this EDC class. Modulation of nuclear receptor activity by MAA could contribute to the testicular toxicity associated with MAA exposure. In the present study, we evaluated the impact of MAA on the transcriptional activity of several nuclear receptors including the androgen receptor (AR), which plays a pivotal role in the development and maturation of spermatocytes. AR transcriptional activity is shown to be increased by MAA through a tyrosine kinase signaling pathway that involves PI3-kinase. In a combinatorial setting with AR antagonists, MAA potentiated the AR response without significantly altering the EC{sub 50} for androgen responsiveness, partially alleviating the antagonistic effect of the anti-androgens. Finally, MAA treatment of TM3 mouse testicular Leydig cells markedly increased the expression of Cyp17a1 and Shbg while suppressing Igfbp3 expression by {approx} 90%. Deregulation of these genes may alter androgen synthesis and action in a manner that contributes to MAA-induced testicular toxicity.

  18. 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

  19. Specific sequences in the fragile X syndrome protein FMR1 and the FXR proteins mediate their binding to 60S ribosomal subunits and the interactions among them.

    PubMed Central

    Siomi, M C; Zhang, Y; Siomi, H; Dreyfuss, G

    1996-01-01

    Fragile X syndrome, the most common form of hereditary mental retardation, usually results from lack of expression of the FMR1 gene. The FMR1 protein is a cytoplasmic RNA-binding protein. The RNA-binding activity of FMR1 is an essential feature of FMR1, as fragile X syndrome can also result from the expression of mutant FMR1 protein that is impaired in RNA binding. Recently, we described two novel cytoplasmic proteins, FXR1 and FXR2, which are both very similar in amino acid sequence to FMR1 and which also interact strongly with FMR1 and with each other. To understand the function of FMR1 and the FXR proteins, we carried out cell fractionation and sedimentation experiments with monoclonal antibodies to these proteins to characterize the complexes they form. Here, we report that the FMR1 and FXR proteins are associated with ribosomes, predominantly with 60S large ribosomal subunits. The FXR proteins are associated with 60S ribosomal subunits even in cells that lack FMR1 and that are derived from a fragile X syndrome patient, indicating that FMR1 is not required for this association. We delineated the regions of FMR1 that mediate its binding to 60S ribosomal subunits and the interactions among the FMR1-FXR family members. Both regions contain sequences predicted to have a high propensity to form coiled coil interactions, and the sequences are highly evolutionarily conserved in this protein family. The association of the FMR1, FXR1, and FXR2 proteins with ribosomes suggests they have functions in translation or mRNA stability. PMID:8668200

  20. Fragmentation of GW4064 led to a highly potent partial farnesoid X receptor agonist with improved drug-like properties.

    PubMed

    Flesch, Daniel; Gabler, Matthias; Lill, Andreas; Gomez, Roberto Carrasco; Steri, Ramona; Schneider, Gisbert; Stark, Holger; Schubert-Zsilavecz, Manfred; Merk, Daniel

    2015-07-01

    The ligand activated transcription factor farnesoid X receptor (FXR) is a crucial regulator of several metabolic and inflammatory pathways and its activation by agonistic ligands seems a valuable therapeutic approach for many disorders. Most known non-steroidal FXR agonists however, have limitations that hinder their clinical development and novel FXR ligands are required. Evaluation of the co-crystal structures of the widely used FXR agonist GW4064 and related compounds in complex with the FXR ligand binding domain indicated that their disubstituted isoxazole moiety is especially relevant for FXR activation. By investigation of GW4064-fragments missing the aromatic tail, we discovered a highly potent and soluble partial FXR agonist (14, ST-1892) as well as a fluorescent FXR ligand (15) as potential pharmacological tool.

  1. FXR antagonism of NSAIDs contributes to drug-induced liver injury identified by systems pharmacology approach

    PubMed Central

    Lu, Weiqiang; Cheng, Feixiong; Jiang, Jing; Zhang, Chen; Deng, Xiaokang; Xu, Zhongyu; Zou, Shien; Shen, Xu; Tang, Yun; Huang, Jin

    2015-01-01

    Non-steroidal anti-inflammatory drugs (NSAIDs) are worldwide used drugs for analgesic, antipyretic, and anti-inflammatory therapeutics. However, NSAIDs often cause several serious liver injuries, such as drug-induced liver injury (DILI), and the molecular mechanisms of DILI have not been clearly elucidated. In this study, we developed a systems pharmacology approach to explore the mechanism-of-action of NSAIDs. We found that the Farnesoid X Receptor (FXR) antagonism of NSAIDs is a potential molecular mechanism of DILI through systematic network analysis and in vitro assays. Specially, the quantitative real-time PCR assay reveals that indomethacin and ibuprofen regulate FXR downstream target gene expression in HepG2 cells. Furthermore, the western blot shows that FXR antagonism by indomethacin induces the phosphorylation of STAT3 (signal transducer and activator of transcription 3), promotes the activation of caspase9, and finally causes DILI. In summary, our systems pharmacology approach provided novel insights into molecular mechanisms of DILI for NSAIDs, which may propel the ways toward the design of novel anti-inflammatory pharmacotherapeutics. PMID:25631039

  2. Glucose-lowering effects of intestinal bile acid sequestration through enhancement of splanchnic glucose utilization.

    PubMed

    Prawitt, Janne; Caron, Sandrine; Staels, Bart

    2014-05-01

    Intestinal bile acid (BA) sequestration efficiently lowers plasma glucose concentrations in type 2 diabetes (T2D) patients. Because BAs act as signaling molecules via receptors, including the G protein-coupled receptor TGR5 and the nuclear receptor FXR (farnesoid X receptor), to regulate glucose homeostasis, BA sequestration, which interrupts the entero-hepatic circulation of BAs, constitutes a plausible action mechanism of BA sequestrants. An increase of intestinal L-cell glucagon-like peptide-1 (GLP-1) secretion upon TGR5 activation is the most commonly proposed mechanism, but recent studies also argue for a direct entero-hepatic action to enhance glucose utilization. We discuss here recent findings on the mechanisms of sequestrant-mediated glucose lowering via an increase of splanchnic glucose utilization through entero-hepatic FXR signaling.

  3. Silymarin Ameliorates Metabolic Dysfunction Associated with Diet-Induced Obesity via Activation of Farnesyl X Receptor

    PubMed Central

    Gu, Ming; Zhao, Ping; Huang, Jinwen; Zhao, Yuanyuan; Wang, Yahui; Li, Yin; Li, Yifei; Fan, Shengjie; Ma, Yue-Ming; Tong, Qingchun; Yang, Li; Ji, Guang; Huang, Cheng

    2016-01-01

    Background and purpose: Silymarin, a standardized extract of the milk thistle seeds, has been widely used to treat chronic hepatitis, cirrhosis, and other types of toxic liver damage. Despite increasing studies on the action of silymarin and its major active constituent, silybin in their therapeutic properties against insulin resistance, diabetes and hyperlipidaemia in vitro and in vivo, the mechanism underlying silymarin action remains unclear. Experimental approach: C57BL/6 mice were fed high-fat diet (HFD) for 3 months to induce obesity, insulin resistance, hyperlipidaemia, and fatty liver. These mice were then continuously treated with HFD alone or mixed with silymarin at 40 mg/100 g for additional 6 weeks. Biochemical analysis was used to test the serum lipid and bile acid profiles. Farnesyl X receptor (FXR) and nuclear factor kappa B (NF-κB) transactivities were analyzed in liver using a gene reporter assay based on quantitative RT-PCR. Key results: Silymarin treatment ameliorated insulin resistance, dyslipidaemia and inflammation, and reconstituted the bile acid pool in liver of diet-induced obesity. Associated with this, silybin and silymarin enhanced FXR transactivity. Consistently, in HepG2 cells, silybin inhibited NF-κB signaling, which was enhanced by FXR activation. Conclusion and implications: Our results suggest that silybin is an effective component of silymarin for treating metabolic syndrome by stimulating FXR signaling. PMID:27733832

  4. Activation of the farnesoid X receptor represses PCSK9 expression in human hepatocytes.

    PubMed

    Langhi, Cédric; Le May, Cédric; Kourimate, Sanae; Caron, Sandrine; Staels, Bart; Krempf, Michel; Costet, Philippe; Cariou, Bertrand

    2008-03-19

    The purpose of this study was to determine whether bile acids (BAs) modulate hepatic pro-protein convertase subtilisin/kexin 9 (PCSK9) gene expression. Immortalized human hepatocytes were treated with various BAs. Chenodeoxycholic acid (CDCA) treatment specifically decreased both PCSK9 mRNA and protein contents. Moreover, activation of the BA-activated farnesoid X receptor (FXR) by its synthetic specific agonist GW4064 also decreased PCSK9 expression. Of functional relevance, coadministration of CDCA counteracted the statin-induced PCSK9 expression, leading to a potentiation of LDL receptor activity. This study suggests that a transcriptional repression of PCSK9 by CDCA or FXR agonists may potentiate the hypolipidemic effect of statins.

  5. Activated FXR Inhibits Leptin Signaling and Counteracts Tumor-promoting Activities of Cancer-Associated Fibroblasts in Breast Malignancy.

    PubMed

    Giordano, Cinzia; Barone, Ines; Vircillo, Valentina; Panza, Salvatore; Malivindi, Rocco; Gelsomino, Luca; Pellegrino, Michele; Rago, Vittoria; Mauro, Loredana; Lanzino, Marilena; Panno, Maria Luisa; Bonofiglio, Daniela; Catalano, Stefania; Andò, Sebastiano

    2016-02-22

    Cancer-associated fibroblasts (CAFs), the principal components of the tumor stroma, play a central role in cancer development and progression. As an important regulator of the crosstalk between breast cancer cells and CAFs, the cytokine leptin has been associated to breast carcinogenesis. The nuclear Farnesoid X Receptor-(FXR) seems to exert an oncosuppressive role in different tumors, including breast cancer. Herein, we demonstrated, for the first time, that the synthetic FXR agonist GW4064, inhibiting leptin signaling, affects the tumor-promoting activities of CAFs in breast malignancy. GW4064 inhibited growth, motility and invasiveness induced by leptin as well as by CAF-conditioned media in different breast cancer cell lines. These effects rely on the ability of activated FXR to increase the expression of the suppressor of the cytokine signaling 3 (SOCS3) leading to inhibition of leptin-activated signaling and downregulation of leptin-target genes. In vivo xenograft studies, using MCF-7 cells alone or co-injected with CAFs, showed that GW4064 administration markedly reduced tumor growth. Interestingly, GW4064-treated tumors exhibited decreased levels of leptin-regulated proteins along with a strong staining intensity for SOCS3. Thus, FXR ligands might represent an emerging potential anti-cancer therapy able to block the tumor supportive role of activated fibroblasts within the breast microenvironment.

  6. Activated FXR Inhibits Leptin Signaling and Counteracts Tumor-promoting Activities of Cancer-Associated Fibroblasts in Breast Malignancy

    PubMed Central

    Giordano, Cinzia; Barone, Ines; Vircillo, Valentina; Panza, Salvatore; Malivindi, Rocco; Gelsomino, Luca; Pellegrino, Michele; Rago, Vittoria; Mauro, Loredana; Lanzino, Marilena; Panno, Maria Luisa; Bonofiglio, Daniela; Catalano, Stefania; Andò, Sebastiano

    2016-01-01

    Cancer-associated fibroblasts (CAFs), the principal components of the tumor stroma, play a central role in cancer development and progression. As an important regulator of the crosstalk between breast cancer cells and CAFs, the cytokine leptin has been associated to breast carcinogenesis. The nuclear Farnesoid X Receptor-(FXR) seems to exert an oncosuppressive role in different tumors, including breast cancer. Herein, we demonstrated, for the first time, that the synthetic FXR agonist GW4064, inhibiting leptin signaling, affects the tumor-promoting activities of CAFs in breast malignancy. GW4064 inhibited growth, motility and invasiveness induced by leptin as well as by CAF-conditioned media in different breast cancer cell lines. These effects rely on the ability of activated FXR to increase the expression of the suppressor of the cytokine signaling 3 (SOCS3) leading to inhibition of leptin-activated signaling and downregulation of leptin-target genes. In vivo xenograft studies, using MCF-7 cells alone or co-injected with CAFs, showed that GW4064 administration markedly reduced tumor growth. Interestingly, GW4064-treated tumors exhibited decreased levels of leptin-regulated proteins along with a strong staining intensity for SOCS3. Thus, FXR ligands might represent an emerging potential anti-cancer therapy able to block the tumor supportive role of activated fibroblasts within the breast microenvironment. PMID:26899873

  7. Nutritional Signaling via Free Fatty Acid Receptors.

    PubMed

    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

  8. 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

  9. Nuclear receptors and cholesterol metabolism in the intestine.

    PubMed

    Moschetta, Antonio

    2015-02-01

    Nuclear receptors are involved in many important function and mediate signaling by factors including hormones, vitamins and a number of endogenous ligands and xenobiotics, several of which are involved in lipid metabolism. This review focuses on the liver X receptor (LXR), which is an important regulator of whole-body cholesterol, fatty acid, and glucose homeostasis that binds to LXR response elements as a heterodimer with retinoid X receptors, and the farnesoid X receptor (FXR), which is a bile acid receptor involved in feedback inhibition of bile acid synthesis, and thus cholesterol catabolism. These nuclear receptors regulate gene programs that control intestinal and hepatic lipid homeostasis through their effects on cholesterol transport and catabolism.

  10. FXR agonist activity of conformationally constrained analogs of GW 4064

    SciTech Connect

    Akwabi-Ameyaw, Adwoa; Bass, Jonathan Y.; Caldwell, Richard D.; Caravella, Justin A.; Chen, Lihong; Creech, Katrina L.; Deaton, David N.; Madauss, Kevin P.; Marr, Harry B.; McFadyen, Robert B.; Miller, Aaron B.; Navas, III, Frank; Parks, Derek J.; Spearing, Paul K.; Todd, Dan; Williams, Shawn P.; Wisely, G. Bruce

    2010-09-27

    Two series of conformationally constrained analogs of the FXR agonist GW 4064 1 were prepared. Replacement of the metabolically labile stilbene with either benzothiophene or naphthalene rings led to the identification of potent full agonists 2a and 2g.

  11. FXR agonist activity of conformationally constrained analogs of GW 4064.

    PubMed

    Akwabi-Ameyaw, Adwoa; Bass, Jonathan Y; Caldwell, Richard D; Caravella, Justin A; Chen, Lihong; Creech, Katrina L; Deaton, David N; Madauss, Kevin P; Marr, Harry B; McFadyen, Robert B; Miller, Aaron B; Navas, Frank; Parks, Derek J; Spearing, Paul K; Todd, Dan; Williams, Shawn P; Bruce Wisely, G

    2009-08-15

    Two series of conformationally constrained analogs of the FXR agonist GW 4064 1 were prepared. Replacement of the metabolically labile stilbene with either benzothiophene or naphthalene rings led to the identification of potent full agonists 2a and 2g.

  12. Chenodeoxycholic Acid Reduces Hypoxia Inducible Factor-1α Protein and Its Target Genes.

    PubMed

    Moon, Yunwon; Choi, Su Mi; Chang, Soojeong; Park, Bongju; Lee, Seongyeol; Lee, Mi-Ock; Choi, Hueng-Sik; Park, Hyunsung

    2015-01-01

    This study evaluated HIF-1α inhibitors under different hypoxic conditions, physiological hypoxia (5% O2) and severe hypoxia (0.1% O2). We found that chenodeoxy cholic acid (CDCA) reduced the amount of HIF-1α protein only under physiological hypoxia but not under severe hypoxia without decreasing its mRNA level. By using a proteasome inhibitor MG132 and a translation inhibitor cyclohexamide, we showed that CDCA reduced HIF-1α protein by decreasing its translation but not by enhancing its degradation. The following findings indicated that farnesoid X receptor (FXR), a CDCA receptor and its target gene, Small heterodimer partner (SHP) are not involved in this effect of CDCA. Distinctly from CDCA, MG132 prevented SHP and an exogenous FXR agonist, GW4064 from reducing HIF-1α protein. Furthermore a FXR antagonist, guggulsterone failed to prevent CDCA from decreasing HIF-1α protein. Furthermore, guggulsterone by itself reduced HIF-1α protein even in the presence of MG132. These findings suggested that CDCA and guggulsterone reduced the translation of HIF-1α in a mechanism which FXR and SHP are not involved. This study reveals novel therapeutic functions of traditional nontoxic drugs, CDCA and guggulsterone, as inhibitors of HIF-1α protein.

  13. Chenodeoxycholic Acid Reduces Hypoxia Inducible Factor-1α Protein and Its Target Genes

    PubMed Central

    Moon, Yunwon; Choi, Su Mi; Chang, Soojeong; Park, Bongju; Lee, Seongyeol; Lee, Mi-Ock; Choi, Hueng-Sik; Park, Hyunsung

    2015-01-01

    This study evaluated HIF-1α inhibitors under different hypoxic conditions, physiological hypoxia (5% O2) and severe hypoxia (0.1% O2). We found that chenodeoxy cholic acid (CDCA) reduced the amount of HIF-1α protein only under physiological hypoxia but not under severe hypoxia without decreasing its mRNA level. By using a proteasome inhibitor MG132 and a translation inhibitor cyclohexamide, we showed that CDCA reduced HIF-1α protein by decreasing its translation but not by enhancing its degradation. The following findings indicated that farnesoid X receptor (FXR), a CDCA receptor and its target gene, Small heterodimer partner (SHP) are not involved in this effect of CDCA. Distinctly from CDCA, MG132 prevented SHP and an exogenous FXR agonist, GW4064 from reducing HIF-1α protein. Furthermore a FXR antagonist, guggulsterone failed to prevent CDCA from decreasing HIF-1α protein. Furthermore, guggulsterone by itself reduced HIF-1α protein even in the presence of MG132. These findings suggested that CDCA and guggulsterone reduced the translation of HIF-1α in a mechanism which FXR and SHP are not involved. This study reveals novel therapeutic functions of traditional nontoxic drugs, CDCA and guggulsterone, as inhibitors of HIF-1α protein. PMID:26098428

  14. 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

  15. Pharmacology of bile acid receptors: Evolution of bile acids from simple detergents to complex signaling molecules.

    PubMed

    Copple, Bryan L; Li, Tiangang

    2016-02-01

    For many years, bile acids were thought to only function as detergents which solubilize fats and facilitate the uptake of fat-soluble vitamins in the intestine. Many early observations; however, demonstrated that bile acids regulate more complex processes, such as bile acids synthesis and immune cell function through activation of signal transduction pathways. These studies were the first to suggest that receptors may exist for bile acids. Ultimately, seminal studies by many investigators led to the discovery of several bile acid-activated receptors including the farnesoid X receptor, the vitamin D receptor, the pregnane X receptor, TGR5, α5 β1 integrin, and sphingosine-1-phosphate receptor 2. Several of these receptors are expressed outside of the gastrointestinal system, indicating that bile acids may have diverse functions throughout the body. Characterization of the functions of these receptors over the last two decades has identified many important roles for these receptors in regulation of bile acid synthesis, transport, and detoxification; regulation of glucose utilization; regulation of fatty acid synthesis and oxidation; regulation of immune cell function; regulation of energy expenditure; and regulation of neural processes such as gastric motility. Through these many functions, bile acids regulate many aspects of digestion ranging from uptake of essential vitamins to proper utilization of nutrients. Accordingly, within a short time period, bile acids moved beyond simple detergents and into the realm of complex signaling molecules. Because of the important processes that bile acids regulate through activation of receptors, drugs that target these receptors are under development for the treatment of several diseases, including cholestatic liver disease and metabolic syndrome. In this review, we will describe the various bile acid receptors, the signal transduction pathways activated by these receptors, and briefly discuss the physiological processes that

  16. Ménage-à-trois of bariatric surgery, bile acids and the gut microbiome.

    PubMed

    Raghow, Rajendra

    2015-04-15

    Bariatric surgeries have emerged as highly effective treatments for obesity associated type-2 diabetes mellitus. Evidently, the desired therapeutic endpoints such as rates of weight loss, lower levels of glycated hemoglobin and remission of diabetes are achieved more rapidly and last longer following bariatric surgery, as opposed to drug therapies alone. In light of these findings, it has been suspected that in addition to causing weight loss dependent glucose intolerance, bariatric surgery induces other physiological changes that contribute to the alleviation of diabetes. However, the putative post-surgical neuro-hormonal pathways that underpin the therapeutic benefits of bariatric surgery remain undefined. In a recent report, Ryan and colleagues shed new light on the potential mechanisms that determine the salutary effects of bariatric surgery in mice. The authors demonstrated that the improved glucose tolerance and weight loss in mice after vertical sleeve gastrectomy (VSG) surgery were likely to be caused by post-surgical changes in circulating bile acids and farnesoid-X receptor (FXR) signaling, both of which were also mechanistically linked to changes in the microbial ecology of the gut. The authors arrived at this conclusion from a comparison of genome-wide, metabolic consequences of VSG surgery in obese wild type (WT) and FXR knockout mice. Gene expression in the distal small intestines of WT and FXR knockout mice revealed that the pathways regulating bile acid composition, nutrient metabolism and anti-oxidant defense were differentially altered by VSG surgery in WT and FXR(-/-) mice. Based on these data Ryan et al, hypothesized that bile acid homeostasis and FXR signaling were mechanistically linked to the gut microbiota that played a role in modulating post-surgical changes in total body mass and glucose tolerance. The authors' data provide a plausible explanation for putative weight loss-independent benefits of bariatric surgery and its relationship with

  17. Anatomical and pharmacological characterization of excitatory amino acid receptors

    SciTech Connect

    Monaghan, D.T.

    1985-01-01

    The majority of the excitatory neurotransmission in the vertebrate Central Nervous System is thought to be mediated by acidic amino acid neurotransmitters. However, relatively little is known about the excitatory amino acid receptors and their distribution within the CNS. By analyzing radioligand binding to purified synaptic plasma membranes and to thin tissue sections processed for autoradiography, multiple distinct binding sites were found. These binding sites exhibited the pharmacological properties indicative of the excitatory amino acid receptors, which had been identified by electrophysiological techniques. Specifically, L-(/sup 3/H)-glutamate and D-(/sup 3/H)-amino-5-phosphonopentanoate appear to label N-methyl-D-aspartate receptors, L-(/sup 3/H)-glutamate and (/sup 3/H)-kainic acid appear to label kainic acid receptors, and L-(/sup 3/H)-glutamate and (/sup 3/H)-amino-3-hydroxy-5-methyl-4-isoxazolepropionate appear to label quisqualate receptors. Together, these results confirm the three receptor scheme proposed for excitatory amino acid neurotransmission. These results also show that these transmitter-receptor systems are differentially distributed in the brain, and that the total distribution is consistent with that found by other markers for excitatory amino acid-using neurons.

  18. 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

  19. Comparative analyses of lysophosphatidic acid receptor-mediated signaling.

    PubMed

    Fukushima, Nobuyuki; Ishii, Shoichi; Tsujiuchi, Toshifumi; Kagawa, Nao; Katoh, Kazutaka

    2015-06-01

    Lysophosphatidic acid (LPA) is a bioactive lipid mediator that activates G protein-coupled LPA receptors to exert fundamental cellular functions. Six LPA receptor genes have been identified in vertebrates and are classified into two subfamilies, the endothelial differentiation genes (edg) and the non-edg family. Studies using genetically engineered mice, frogs, and zebrafish have demonstrated that LPA receptor-mediated signaling has biological, developmental, and pathophysiological functions. Computational analyses have also identified several amino acids (aa) critical for LPA recognition by human LPA receptors. This review focuses on the evolutionary aspects of LPA receptor-mediated signaling by comparing the aa sequences of vertebrate LPA receptors and LPA-producing enzymes; it also summarizes the LPA receptor-dependent effects commonly observed in mouse, frog, and fish. PMID:25732591

  20. 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.

  1. 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. PMID:25010412

  2. [Antinociceptive effect of docosahexaenoic acid (DHA) through long fatty acid receptor G protein-coupled receptor 40 (GPR40)].

    PubMed

    Nakamoto, Kazuo; Nishinaka, Takashi; Sato, Naoya; Mankura, Mitsumasa; Koyama, Yutaka; Tokuyama, Shogo

    2014-01-01

    Fatty acids, one class of essential nutrients for humans, are an important source of energy and an essential component of cell membranes. They also function as signal transduction molecules in a variety of biological phenomena. The important functional role of fatty acids in both onset and suppression of pain has become increasingly apparent in recent years. Recently, we have also demonstrated that the release of an endogenous opioid peptide, β-endorphin, plays an important role in the induction of docosahexaenoic acid (DHA)-induced antinociception. It is well known that fatty acids affect intracellular and intercellular signaling as well as the membrane fluidity of neurons. In addition to intracellular actions, unbound free fatty acids (FFAs) can also carry out extracellular signaling by stimulating the G protein-coupled receptor (GPCR). Among these receptors, G protein-coupled receptor 40 (GPR40) has been reported to be activated by long-chain fatty acids such as DHA, eicosapentaenoic acid (EPA) and arachidonic acid. In the peripheral area, GPR40 is preferentially expressed in pancreatic β-cells and is known to relate to the secretion of hormone and peptides. On the other hand, even though this receptor is widely distributed in the central nervous system, reports studying the role and functions of GPR40 in the brain have not been found. In this review, we summarize the findings of our recent study about the long-chain fatty acid receptor GPR40 as a novel pain regulatory system. PMID:24584021

  3. Recognition of Legionella pneumophila nucleic acids by innate immune receptors.

    PubMed

    Cunha, Larissa D; Zamboni, Dario S

    2014-12-01

    Innate immune receptors evolved to sense conserved molecules that are present in microbes or are released during non-physiological conditions. Activation of these receptors is essential for early restriction of microbial infections and generation of adaptive immunity. Among the conserved molecules sensed by innate immune receptors are the nucleic acids, which are abundantly contained in all infectious organisms including virus, bacteria, fungi and parasites. In this review we focus in the innate immune proteins that function to sense nucleic acids from the intracellular bacterial pathogen Legionella pneumophila and the importance of these processes to the outcome of the infection.

  4. 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.

  5. Upregulation of early growth response factor-1 by bile acids requires mitogen-activated protein kinase signaling

    SciTech Connect

    Allen, Katryn; Kim, Nam Deuk; Moon, Jeon-OK; Copple, Bryan L.

    2010-02-15

    Cholestasis results when excretion of bile acids from the liver is interrupted. Liver injury occurs during cholestasis, and recent studies showed that inflammation is required for injury. Our previous studies demonstrated that early growth response factor-1 (Egr-1) is required for development of inflammation in liver during cholestasis, and that bile acids upregulate Egr-1 in hepatocytes. What remains unclear is the mechanism by which bile acids upregulate Egr-1. Bile acids modulate gene expression in hepatocytes by activating the farnesoid X receptor (FXR) and through activation of mitogen-activated protein kinase (MAPK) signaling. Accordingly, the hypothesis was tested that bile acids upregulate Egr-1 in hepatocytes by FXR and/or MAPK-dependent mechanisms. Deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA) stimulated upregulation of Egr-1 to the same extent in hepatocytes isolated from wild-type mice and FXR knockout mice. Similarly, upregulation of Egr-1 in the livers of bile duct-ligated (BDL) wild-type and FXR knockout mice was not different. Upregulation of Egr-1 in hepatocytes by DCA and CDCA was prevented by the MEK inhibitors U0126 and SL-327. Furthermore, pretreatment of mice with U0126 prevented upregulation of Egr-1 in the liver after BDL. Results from these studies demonstrate that activation of MAPK signaling is required for upregulation of Egr-1 by bile acids in hepatocytes and for upregulation of Egr-1 in the liver during cholestasis. These studies suggest that inhibition of MAPK signaling may be a novel therapy to prevent upregulation of Egr-1 in liver during cholestasis.

  6. A thermodynamic switch modulates abscisic acid receptor sensitivity

    PubMed Central

    Dupeux, Florine; Santiago, Julia; Betz, Katja; Twycross, Jamie; Park, Sang-Youl; Rodriguez, Lesia; Gonzalez-Guzman, Miguel; Jensen, Malene Ringkjøbing; Krasnogor, Natalio; Blackledge, Martin; Holdsworth, Michael; Cutler, Sean R; Rodriguez, Pedro L; Márquez, José Antonio

    2011-01-01

    Abscisic acid (ABA) is a key hormone regulating plant growth, development and the response to biotic and abiotic stress. ABA binding to pyrabactin resistance (PYR)/PYR1-like (PYL)/Regulatory Component of Abscisic acid Receptor (RCAR) intracellular receptors promotes the formation of stable complexes with certain protein phosphatases type 2C (PP2Cs), leading to the activation of ABA signalling. The PYR/PYL/RCAR family contains 14 genes in Arabidopsis and is currently the largest plant hormone receptor family known; however, it is unclear what functional differentiation exists among receptors. Here, we identify two distinct classes of receptors, dimeric and monomeric, with different intrinsic affinities for ABA and whose differential properties are determined by the oligomeric state of their apo forms. Moreover, we find a residue in PYR1, H60, that is variable between family members and plays a key role in determining oligomeric state. In silico modelling of the ABA activation pathway reveals that monomeric receptors have a competitive advantage for binding to ABA and PP2Cs. This work illustrates how receptor oligomerization can modulate hormonal responses and more generally, the sensitivity of a ligand-dependent signalling system. PMID:21847091

  7. Characterization of the hydroxycarboxylic acid receptor 2 in cats.

    PubMed

    Graff, E C; Norris, O C; Sandey, M; Kemppainen, R J; Judd, R L

    2015-10-01

    The hydroxycarboxylic acid receptor 2 (HCA2) belongs to a family of nutrient-sensing receptors that bind β-hydroxybutyrate, an alternative fuel source produced during a negative energy balance. The HCA2 receptor has not been identified or characterized in cats. Therefore, the following were the objectives of this study: (1) identify the feline HCA2 receptor protein sequence and compare against known human and rodent sequences, (2) determine tissue distribution and relative expression in lean, healthy cats, and (3) demonstrate in vitro functionality in feline adipose tissue. Tissues (n = 6) and primary adipocytes (n = 4) were collected from lean, healthy, female cats. The published genomic sequence for cats was used to design primers for polymerase chain reaction isolation of HCA2. Relative tissue distribution was evaluated using reverse transcriptase-polymerase chain reaction with RNA isolated from 9 different tissues (spleen, pancreas, lymph node, jejunum, kidney, liver, heart, and subcutaneous and abdominal adipose tissue). Receptor function was evaluated in primary feline adipocyte culture, and changes were compared with basal lipolysis. The in silico predicted feline HCA2 protein sequence exhibited 83.1% and 86.5% amino acid similarity to human and mouse sequences, respectively. The feline HCA2 receptor is predominantly expressed in adipose tissue and spleen. Exposure of feline adipocytes to niacin, a pharmacologic ligand of HCA2, inhibited lipolysis to a similar degree as insulin, a potent lipolytic inhibitor. In conclusion, the feline HCA2 receptor is similar to human and murine receptors in sequence, distribution, and functionality. By gaining a better understanding of the HCA2 receptor in cats, we will be able to better manage feline patients.

  8. Chronic stimulation of farnesoid X receptor impairs nitric oxide sensitivity of vascular smooth muscle.

    PubMed

    Kida, Taiki; Murata, Takahisa; Hori, Masatoshi; Ozaki, Hiroshi

    2009-01-01

    Farnesoid X receptor (FXR), a member of the nuclear receptor superfamily that is highly expressed in enterohepatic tissue, is implicated in bile acid, lipid, and glucose metabolisms. Although recent studies showed that FXR is also expressed in vascular endothelial cells and smooth muscle cells, its physiological and/or pathological roles in vasculature tissue remain unknown. The aim of this study is to examine the chronic effect of synthetic FXR agonist GW4064 on vascular contraction and endothelium-dependent relaxation using tissue culture procedure. In cultured rabbit mesenteric arteries, the treatment with 0.1-10 microM GW4064 for 7 days did not influence vascular contractility induced by high K(+) (15-65 mM), norepinephrine (0.1-100 microM), and endothelin-1 (0.1-100 nM). However, the chronic treatment with GW4064 (1-10 microM for 7 days) dose dependently impaired endothelium-dependent relaxation induced by substance P (0.1-30 nM). In hematoxylin-eosin cross sectioning and en face immunostaining, GW4064 had no effects on the morphology of endothelial and smooth muscle cells. In endothelium-denuded arteries treated with GW4064 (1-10 microM) for 7 days, 3 nM-100 microM sodium nitroprusside-induced vasorelaxation, but not membrane-permeable cGMP analog 8-bromoguanosine-cGMP (8-Br-cGMP; 1-100 microM)-induced vasorelaxation, was significantly impaired. In these GW4064-treated arteries, 1 muM sodium nitroprusside-induced intracellular cGMP elevations were impaired. In RT-PCR, any changes were detected in mRNA expression level of alpha(1)- and beta(1)-subunit of soluble guanylyl cyclase. These results suggest that chronic stimulation of FXR impairs endothelium-dependent relaxation, which is due to decreased sensitivity of smooth muscle cells to nitric oxide.

  9. FXR1, an autosomal homolog of the fragile X mental retardation gene.

    PubMed Central

    Siomi, M C; Siomi, H; Sauer, W H; Srinivasan, S; Nussbaum, R L; Dreyfuss, G

    1995-01-01

    Fragile X mental retardation syndrome, the most common cause of hereditary mental retardation, is directly associated with the FMR1 gene at Xq27.3. FMR1 encodes an RNA binding protein and the syndrome results from lack of expression of FMR1 or expression of a mutant protein that is impaired in RNA binding. We found a novel gene, FXR1, that is highly homologous to FMR1 and located on chromosome 12 at 12q13. FXR1 encodes a protein which, like FMR1, contains two KH domains and is highly conserved in vertebrates. The 3' untranslated regions (3'UTRs) of the human and Xenopus laevis FXR1 mRNAs are strikingly conserved (approximately 90% identity), suggesting conservation of an important function. The KH domains of FXR1 and FMR1 are almost identical, and the two proteins have similar RNA binding properties in vitro. However, FXR1 and FMR1 have very different carboxy-termini. FXR1 and FMR1 are expressed in many tissues, and both proteins, which are cytoplasmic, can be expressed in the same cells. Interestingly, cells from a fragile X patient that do not have any detectable FMR1 express normal levels of FXR1. These findings demonstrate that FMR1 and FXR1 are members of a gene family and suggest a biological role for FXR1 that is related to that of FMR1. Images PMID:7781595

  10. Cross-talk between bile acids and intestinal microbiota in host metabolism and health*

    PubMed Central

    Nie, Yang-fan; Hu, Jun; Yan, Xiang-hua

    2015-01-01

    Bile acid (BA) is de novo synthesized exclusively in the liver and has direct or indirect antimicrobial effects. On the other hand, the composition and size of the BA pool can be altered by intestinal microbiota via the biotransformation of primary BAs to secondary BAs, and subsequently regulate the nuclear farnesoid X receptor (FXR; NR1H4). The BA-activated FXR plays important roles in BA synthesis and metabolism, glucose and lipid metabolism, and even hepatic autophagy. BAs can also play a role in the interplays among intestinal microbes. In this review, we mainly discuss the interactions between BAs and intestinal microbiota and their roles in regulating host metabolism, and probably the autophagic signaling pathway. PMID:26055905

  11. FXR LIA Optimization - Time-resolved OTR Emittance Measurement

    SciTech Connect

    Jacob, J; Ong, M; Wargo, P; LeSage, G

    2005-07-21

    The Flash X-Ray Radiography (FXR) facility at Lawrence Livermore National Laboratory utilizes a high current, long pulse linear induction accelerator to produce high doses of x-ray radiation. Accurate characterization of the transverse beam emittance is required in order to facilitate accelerator modeling and tuning efforts and, ultimately, to optimize the final focus spot size, yielding higher resolution radiographs. In addition to conventional magnet scan, pepper-pot, and multiple screen techniques, optical transition radiation (OTR) has been proven as a useful emittance measurement diagnostic and is particularly well suited to the FXR accelerator. We shall discuss the time-resolved emittance characterization of an induction linac electron beam using OTR, and we will present our experimental apparatus and analysis software. We shall also develop the theoretical background of beam emittance and transition radiation.

  12. 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.

  13. Advances in bile acid medicinal chemistry.

    PubMed

    Sharma, R; Long, A; Gilmer, J F

    2011-01-01

    Bile acids (BAs) are a family of steroidal molecules derived from cholesterol and biosynthesised in the pericentral hepatocytes of the liver. Structurally they may be regarded as consisting of two components, a rigid steroid nucleus and a short aliphatic side chain terminating in an alcohol or carboxyl group. Traditionally BAs are known for their ability to act as solubilising agents in the gut, aiding in the absorption of dietary lipids through the formation of mixed micelles. However the identification of BAs as ligands of the farnesoid X receptor (FXR) has lead to the realisation that these molecules have a wider range of biological effects. BAs regulate lipid and glucose homeostasis through activation of the FXR and the G-protein coupled receptor, TGR5. They can activate apoptotic, inflammatory and carcinogenic signalling pathways. BAs have also been shown to have anti-inflammatory effects. Interestingly, BAs are not restricted to the hepatic-intestinal system. Plasma BAs regulate BA synthesis and metabolism. BAs have recently been identified in cerebrospinal fluid. The BA, ursodeoxycholic acid has a potential role as a neuroprotectant in Huntington's disease and its taurine conjugate exhibits neuro-protective effects in vitro that may be relevant to Alzheimer's disease. This renaissance in BA biology has lead to the development of numerous medicinal chemistry programmes with different therapeutic targets, using BAs as lead structures. BA derivatives with increased efficacy and potency for FXR and TGR5 hold significant promise for the treatment of metabolic disorders. The peculiar effects of BAs on cell viability have been exploited for the design of selective cytocidal agents for treatment of various cancers. BA derivatives have also been screened with much success for anti-microbial and antifungal properties. Other targets include carbonic anhydrase for treatment of glaucoma and the glucocorticoid receptor for antiinflammatory effects. In this review

  14. High affinity retinoic acid receptor antagonists: analogs of AGN 193109.

    PubMed

    Johnson, A T; Wang, L; Gillett, S J; Chandraratna, R A

    1999-02-22

    A series of high affinity retinoic acid receptor (RAR) antagonists were prepared based upon the known antagonist AGN 193109 (2). Introduction of various phenyl groups revealed a preference for substitution at the para-position relative to the meta-site. Antagonists with the highest affinities for the RARs possessed hydrophobic groups, however, the presence of polar functionality was also well tolerated.

  15. 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.

  16. CMP-N-acetylneuraminic acid synthetase interacts with fragile X related protein 1

    PubMed Central

    Ma, Yun; Tian, Shuai; Wang, Zongbao; Wang, Changbo; Chen, Xiaowei; Li, Wei; Yang, Yang; He, Shuya

    2016-01-01

    Fragile X mental retardation protein (FMRP), fragile X related 1 protein (FXR1P) and FXR2P are the members of the FMR protein family. These proteins contain two KH domains and a RGG box, which are characteristic of RNA binding proteins. The absence of FMRP, causes fragile X syndrome (FXS), the leading cause of hereditary mental retardation. FXR1P is expressed throughout the body and important for normal muscle development, and its absence causes cardiac abnormality. To investigate the functions of FXR1P, a screen was performed to identify FXR1P-interacting proteins and determine the biological effect of the interaction. The current study identified CMP-N-acetylneuraminic acid synthetase (CMAS) as an interacting protein using the yeast two-hybrid system, and the interaction between FXR1P and CMAS was validated in yeast using a β-galactosidase assay and growth studies with selective media. Furthermore, co-immunoprecipitation was used to analyze the FXR1P/CMAS association and immunofluorescence microscopy was performed to detect expression and intracellular localization of the proteins. The results of the current study indicated that FXR1P and CMAS interact, and colocalize in the cytoplasm and the nucleus of HEK293T and HeLa cells. Accordingly, a fragile X related 1 (FXR1) gene overexpression vector was constructed to investigate the effect of FXR1 overexpression on the level of monosialotetrahexosylganglioside 1 (GM1). The results of the current study suggested that FXR1P is a tissue-specific regulator of GM1 levels in SH-SY5Y cells, but not in HEK293T cells. Taken together, the results initially indicate that FXR1P interacts with CMAS, and that FXR1P may enhance the activation of sialic acid via interaction with CMAS, and increase GM1 levels to affect the development of the nervous system, thus providing evidence for further research into the pathogenesis of FXS. PMID:27357083

  17. E297G mutated bile salt export pump (BSEP) function enhancers derived from GW4064: structural development study and separation from farnesoid X receptor-agonistic activity.

    PubMed

    Misawa, Takashi; Hayashi, Hisamitsu; Makishima, Makoto; Sugiyama, Yuichi; Hashimoto, Yuichi

    2012-06-15

    Bile salt export pump (BSEP) is a member of the ATP-binding cassette transmembrane transporter family and mediates biliary excretion of bile acids from hepatocytes. Several BSEP mutants, including Glu297Gly (E297G) and Asp482Gly (D482G), cause progressive familial intrahepatic cholestasis type 2. We previously found that compounds based on GW4064, a representative farnesoid X receptor (FXR) agonist, enhanced E297G BSEP transport activity. Here, we conducted a structure-activity relationship analysis of GW4064 derivatives aimed at separating E297G BSEP-function-promoting activity and FXR-agonistic activity. Among newly synthesized reversed-amide derivatives of previously reported GW4064 analogs 2a-2f, we identified 7c as a selective BSEP function enhancer.

  18. Activation of farnesoid X receptor downregulates visfatin and attenuates diabetic nephropathy.

    PubMed

    Zhou, Baoshang; Feng, Bing; Qin, Zhexue; Zhao, Youguang; Chen, Yu; Shi, Zhengmin; Gong, Yi; Zhang, Jing; Yuan, Fahuan; Mu, Jiao

    2016-01-01

    Visfatin, a recently discovered adipocytokine, has been shown to have an important role in the pathogenesis of diabetic nephropathy (DN). The farnesoid X receptor (FXR), a ligand-activated nuclear receptor, plays a protective role in DN. The regulation between FXR and visfatin and their interaction in DN has not been well established. In this study, we reported that FXR agonist GW4064 reduced high glucose induced human mesangial cells (HMCs) inflammation, fibrosis and proliferation by downregulating visfatin expression, which can be blunted by exogenous visfatin treatment. Moreover, luciferase reporter assay showed FXR regulated visfatin transcription activity probably by binding to the -1607 bp and -1192 bp region of the visfatin promoter. In vivo study also showed that GW4064 ameliorated the progression of DN in db/db mice with a decreased visfatin expression. These findings suggest that FXR activation delayed the progression of diabetic nephropathy and this effect is through downregulating visfatin.

  19. Ligand regulation of retinoic acid receptor-related orphan receptors: implications for development of novel therapeutics

    PubMed Central

    Solt, Laura A.; Griffin, Patrick R.; Burris, Thomas P.

    2016-01-01

    Purpose of review In the late 1980s, the cloning of several nuclear receptors led to the intense search and isolation of new members of this superfamily. Despite their identification, many of these receptors were dubbed ‘orphan’ receptors, as their physiological ligands remained unknown. Recent reports have presented evidence for one family of orphan receptors, the retinoic acid receptor-related orphan receptors (RORs), in several pathologies, including osteoporosis, several autoimmune diseases, asthma, cancer, diabetes and obesity. The present review summarizes the studies identifying ligands for the RORs and evaluates their role as targets for potential therapeutics. Recent findings Significant progress was made in the initial identification of ligands for the RORs when X-ray crystallographic studies identified several molecules within the ligand-binding pockets of RORα and RORβ. Recently, we identified endogenous and synthetic ligands for RORα and RORγ, thereby solidifying their function as ligand-dependent transcription factors. Summary Recent studies have established roles for the RORs in physiological development and the advent of disease. Identification of ligands for the RORs, both endogenous and synthetic, has established these receptors as attractive new therapeutic targets for the treatment of ROR-related diseases. PMID:20463469

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

    PubMed

    Chang, Pishan; Augustin, Katrin; Boddum, Kim; Williams, Sophie; Sun, Min; Terschak, John A; Hardege, Jörg D; Chen, Philip E; Walker, Matthew C; Williams, Robin S B

    2016-02-01

    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

  1. Lysophosphatidic acid receptor (LPAR) modulators: The current pharmacological toolbox.

    PubMed

    Llona-Minguez, Sabin; Ghassemian, Artin; Helleday, Thomas

    2015-04-01

    Lysophosphatidic acids (LPA) are key lipid-signalling molecules that regulate a remarkably diverse set of cellular events, such as motility, chemotaxis, cell cycle progression, viability, and wound healing. The physiological and pathophysiological consequences of LPA signalling are evident and misregulation of LPA signalling can lead to pathologies like cancer, atherosclerosis, ischaemia, and fibrosis. LPA exerts its biological actions mainly through several types of G protein-coupled receptors, some of which display opposing or redundant effects. For this reason, selective LPA receptor small-molecule ligands can shine light on LPA biology and present an exciting opportunity for drug discovery endeavours. This review provides insights into the detailed chemical nature and pharmacological profile of the small-molecules thus far developed as LPA receptor modulators, as well as information on the preparation of key pharmaceuticals. This summary will facilitate future research efforts and nurture collaboration between chemists and biologists working in this emerging field. PMID:25704399

  2. 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.

  3. Retinoic acid receptor alpha (RAralpha) Mutations in Human Leukemia.

    PubMed

    Parrado, A; Chomienne, C; Padua, R A

    2000-10-01

    The retinoic acid receptor alpha (RARalpha) plays a central role in the biology of the myeloid cellular compartment. Chromosomal translocations involving the RARalpha locus probably represent the malignant initiating events in acute promyelocytic leukemia (APL). Recent studies that identify novel interactions between RARalpha and the nuclear receptor co-activators and co-repressors, new functions of the oncogenic RARalpha fusion proteins and their catabolism in retinoic acid-induced differentiation, and the availability of new transgenic mice models have provided important insights into our understanding of the mechanisms by which mutant forms of RARalpha can be implicated in the development of leukemia. Novel alterations of the RARalpha gene identified in hematopoietic malignant disorders other than APL, such as myelodysplastic syndromes, non-APL acute myeloid leukemias and B-chronic lymphocytic leukemias, suggest that disruption of the RARalpha gene might predispose to myeloid and lymphoid disorders.

  4. 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.

  5. A third human retinoic acid receptor, hRAR-. gamma

    SciTech Connect

    Krust, A.; Kastner, Ph.; Petkovich, M.; Zelent, A.; Chambon, P. )

    1989-07-01

    Retinoic acid receptors (RARs) are retinoic acid (RA)-inducible enhancer factors belonging to the superfamily of steroid/thyroid nuclear receptors. The authors have previously characterized two human RAR (hRAR-{alpha} and hRAR-{beta}) cDNAs and have recently cloned their murine cognates (mRAR-{alpha} and mRAR-{beta}) together with a third RAR (mRAR-{gamma}) whose RNA was detected predominantly in skin, a well-known target for RA. mRAR-{gamma} cDNA was used here to clone its human counterpart (hRAR-{gamma}) from a T47D breast cancer cell cDNA library. Using a transient transfection assay in HeLa cells and a reporter gene harboring a synthetic RA responsive element, they demonstrate that hRAR-{gamma} cDNA indeed encodes a RA-inducible transcriptional trans-activator. Interestingly, comparisons of the amino acid sequences of all six human and mouse RARs indicate that the interspecies conservation of a given member of the RAR subfamily (either {alpha}, {beta}, or {gamma}) is much higher than the conservation of all three receptors within a given species. These observations indicate that RAR-{alpha}, -{beta}, and -{gamma} may perform specific functions. They show also that hRAR-{gamma} RNA is the predominant RAR RNA species in human skin, which suggests that hRAR-{gamma} mediates some of the retinoid effects in this tissue.

  6. Adenovirus Type 37 Uses Sialic Acid as a Cellular Receptor

    PubMed Central

    Arnberg, Niklas; Edlund, Karin; Kidd, Alistair H.; Wadell, Göran

    2000-01-01

    Two cellular receptors for adenovirus, coxsackievirus-adenovirus receptor (CAR) and major histocompatibility complex class I (MHC-I) α2, have recently been identified. In the absence of CAR, MHC-I α2 has been suggested to serve as a cellular attachment protein for subgenus C adenoviruses, while members from all subgenera except subgenus B have been shown to interact with CAR. We have found that adenovirus type 37 (Ad37) attachment to CAR-expressing CHO cells was no better than that to CHO cells lacking CAR expression, suggesting that CAR is not used by Ad37 during attachment. Instead, we have identified sialic acid as a third adenovirus receptor moiety. First, Ad37 attachment to both CAR-expresing CHO cells and MHC-I α2-expressing Daudi cells was sensitive to neuraminidase treatment, which eliminates sialic acid on the cell surface. Second, Ad37 attachment to sialic acid-expressing Pro-5 cells was more than 10-fold stronger than that to the Pro-5 subline Lec2, which is deficient in sialic acid expression. Third, neuraminidase treatment of A549 cells caused a 60% decrease in Ad37 replication in a fluorescent-focus assay. Moreover, the receptor sialoconjugate is most probably a glycoprotein rather than a ganglioside, since Ad37 attachment to sialic acid-expressing Pro-5 cells was sensitive to protease treatment. Ad37 attachment to Pro-5 cells occurs via α(2→3)-linked sialic acid saccharides rather than α(2→6)-linked ones, since (i) α(2→3)-specific but not α(2→6)-specific lectins blocked Ad37 attachment to Pro-5 cells and (ii) pretreatment of Pro-5 cells with α(2→3)-specific neuraminidase resulted in decreased Ad37 binding. Taken together, these results suggest that, unlike Ad5, Ad37 makes use of α(2→3)-linked sialic acid saccharides on glycoproteins for entry instead of using CAR or MHC-I α2. PMID:10590089

  7. 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

  8. Development of novel silicon-containing inverse agonists of retinoic acid receptor-related orphan receptors.

    PubMed

    Toyama, Hirozumi; Nakamura, Masaharu; Nakamura, Masahiko; Matsumoto, Yotaro; Nakagomi, Madoka; Hashimoto, Yuichi

    2014-03-15

    Retinoic acid receptor (RAR)-related orphan receptors (RORs) regulate a variety of physiological processes, including hepatic gluconeogenesis, lipid metabolism, circadian rhythm and immune function. The RAR agonist: all-trans retinoic acid was reported to be an RORβ inverse agonist, but no information is available regarding ROR activity of its synthetic analogue Am580. Therefore, we screened Am580 and some related tetramethyltetrahydronaphthalene derivatives and carried out structural development studies, including substitution of carbon atoms with silicon, with the aim of creating a potent ROR transcriptional inhibitor. The phenyl amide disila compound 22 showed the most potent ROR-inhibitory activity among the compounds examined. Its activity towards RORα, RORβ and RORγ was increased compared to that of Am580. The IC₅₀ values for RORα, RORβ and RORγ are 1.3, >10 and 4.5 μM, respectively.

  9. FXR Agonist INT-747 Upregulates DDAH Expression and Enhances Insulin Sensitivity in High-Salt Fed Dahl Rats

    PubMed Central

    Ghebremariam, Yohannes T.; Yamada, Keisuke; Lee, Jerry C.; Johnson, Christine L. C.; Atzler, Dorothee; Anderssohn, Maike; Agrawal, Rani; Higgins, John P.; Patterson, Andrew J.; Böger, Rainer H.; Cooke, John P

    2013-01-01

    Aims Genetic and pharmacological studies have shown that impairment of the nitric oxide (NO) synthase (NOS) pathway is associated with hypertension and insulin-resistance (IR). In addition, inhibition of NOS by the endogenous inhibitor, asymmetric dimethylarginine (ADMA), may also result in hypertension and IR. On the other hand, overexpression of dimethylarginine dimethylaminohydrolase (DDAH), an enzyme that metabolizes ADMA, in mice is associated with lower ADMA, increased NO and enhanced insulin sensitivity. Since DDAH carries a farnesoid X receptor (FXR)-responsive element, we aimed to upregulate its expression by an FXR-agonist, INT-747, and evaluate its effect on blood pressure and insulin sensitivity. Methods and Results In this study, we evaluated the in vivo effect of INT-747 on tissue DDAH expression and insulin sensitivity in the Dahl rat model of salt-sensitive hypertension and IR (Dahl-SS). Our data indicates that high salt (HS) diet significantly increased systemic blood pressure. In addition, HS diet downregulated tissue DDAH expression while INT-747 protected the loss in DDAH expression and enhanced insulin sensitivity compared to vehicle controls. Conclusion Our study may provide the basis for a new therapeutic approach for IR by modulating DDAH expression and/or activity using small molecules. PMID:23593273

  10. 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.

  11. Dehydroepiandrosterone sulfotransferase is a target for transcriptional induction by the vitamin D receptor.

    PubMed

    Echchgadda, Ibtissam; Song, Chung S; Roy, Arun K; Chatterjee, Bandana

    2004-03-01

    Dehydroepiandrosterone sulfotransferase (SULT2A1) is a cytosolic enzyme that mediates sulfo-conjugation of endogenous hydroxysteroids (dehydroepiandrosterone, testosterone, bile acids), and diverse xenobiotic compounds. Upon sulfonation, SULT2A1 substrates become polar and water-soluble and thus suitable for rapid excretion. SULT2A1 is abundantly expressed in the liver and intestine. Recent evidence has shown that the ligand-activated vitamin D receptor (VDR) can transcriptionally induce the xenobiotic-metabolizing cytochrome P450 enzymes. Herein, we report that VDR also targets SULT2A1 for transcriptional activation. Vitamin D stimulated endogenous SULT2A1 expression and induced transfected human, mouse, and rat SULT2A1 promoters in liver and intestinal cells upon cotransfection with VDR. An inverted repeat DNA element (IR0), located within -191 to -168 positions of mouse and rat Sult2A1, mediates VDR induction of Sult2A1. DNase1 footprinting, competition EMSA, and antibody supershift assay showed that the IR0 is a binding site for the RXR-alpha/VDR heterodimer. Point mutations within the IR0 prevented RXR/VDR binding and abolished VDR-mediated Sult2A1 induction. The IR0 element conferred VDR responsiveness on a thymidine kinase promoter. Thus, VDR-mediated nuclear signaling may be important in the phase II metabolism involving Sult2A1. The rodent Sult2A1 gene is also induced by the farnesoid X receptor (FXR) and pregnane X receptor (PXR) through the same IR0. In competition transfections, FXR or PXR inhibited VDR induction of the IR0. Competitive functional interactions among VDR, PXR, and FXR suggest that the intracellular hormonal and metabolic milieu may determine the extent to which a specific nuclear receptor pathway would influence steroid/xenobiotic metabolism using dehydroepiandrosterone sulfotransferase.

  12. N-Myristoylation regulates the axonal distribution of the fragile X-related protein FXR2P

    PubMed Central

    Stackpole, Emily E.; Akins, Michael R.; Fallon, Justin R.

    2014-01-01

    Fragile X Syndrome, the leading cause of inherited intellectual disability and autism, is caused by loss of function of Fragile X mental retardation protein (FMRP). FMRP is an RNA binding protein that regulates local protein synthesis in the somatodendritic compartment. However, emerging evidence also indicates important roles for FMRP in axonal and presynaptic function. In particular, FMRP and its homolog FXR2P localize axonally and presynaptically to discrete endogenous structures in the brain termed Fragile X granules (FXGs). FXR2P is a component of all FXGs and is necessary for the axonal and presynaptic localization of FMRP to these structures. We therefore sought to identify and characterize structural features of FXR2P that regulate its axonal localization. Sequence analysis reveals that FXR2P harbors a consensus N-terminal myristoylation sequence (MGXXXS) that is absent in FMRP. Using click chemistry with wild type and an unmyristoylatable G2A mutant we demonstrate that FXR2P is N-myristoylated on glycine 2, establishing it as a lipid-modified RNA binding protein. To investigate the role of FXR2P N-myristoylation in neurons we generated fluorescently tagged wild type and unmyristoylatable FXR2P (WT and G2A, respectively) and expressed them in primary cortical cultures. Both FXR2PWT and FXR2PG2A are expressed at equivalent overall levels and are capable of forming FMRP-containing axonal granules. However, FXR2PWT granules are largely restricted to proximal axonal segments while granules formed with unmyristoylatable FXR2PG2A are localized throughout the axonal arbor, including in growth cones. These studies indicate that N-terminal myristoylation of the RNA binding protein FXR2P regulates its localization within the axonal arbor. Moreover, since FMRP localization within axonal domains requires its association with FXR2P, these findings suggest that FXR2P lipid modification is a control point for the axonal and presynaptic distribution of FMRP. PMID:25109237

  13. 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'.

  14. Retinoic acid receptor alpha is associated with tamoxifen resistance in breast cancer

    PubMed Central

    Johansson, Henrik J.; Sanchez, Betzabe C.; Mundt, Filip; Forshed, Jenny; Kovacs, Aniko; Panizza, Elena; Hultin-Rosenberg, Lina; Lundgren, Bo; Martens, Ulf; Máthé, Gyöngyvér; Yakhini, Zohar; Helou, Khalil; Krawiec, Kamilla; Kanter, Lena; Hjerpe, Anders; Stål, Olle; Linderholm, Barbro K.; Lehtiö, Janne

    2013-01-01

    About one-third of oestrogen receptor alpha-positive breast cancer patients treated with tamoxifen relapse. Here we identify the nuclear receptor retinoic acid receptor alpha as a marker of tamoxifen resistance. Using quantitative mass spectrometry-based proteomics, we show that retinoic acid receptor alpha protein networks and levels differ in a tamoxifen-sensitive (MCF7) and a tamoxifen-resistant (LCC2) cell line. High intratumoural retinoic acid receptor alpha protein levels also correlate with reduced relapse-free survival in oestrogen receptor alpha-positive breast cancer patients treated with adjuvant tamoxifen solely. A similar retinoic acid receptor alpha expression pattern is seen in a comparable independent patient cohort. An oestrogen receptor alpha and retinoic acid receptor alpha ligand screening reveals that tamoxifen-resistant LCC2 cells have increased sensitivity to retinoic acid receptor alpha ligands and are less sensitive to oestrogen receptor alpha ligands compared with MCF7 cells. Our data indicate that retinoic acid receptor alpha may be a novel therapeutic target and a predictive factor for oestrogen receptor alpha-positive breast cancer patients treated with adjuvant tamoxifen. PMID:23868472

  15. The LLNL Flash X-Ray Induction Linear Accelerator (FXR)

    SciTech Connect

    Multhauf, L G

    2002-09-19

    The FXR is an induction linear accelerator used for high-speed radiography at the Lawrence Livermore National Laboratory's Experimental Test Site. It was designed specifically for the radiography of very thick explosive objects. Since its completion in 1982, it has been very actively used for a large variety of explosives tests, and has been periodically upgraded to achieve higher performance. Upgrades have addressed machine reliability, radiographic sensitivity and resolution, two-frame imaging by double pulsing improvements that are described in detail in the paper. At the same time, the facility in which it was installed has also been extensively upgraded, first by adding space for optical and interferometric diagnostics, and more recently by adding a containment chamber to prevent the environmental dispersal of hazardous and radioactive materials. The containment addition also further expands space for new non-radiographic diagnostics. The new Contained Firing Facility is still in the process of activation. At the same time, FXR is continuing to undergo modifications aimed primarily at further increasing radiographic resolution and sensitivity, and at improving double-pulsed performance.

  16. An Improved SF6 System for the FXR Induction Linac Blumlein Switches

    SciTech Connect

    DeHope, W; Kihara, R; Griffin, K L; Ong, M; Ross, T

    2007-06-16

    The now-mature FXR (Flash X-Ray) radiographic facility at Lawrence Livermore National Laboratory will be briefly described with emphasis on its pulsed power system. The heart of each accelerating cell's pulse forming Blumlein is it's sulfur hexafluoride-based triggered closing switch. FXR's recent upgrade to a recirculating SF{sub 6} gas reclamation system will be described and the resulting accelerator performance and reliability improvements documented. This was accompanied by a detailed switch breakdown study on FXR's Test Stand and the recent analysis of the resulting statistics will be shown.

  17. Identification of an N-oxide pyridine GW4064 analog as a potent FXR agonist.

    PubMed

    Feng, Song; Yang, Minmin; Zhang, Zhenshan; Wang, Zhanguo; Hong, Di; Richter, Hans; Benson, Gregory Martin; Bleicher, Konrad; Grether, Uwe; Martin, Rainer E; Plancher, Jean-Marc; Kuhn, Bernd; Rudolph, Markus Georg; Chen, Li

    2009-05-01

    According to the docking studies and the analysis of a co-crystal structure of GW4064 with FXR, a series of 3-aryl heterocyclic isoxazole analogs were designed and synthesized. N-Oxide pyridine analog (7b) was identified as a promising FXR agonist with potent binding affinity and good efficacy, supporting our hypothesis that through an additional hydrogen bond interaction between the pyridine substituent of isoxazole analogs and Tyr373 and Ser336 of FXR, binding affinity and functional activity could be improved.

  18. 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

  19. 1alpha,25-Dihydroxyvitamin D(3) triggered vitamin D receptor and farnesoid X receptor-like effects in rat intestine and liver in vivo.

    PubMed

    Chow, Edwin C Y; Maeng, Han-Joo; Liu, Shanjun; Khan, Ansar A; Groothuis, Geny M M; Pang, K Sandy

    2009-11-01

    1alpha,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), a natural ligand of the vitamin D receptor (VDR), was found to increase the rat ileal Asbt and bile acid absorption. The effects of VDR, whose expression is low in liver, on hepatic transporters and enzymes are unknown. Protein and mRNA levels of target genes in the small intestine, colon and liver after intraperitoneal dosing of 1,25(OH)(2)D(3) (0-2.56 nmol/kg/day for 4 days) to the rat were determined by Western blotting and qPCR, respectively. The 1,25(OH)(2)D(3) treatment increased total Cyp3a protein and Cyp3a1 mRNA expressions in the proximal small intestine, and the short heterodimer partner (SHP), the fibroblast growth factor 15 (FGF15), organic solute transporter (Ostalpha and Ostbeta) mRNA and Asbt protein expressions in the ileum. About 50% higher portal bile acid concentration (65.1+/-14.9 vs 41.9+/-7.8 microm, p<0.05) and elevated expressions of the hepatic farnesoid X receptor (FXR) and SHP mRNA resulted with 1,25(OH)(2)D(3) treatment. Increased Bsep and Ostalpha mRNA expressions in liver and a>50% reduction in the Cyp7a1 protein level (p<0.05) and cholesterol metabolism in rat liver microsomes (p=0.002), likely consequences of the bile acid-FXR-SHP cascade and activation of the signaling pathway for Cyp7a1 inhibition by FGF15, were found. Increased hepatic multidrug resistance-associated protein (Mrp3) and multidrug resistance protein 1a (Mdr1a) mRNA and P-gp protein were also observed. It was concluded that the changes in hepatic transporters and enzymes in the rat were indirect, secondary effects of the liver FXR-SHP cascade due to increased intestinal absorption of bile acids and elevated levels of FGF15, events that led to the activation of FXR.

  20. Retinoic acid receptors: from molecular mechanisms to cancer therapy.

    PubMed

    di Masi, Alessandra; Leboffe, Loris; De Marinis, Elisabetta; Pagano, Francesca; Cicconi, Laura; Rochette-Egly, Cécile; Lo-Coco, Francesco; Ascenzi, Paolo; Nervi, Clara

    2015-02-01

    Retinoic acid (RA), the major bioactive metabolite of retinol or vitamin A, induces a spectrum of pleiotropic effects in cell growth and differentiation that are relevant for embryonic development and adult physiology. The RA activity is mediated primarily by members of the retinoic acid receptor (RAR) subfamily, namely RARα, RARβ and RARγ, which belong to the nuclear receptor (NR) superfamily of transcription factors. RARs form heterodimers with members of the retinoid X receptor (RXR) subfamily and act as ligand-regulated transcription factors through binding specific RA response elements (RAREs) located in target genes promoters. RARs also have non-genomic effects and activate kinase signaling pathways, which fine-tune the transcription of the RA target genes. The disruption of RA signaling pathways is thought to underlie the etiology of a number of hematological and non-hematological malignancies, including leukemias, skin cancer, head/neck cancer, lung cancer, breast cancer, ovarian cancer, prostate cancer, renal cell carcinoma, pancreatic cancer, liver cancer, glioblastoma and neuroblastoma. Of note, RA and its derivatives (retinoids) are employed as potential chemotherapeutic or chemopreventive agents because of their differentiation, anti-proliferative, pro-apoptotic, and anti-oxidant effects. In humans, retinoids reverse premalignant epithelial lesions, induce the differentiation of myeloid normal and leukemic cells, and prevent lung, liver, and breast cancer. Here, we provide an overview of the biochemical and molecular mechanisms that regulate the RA and retinoid signaling pathways. Moreover, mechanisms through which deregulation of RA signaling pathways ultimately impact on cancer are examined. Finally, the therapeutic effects of retinoids are reported. PMID:25543955

  1. Molecular basis for amino acid sensing by family C G-protein-coupled receptors

    PubMed Central

    Wellendorph, P; Bräuner-Osborne, H

    2009-01-01

    Family C of human G-protein-coupled receptors (GPCRs) is constituted by eight metabotropic glutamate receptors, two γ-aminobutyric acid type B (GABAB1–2) subunits forming the heterodimeric GABAB receptor, the calcium-sensing receptor, three taste1 receptors (T1R1–3), a promiscuous L-α-amino acid receptor G-protein-coupled receptor family C, group 6, subtype A (GPRC6A) and seven orphan receptors. Aside from the orphan receptors, the family C GPCRs are dimeric receptors characterized by a large extracellular Venus flytrap domain which bind the endogenous agonists. Except from the GABAB1–2 and T1R2–3 receptor, all receptors are either activated or positively modulated by amino acids. In this review, we outline mutational, biophysical and structural studies which have elucidated the interaction of the amino acids with the Venus flytrap domains, molecular mechanisms of receptor selectivity and the initial steps in receptor activation. PMID:19298394

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

    PubMed

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

    2016-06-14

    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, (13)C 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

  3. 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

  4. Biological roles and therapeutic potential of hydroxy-carboxylic Acid receptors.

    PubMed

    Ahmed, Kashan

    2011-01-01

    In the recent past, deorphanization studies have described intermediates of energy metabolism to activate G protein-coupled receptors and to thereby regulate metabolic functions. GPR81, GPR109A, and GPR109B, formerly known as the nicotinic acid receptor family, are encoded by clustered genes and share a high degree of sequence homology. Recently, hydroxy-carboxylic acids were identified as endogenous ligands of GPR81, GPR109A, and GPR109B, and therefore these receptors have been placed into a novel receptor family of hydroxy-carboxylic acid (HCA) receptors. The HCA(1) receptor (GPR81) is activated by the glycolytic metabolite 2-hydroxy-propionic acid (lactate), the HCA(2) receptor is activated by the ketone body 3-hydroxy-butyric acid, and the HCA(3) receptor (GPR109B) is a receptor for the β-oxidation intermediate 3-hydroxy-octanoic acid. While HCA(1) and HCA(2) receptors are present in most mammalian species, the HCA(3) receptor is exclusively found in humans and higher primates. HCA receptors are expressed in adipose tissue and mediate anti-lipolytic effects in adipocytes through G(i)-type G protein-dependent inhibition of adenylyl cyclase. HCA(2) and HCA(3) inhibit lipolysis during conditions of increased β-oxidation such as prolonged fasting, whereas HCA(1) mediates the anti-lipolytic effects of insulin in the fed state. As HCA(2) is a receptor for the established anti-dyslipidemic drug nicotinic acid, HCA(1) and HCA(3) also represent promising drug targets and several synthetic ligands for HCA receptors have been developed. In this article, we will summarize the deorphanization and pharmacological characterization of HCA receptors. Moreover, we will discuss recent progress in elucidating the physiological and pathophysiological role to further evaluate the therapeutic potential of the HCA receptor family for the treatment of metabolic disease.

  5. A widely used retinoic acid receptor antagonist induces peroxisome proliferator-activated receptor-gamma activity.

    PubMed

    Schupp, Michael; Curtin, Joshua C; Kim, Roy J; Billin, Andrew N; Lazar, Mitchell A

    2007-05-01

    Nuclear receptors (NRs) are transcription factors whose activity is regulated by the binding of small lipophilic ligands, including hormones, vitamins, and metabolites. Pharmacological NR ligands serve as important therapeutic agents; for example, all-trans retinoic acid, an activating ligand for retinoic acid receptor alpha (RARalpha), is used to treat leukemia. Another RARalpha ligand, (E)-S,S-dioxide-4-(2-(7-(heptyloxy)-3,4-dihydro-4,4-dimethyl-2H-1-benzothiopyran-6-yl)-1-propenyl)-benzoic acid (Ro 41-5253), is a potent antagonist that has been a useful and purportedly specific probe of RARalpha function. Here, we report that Ro 41-5253 also activates the peroxisome proliferator-activated receptor gamma (PPARgamma), a master regulator of adipocyte differentiation and target of widely prescribed antidiabetic thiazolidinediones (TZDs). Ro 41-5253 enhanced differentiation of mouse and human preadipocytes and activated PPARgamma target genes in mature adipocytes. Like the TZDs, Ro 41-5253 also down-regulated PPARgamma protein expression in adipocytes. In addition, Ro 41-5253 activated the PPARgamma-ligand binding domain in transiently transfected HEK293T cells. These effects were not prevented by a potent RARalpha agonist or by depleting cells of RARalpha, indicating that PPARgamma activation was not related to RARalpha antagonism. Indeed, Ro 41-5253 was able to compete with TZD ligands for binding to PPARgamma, suggesting that Ro 41-5253 directly affects PPAR activity. These results vividly demonstrate that pharmacological NR ligands may have "off-target" effects on other NRs. Ro 41-5253 is a PPARgamma agonist as well as an RARalpha antagonist whose pleiotropic effects on NRs may signify a unique spectrum of biological responses.

  6. Structure-activity relationship study of betulinic acid, a novel and selective TGR5 agonist, and its synthetic derivatives: potential impact in diabetes.

    PubMed

    Genet, Cédric; Strehle, Axelle; Schmidt, Céline; Boudjelal, Geoffrey; Lobstein, Annelise; Schoonjans, Kristina; Souchet, Michel; Auwerx, Johan; Saladin, Régis; Wagner, Alain

    2010-01-14

    We describe here the biological screening of a collection of natural occurring triterpenoids against the G protein-coupled receptor TGR5, known to be activated by bile acids and which mediates some important cell functions. This work revealed that betulinic (1), oleanolic (2), and ursolic acid (3) exhibited TGR5 agonist activity in a selective manner compared to bile acids, which also activated FXR, the nuclear bile acid receptor. The most potent natural triterpenoid betulinic acid was chosen as a reference compound for an SAR study. Hemisyntheses were performed on the betulinic acid scaffold, and we focused on structural modifications of the C-3 alcohol, the C-17 carboxylic acid, and the C-20 alkene. In particular, structural variations around the C-3 position gave rise to major improvements of potency exemplified with derivatives 18 dia 2 (RG-239) and 19 dia 2. The best derivative was tested in vitro and in vivo, and its biological profile is discussed.

  7. Thyroid hormone receptor can modulate retinoic acid-mediated axis formation in frog embryogenesis.

    PubMed Central

    Banker, D E; Eisenman, R N

    1993-01-01

    Thyroid hormone receptor acts as a hormone-dependent transcriptional transactivator and as a transcriptional repressor in the absence of thyroid hormone. Specifically, thyroid hormone receptor can repress retinoic acid-induced gene expression through interactions with retinoic acid receptor. (Retinoic acid is a potent teratogen in the frog Xenopus laevis, acting at early embryonic stages to interfere with the formation of anterior structures. Endogenous retinoic acid is thought to act in normal anterior-posterior axis formation.) We have previously shown that thyroid hormone receptor RNA (alpha isotype) is expressed and polysome-associated during Xenopus embryogenesis preceding thyroid gland maturation and endogenous thyroid hormone production (D. E. Banker, J. Bigler, and R. N. Eisenman, Mol. Cell. Biol. 11:5079-5089, 1991). To determine whether thyroid hormone receptor might influence the effects of retinoic acid in early frog development, we have examined the results of ectopic thyroid hormone receptor expression on retinoic acid teratogenesis. We demonstrate that microinjections of full-length thyroid hormone receptor RNA protect injected embryos from retinoic acid teratogenesis. DNA binding is apparently essential to this protective function, as truncated thyroid hormone receptors, lacking DNA-binding domains but including hormone-binding and dimerization domains, do not protect from retinoic acid. We have shown that microinjections of these dominant-interfering thyroid hormone receptors, as well as anti-thyroid hormone receptor antibodies, increase retinoic acid teratogenesis in injected embryos, presumably by inactivating endogenous thyroid hormone receptor. This finding suggests that endogenous thyroid hormone receptors may act to limit retinoic acid sensitivity. On the other hand, after thyroid hormone treatment, ectopic thyroid hormone receptor mediates teratogenesis that is indistinguishable from the dorsoanterior deficiencies produced in retinoic acid

  8. FXR1P is a GSK3β substrate regulating mood and emotion processing.

    PubMed

    Del'Guidice, Thomas; Latapy, Camille; Rampino, Antonio; Khlghatyan, Jivan; Lemasson, Morgane; Gelao, Barbara; Quarto, Tiziana; Rizzo, Giuseppe; Barbeau, Annie; Lamarre, Claude; Bertolino, Alessandro; Blasi, Giuseppe; Beaulieu, Jean-Martin

    2015-08-18

    Inhibition of glycogen synthase kinase 3β (GSK3β) is a shared action believed to be involved in the regulation of behavior by psychoactive drugs such as antipsychotics and mood stabilizers. However, little is known about the identity of the substrates through which GSK3β affects behavior. We identified fragile X mental retardation-related protein 1 (FXR1P), a RNA binding protein associated to genetic risk for schizophrenia, as a substrate for GSK3β. Phosphorylation of FXR1P by GSK3β is facilitated by prior phosphorylation by ERK2 and leads to its down-regulation. In contrast, behaviorally effective chronic mood stabilizer treatments in mice inhibit GSK3β and increase FXR1P levels. In line with this, overexpression of FXR1P in the mouse prefrontal cortex also leads to comparable mood-related responses. Furthermore, functional genetic polymorphisms affecting either FXR1P or GSK3β gene expression interact to regulate emotional brain responsiveness and stability in humans. These observations uncovered a GSK3β/FXR1P signaling pathway that contributes to regulating mood and emotion processing. Regulation of FXR1P by GSK3β also provides a mechanistic framework that may explain how inhibition of GSK3β can contribute to the regulation of mood by psychoactive drugs in mental illnesses such as bipolar disorder. Moreover, this pathway could potentially be implicated in other biological functions, such as inflammation and cell proliferation, in which FXR1P and GSK3 are known to play a role. PMID:26240334

  9. FXR1P is a GSK3β substrate regulating mood and emotion processing

    PubMed Central

    Del’Guidice, Thomas; Latapy, Camille; Rampino, Antonio; Khlghatyan, Jivan; Lemasson, Morgane; Gelao, Barbara; Quarto, Tiziana; Rizzo, Giuseppe; Barbeau, Annie; Lamarre, Claude; Bertolino, Alessandro; Blasi, Giuseppe; Beaulieu, Jean-Martin

    2015-01-01

    Inhibition of glycogen synthase kinase 3β (GSK3β) is a shared action believed to be involved in the regulation of behavior by psychoactive drugs such as antipsychotics and mood stabilizers. However, little is known about the identity of the substrates through which GSK3β affects behavior. We identified fragile X mental retardation-related protein 1 (FXR1P), a RNA binding protein associated to genetic risk for schizophrenia, as a substrate for GSK3β. Phosphorylation of FXR1P by GSK3β is facilitated by prior phosphorylation by ERK2 and leads to its down-regulation. In contrast, behaviorally effective chronic mood stabilizer treatments in mice inhibit GSK3β and increase FXR1P levels. In line with this, overexpression of FXR1P in the mouse prefrontal cortex also leads to comparable mood-related responses. Furthermore, functional genetic polymorphisms affecting either FXR1P or GSK3β gene expression interact to regulate emotional brain responsiveness and stability in humans. These observations uncovered a GSK3β/FXR1P signaling pathway that contributes to regulating mood and emotion processing. Regulation of FXR1P by GSK3β also provides a mechanistic framework that may explain how inhibition of GSK3β can contribute to the regulation of mood by psychoactive drugs in mental illnesses such as bipolar disorder. Moreover, this pathway could potentially be implicated in other biological functions, such as inflammation and cell proliferation, in which FXR1P and GSK3 are known to play a role. PMID:26240334

  10. 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 ...

  11. 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

  12. EMBO Retinoids 2011: mechanisms, biology and pathology of signaling by retinoic acid and retinoic acid receptors

    PubMed Central

    McKenna, Neil J.

    2012-01-01

    Retinoic acid (RA) is one of the principal active metabolites of vitamin A (retinol) which mediates a spectrum of critical physiological and developmental processes. Transcriptional regulation by RA is mediated primarily by members of the retinoic acid receptor (RAR) subfamily of the nuclear receptor (NR) superfamily of transcription factors. NRs bind specific genomic DNA sequence motifs and engage coregulators and components of the basal transcription machinery to effect transcriptional regulation at target gene promoters. Disruption of signaling by retinoic acid is thought to underlie the etiology of a number of inflammatory and neoplastic diseases including breast cancer and haematological malignancies. A meeting of international researchers in retinoid signaling was convened in Strasbourg in September 2011 under the auspices of the European Molecular Biology Organization (EMBO). Retinoids 2011 encompassed myriad mechanistic, biological and pathological aspects of these hormones and their cognate receptors, as well as setting these advances in the context of wider current questions on signaling by members of the NR superfamily. PMID:22438793

  13. Medium-chain Fatty Acid-sensing Receptor, GPR84, Is a Proinflammatory Receptor

    PubMed Central

    Suzuki, Masakatsu; Takaishi, Sachiko; Nagasaki, Miyuki; Onozawa, Yoshiko; Iino, Ikue; Maeda, Hiroaki; Komai, Tomoaki; Oda, Tomiichiro

    2013-01-01

    G protein-coupled receptor 84 (GPR84) is a putative receptor for medium-chain fatty acids (MCFAs), whose pathophysiological roles have not yet been clarified. Here, we show that GPR84 was activated by MCFAs with the hydroxyl group at the 2- or 3-position more effectively than nonhydroxylated MCFAs. We also identified a surrogate agonist, 6-n-octylaminouracil (6-OAU), for GPR84. These potential ligands and the surrogate agonist, 6-OAU, stimulated [35S]GTP binding and accumulated phosphoinositides in a GPR84-dependent manner. The surrogate agonist, 6-OAU, internalized GPR84-EGFP from the cell surface. Both the potential ligands and 6-OAU elicited chemotaxis of human polymorphonuclear leukocytes (PMNs) and macrophages and amplified LPS-stimulated production of the proinflammatory cytokine IL-8 from PMNs and TNFα from macrophages. Furthermore, the intravenous injection of 6-OAU raised the blood CXCL1 level in rats, and the inoculation of 6-OAU into the rat air pouch accumulated PMNs and macrophages in the site. Our results indicate a proinflammatory role of GPR84, suggesting that the receptor may be a novel target to treat chronic low grade inflammation associated-disease. PMID:23449982

  14. Review: Mechanisms of How the Intestinal Microbiota Alters the Effects of Drugs and Bile Acids.

    PubMed

    Klaassen, Curtis D; Cui, Julia Yue

    2015-10-01

    Information on the intestinal microbiota has increased exponentially this century because of technical advancements in genomics and metabolomics. Although information on the synthesis of bile acids by the liver and their transformation to secondary bile acids by the intestinal microbiota was the first example of the importance of the intestinal microbiota in biotransforming chemicals, this review will discuss numerous examples of the mechanisms by which the intestinal microbiota alters the pharmacology and toxicology of drugs and other chemicals. More specifically, the altered pharmacology and toxicology of salicylazosulfapridine, digoxin, l-dopa, acetaminophen, caffeic acid, phosphatidyl choline, carnitine, sorivudine, irinotecan, nonsteroidal anti-inflammatory drugs, heterocyclic amines, melamine, nitrazepam, and lovastatin will be reviewed. In addition, recent data that the intestinal microbiota alters drug metabolism of the host, especially Cyp3a, as well as the significance and potential mechanisms of this phenomenon are summarized. The review will conclude with an update of bile acid research, emphasizing the bile acid receptors (FXR and TGR5) that regulate not only bile acid synthesis and transport but also energy metabolism. Recent data indicate that by altering the intestinal microbiota, either by diet or drugs, one may be able to minimize the adverse effects of the Western diet by altering the composition of bile acids in the intestine that are agonists or antagonists of FXR and TGR5. Therefore, it may be possible to consider the intestinal microbiota as another drug target.

  15. Review: Mechanisms of How the Intestinal Microbiota Alters the Effects of Drugs and Bile Acids

    PubMed Central

    Cui, Julia Yue

    2015-01-01

    Information on the intestinal microbiota has increased exponentially this century because of technical advancements in genomics and metabolomics. Although information on the synthesis of bile acids by the liver and their transformation to secondary bile acids by the intestinal microbiota was the first example of the importance of the intestinal microbiota in biotransforming chemicals, this review will discuss numerous examples of the mechanisms by which the intestinal microbiota alters the pharmacology and toxicology of drugs and other chemicals. More specifically, the altered pharmacology and toxicology of salicylazosulfapridine, digoxin, l-dopa, acetaminophen, caffeic acid, phosphatidyl choline, carnitine, sorivudine, irinotecan, nonsteroidal anti-inflammatory drugs, heterocyclic amines, melamine, nitrazepam, and lovastatin will be reviewed. In addition, recent data that the intestinal microbiota alters drug metabolism of the host, especially Cyp3a, as well as the significance and potential mechanisms of this phenomenon are summarized. The review will conclude with an update of bile acid research, emphasizing the bile acid receptors (FXR and TGR5) that regulate not only bile acid synthesis and transport but also energy metabolism. Recent data indicate that by altering the intestinal microbiota, either by diet or drugs, one may be able to minimize the adverse effects of the Western diet by altering the composition of bile acids in the intestine that are agonists or antagonists of FXR and TGR5. Therefore, it may be possible to consider the intestinal microbiota as another drug target. PMID:26261286

  16. Regulation of vitamin D receptor expression by retinoic acid receptor alpha in acute myeloid leukemia cells.

    PubMed

    Marchwicka, Aleksandra; Cebrat, Małgorzata; Łaszkiewicz, Agnieszka; Śnieżewski, Łukasz; Brown, Geoffrey; Marcinkowska, Ewa

    2016-05-01

    Acute myeloid leukemia (AML) is the predominant acute leukemia among adults, characterized by an accumulation of malignant immature myeloid precursors. A very promising way to treat AML is differentiation therapy using either all-trans-retinoic acid (ATRA) or 1,25-dihydroxyvitamin D3 (1,25D), or the use of both these differentiation-inducing agents. However, the effect of combination treatment varies in different AML cell lines, and this is due to ATRA either down- or up-regulating transcription of vitamin D receptor (VDR) in the cells examined. The mechanism of transcriptional regulation of VDR in response to ATRA has not been fully elucidated. Here, we show that the retinoic acid receptor α (RARα) is responsible for regulating VDR transcription in AML cells. We have shown that a VDR transcriptional variant, originating in exon 1a, is regulated by RARα agonists in AML cells. Moreover, in cells with a high basal level of RARα protein, the VDR gene is transcriptionally repressed as long as RARα agonist is absent. In these cells down-regulation of the level of RARα leads to increased expression of VDR. We consider that our findings provide a mechanistic background to explain the different outcomes from treating AML cell lines with a combination of ATRA and 1,25D. PMID:26969398

  17. 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".

  18. Tulane virus recognizes sialic acids as cellular receptors.

    PubMed

    Tan, Ming; Wei, Chao; Huang, Pengwei; Fan, Qiang; Quigley, Christina; Xia, Ming; Fang, Hao; Zhang, Xufu; Zhong, Weiming; Klassen, John S; Jiang, Xi

    2015-01-01

    The recent discovery that human noroviruses (huNoVs) recognize sialic acids (SAs) in addition to histo-blood group antigens (HBGAs) pointed to a new direction in studying virus-host interactions during calicivirus infection. HuNoVs remain difficult to study due to the lack of an effective cell culture model. In this study, we demonstrated that Tulane virus (TV), a cultivable primate calicivirus, also recognizes SAs in addition to the previously known TV-HBGA interactions. Evidence supporting this discovery includes that TV virions bound synthetic sialoglycoconjugates (SGCs) and that treatment of TV permissive LLC-MK2 cells with either neuraminidases or SA-binding lectins inhibited TV infectivity. In addition, we found that Maackia amurensis leukoagglutinin (MAL), a lectin that recognizes the α-2,3 linked SAs, bound LLC-MK2 cells, as well as TV, by which MAL promoted TV infectivity in cell culture. Our findings further highlight TV as a valuable surrogate for huNoVs, particularly in studying virus-host interactions that may involve two host carbohydrate receptors or co-receptors for infection. PMID:26146020

  19. Identification of the orphan GPCR, P2Y(10) receptor as the sphingosine-1-phosphate and lysophosphatidic acid receptor.

    PubMed

    Murakami, Masanori; Shiraishi, Akira; Tabata, Kenichi; Fujita, Norihisa

    2008-07-11

    Phylogenetic analysis of transmembrane regions of GPCRs using PHYLIP indicated that the orphan receptor P2Y(10) receptor was classified into the cluster consisting nucleotide and lipid receptors. Based on the results, we studied the abilities of nucleotides and lipids to activate the P2Y(10) receptors. As a result, sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) evoked intracellular Ca(2+) increases in the CHO cells stably expressing the P2Y(10) fused with a G(16alpha) protein. These Ca(2+) responses were inhibited by S1P receptor and LPA receptor antagonists. The introduction of siRNA designed for P2Y(10) receptor into the P2Y(10)-CHO cells effectively blocked both S1P- and LPA-induced Ca(2+) increases. RT-PCR analysis showed that the mouse P2Y(10) was expressed in reproductive organs, brain, lung and skeletal muscle, suggesting the receptor plays physiological roles throughout the whole body. In conclusion, the P2Y(10) receptor is the first receptor identified as a dual lysophospholipid receptor. PMID:18466763

  20. Cinnabarinic acid, an endogenous metabolite of the kynurenine pathway, activates type 4 metabotropic glutamate receptors.

    PubMed

    Fazio, F; Lionetto, L; Molinaro, G; Bertrand, H O; Acher, F; Ngomba, R T; Notartomaso, S; Curini, M; Rosati, O; Scarselli, P; Di Marco, R; Battaglia, G; Bruno, V; Simmaco, M; Pin, J P; Nicoletti, F; Goudet, C

    2012-05-01

    Cinnabarinic acid is an endogenous metabolite of the kynurenine pathway that meets the structural requirements to interact with glutamate receptors. We found that cinnabarinic acid acts as a partial agonist of type 4 metabotropic glutamate (mGlu4) receptors, with no activity at other mGlu receptor subtypes. We also tested the activity of cinnabarinic acid on native mGlu4 receptors by examining 1) the inhibition of cAMP formation in cultured cerebellar granule cells; 2) protection against excitotoxic neuronal death in mixed cultures of cortical cells; and 3) protection against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity in mice after local infusion into the external globus pallidus. In all these models, cinnabarinic acid behaved similarly to conventional mGlu4 receptor agonists, and, at least in cultured neurons, the action of low concentrations of cinnabarinic acid was largely attenuated by genetic deletion of mGlu4 receptors. However, high concentrations of cinnabarinic acid were still active in the absence of mGlu4 receptors, suggesting that the compound may have off-target effects. Mutagenesis and molecular modeling experiments showed that cinnabarinic acid acts as an orthosteric agonist interacting with residues of the glutamate binding pocket of mGlu4. Accordingly, cinnabarinic acid did not activate truncated mGlu4 receptors lacking the N-terminal Venus-flytrap domain, as opposed to the mGlu4 receptor enhancer, N-phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC). Finally, we could detect endogenous cinnabarinic acid in brain tissue and peripheral organs by high-performance liquid chromatography-tandem mass spectrometry analysis. Levels increased substantially during inflammation induced by lipopolysaccharide. We conclude that cinnabarinic acid is a novel endogenous orthosteric agonist of mGlu4 receptors endowed with neuroprotective activity. PMID:22311707

  1. 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.

  2. 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

  3. Abolished synthesis of cholic acid reduces atherosclerotic development in apolipoprotein E knockout mice.

    PubMed

    Slätis, Katharina; Gåfvels, Mats; Kannisto, Kristina; Ovchinnikova, Olga; Paulsson-Berne, Gabrielle; Parini, Paolo; Jiang, Zhao-Yan; Eggertsen, Gösta

    2010-11-01

    To investigate the effects of abolished cholic acid (CA) synthesis in the ApoE knockout model [apolipoprotein E (apoE) KO],a double-knockout (DKO) mouse model was created by crossbreeding Cyp8b1 knockout mice (Cyp8b1 KO), unable to synthesize the primary bile acid CA, with apoE KO mice. After 5 months of cholesterol feeding, the development of atherosclerotic plaques in the proximal aorta was 50% less in the DKO mice compared with the apoE KO mice. This effect was associated with reduced intestinal cholesterol absorption, decreased levels of apoB-containing lipoproteins in the plasma, enhanced bile acid synthesis, reduced hepatic cholesteryl esters, and decreased hepatic activity of ACAT2. The upregulation of Cyp7a1 in DKO mice seemed primarily caused by reduced expression of the intestinal peptide FGF15. Treatment of DKO mice with the farnesoid X receptor (FXR) agonist GW4064 did not alter the intestinal cholesterol absorption, suggesting that the action of CA in this process is confined mainly to formation of intraluminal micelles and less to its ability to activate the nuclear receptor FXR. Inhibition of CA synthesis may offer a therapeutic strategy for the treatment of hyperlipidemic conditions that lead to atherosclerosis.

  4. Direct activation of GABAA receptors by substances in the organic acid fraction of Japanese sake.

    PubMed

    Izu, Hanae; Shigemori, Kensuke; Eguchi, Masaya; Kawane, Shuhei; Fujii, Shouko; Kitamura, Yuji; Aoshima, Hitoshi; Yamada, Yasue

    2017-01-01

    We investigated the effect of substances present in Japanese sake on the response of ionotropic γ-aminobutyric acid (GABA)A receptors expressed in Xenopus oocytes. Sake was fractionated by ion-exchange chromatography. The fraction containing organic acids (OA fraction) showed agonist activities on the GABAA receptor. OA fractions from sake were analyzed by capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS). Of the 64 compounds identified, 13 compounds showed GABAA receptor agonist activities. Especially, l-lactic acid showed high agonist activity and its EC50 value was 37μM. Intraperitoneal injections of l-lactic acid, gluconic acid, and pyruvic acid (10, 10, and 5mg/kg BW, respectively), which showed agonistic activity on the GABAA receptor, led to significant anxiolytic effects during an elevated plus-maze test in mice. PMID:27507485

  5. Inhibition of ileal bile acid uptake protects against nonalcoholic fatty liver disease in high-fat diet-fed mice.

    PubMed

    Rao, Anuradha; Kosters, Astrid; Mells, Jamie E; Zhang, Wujuan; Setchell, Kenneth D R; Amanso, Angelica M; Wynn, Grace M; Xu, Tianlei; Keller, Brad T; Yin, Hong; Banton, Sophia; Jones, Dean P; Wu, Hao; Dawson, Paul A; Karpen, Saul J

    2016-09-21

    Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in the Western world, and safe and effective therapies are needed. Bile acids (BAs) and their receptors [including the nuclear receptor for BAs, farnesoid X receptor (FXR)] play integral roles in regulating whole-body metabolism and hepatic lipid homeostasis. We hypothesized that interruption of the enterohepatic BA circulation using a luminally restricted apical sodium-dependent BA transporter (ASBT) inhibitor (ASBTi; SC-435) would modify signaling in the gut-liver axis and reduce steatohepatitis in high-fat diet (HFD)-fed mice. Administration of this ASBTi increased fecal BA excretion and messenger RNA (mRNA) expression of BA synthesis genes in liver and reduced mRNA expression of ileal BA-responsive genes, including the negative feedback regulator of BA synthesis, fibroblast growth factor 15. ASBT inhibition resulted in a marked shift in hepatic BA composition, with a reduction in hydrophilic, FXR antagonistic species and an increase in FXR agonistic BAs. ASBT inhibition restored glucose tolerance, reduced hepatic triglyceride and total cholesterol concentrations, and improved NAFLD activity score in HFD-fed mice. These changes were associated with reduced hepatic expression of lipid synthesis genes (including liver X receptor target genes) and normalized expression of the central lipogenic transcription factor, Srebp1c Accumulation of hepatic lipids and SREBP1 protein were markedly reduced in HFD-fed Asbt(-/-) mice, providing genetic evidence for a protective role mediated by interruption of the enterohepatic BA circulation. Together, these studies suggest that blocking ASBT function with a luminally restricted inhibitor can improve both hepatic and whole body aspects of NAFLD.

  6. Inhibition of ileal bile acid uptake protects against nonalcoholic fatty liver disease in high-fat diet-fed mice.

    PubMed

    Rao, Anuradha; Kosters, Astrid; Mells, Jamie E; Zhang, Wujuan; Setchell, Kenneth D R; Amanso, Angelica M; Wynn, Grace M; Xu, Tianlei; Keller, Brad T; Yin, Hong; Banton, Sophia; Jones, Dean P; Wu, Hao; Dawson, Paul A; Karpen, Saul J

    2016-09-21

    Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in the Western world, and safe and effective therapies are needed. Bile acids (BAs) and their receptors [including the nuclear receptor for BAs, farnesoid X receptor (FXR)] play integral roles in regulating whole-body metabolism and hepatic lipid homeostasis. We hypothesized that interruption of the enterohepatic BA circulation using a luminally restricted apical sodium-dependent BA transporter (ASBT) inhibitor (ASBTi; SC-435) would modify signaling in the gut-liver axis and reduce steatohepatitis in high-fat diet (HFD)-fed mice. Administration of this ASBTi increased fecal BA excretion and messenger RNA (mRNA) expression of BA synthesis genes in liver and reduced mRNA expression of ileal BA-responsive genes, including the negative feedback regulator of BA synthesis, fibroblast growth factor 15. ASBT inhibition resulted in a marked shift in hepatic BA composition, with a reduction in hydrophilic, FXR antagonistic species and an increase in FXR agonistic BAs. ASBT inhibition restored glucose tolerance, reduced hepatic triglyceride and total cholesterol concentrations, and improved NAFLD activity score in HFD-fed mice. These changes were associated with reduced hepatic expression of lipid synthesis genes (including liver X receptor target genes) and normalized expression of the central lipogenic transcription factor, Srebp1c Accumulation of hepatic lipids and SREBP1 protein were markedly reduced in HFD-fed Asbt(-/-) mice, providing genetic evidence for a protective role mediated by interruption of the enterohepatic BA circulation. Together, these studies suggest that blocking ASBT function with a luminally restricted inhibitor can improve both hepatic and whole body aspects of NAFLD. PMID:27655848

  7. Novel heterocyclic scaffolds of GW4064 as farnesoid X receptor agonists.

    PubMed

    Smalley, Terrence L; Boggs, Sharon; Caravella, Justin A; Chen, Lihong; Creech, Katrina L; Deaton, David N; Kaldor, Istvan; Parks, Derek J

    2015-01-15

    The farnesoid X receptor (FXR) may play a crucial role in a number of metabolic diseases and, as such, could potentially serve as a target for the development of therapeutics as a treatment for those diseases. Previous work has described GW4064 as an FXR agonist with an interesting activity profile. This manuscript will describe the synthesis of novel analogs of GW4064 and the activity profile of those analogs.

  8. Identification of Hydroxybenzoic Acids as Selective Lactate Receptor (GPR81) Agonists with Antilipolytic Effects.

    PubMed

    Dvorak, Curt A; Liu, Changlu; Shelton, Jonathan; Kuei, Chester; Sutton, Steven W; Lovenberg, Timothy W; Carruthers, Nicholas I

    2012-08-01

    Following the characterization of the lactate receptor (GPR81), a focused screening effort afforded 3-hydroxybenzoic acid 1 as a weak agonist of both GPR81 and GPR109a (niacin receptor). An examination of structurally similar arylhydroxy acids led to the identification of 3-chloro-5-hydroxybenzoic acid 2, a selective GPR81 agonist that exhibited favorable in vivo effects on lipolysis in a mouse model of obesity.

  9. Identification of Hydroxybenzoic Acids as Selective Lactate Receptor (GPR81) Agonists with Antilipolytic Effects

    PubMed Central

    2012-01-01

    Following the characterization of the lactate receptor (GPR81), a focused screening effort afforded 3-hydroxybenzoic acid 1 as a weak agonist of both GPR81 and GPR109a (niacin receptor). An examination of structurally similar arylhydroxy acids led to the identification of 3-chloro-5-hydroxybenzoic acid 2, a selective GPR81 agonist that exhibited favorable in vivo effects on lipolysis in a mouse model of obesity. PMID:24900524

  10. Bile Acid Modifications at the Microbe-Host Interface: Potential for Nutraceutical and Pharmaceutical Interventions in Host Health.

    PubMed

    Joyce, Susan A; Gahan, Cormac G M

    2016-01-01

    Bile acids have emerged as important signaling molecules in the host, as they interact either locally or systemically with specific cellular receptors, in particular the farnesoid X receptor (FXR) and TGR5. These signaling functions influence systemic lipid and cholesterol metabolism, energy metabolism, immune homeostasis, and intestinal electrolyte balance. Through defined enzymatic activities, the gut microbiota can significantly modify the signaling properties of bile acids and therefore can have an impact upon host health. Alterations to the gut microbiota that influence bile acid metabolism are associated with metabolic disease, obesity, diarrhea, inflammatory bowel disease (IBD), Clostridium difficile infection, colorectal cancer, and hepatocellular carcinoma. Here, we examine the regulation of this gut-microbiota-liver axis in the context of bile acid metabolism and indicate how this pathway represents an important target for the development of new nutraceutical (diet and/or probiotics) and targeted pharmaceutical interventions. PMID:26772409

  11. Resveratrol Attenuates Trimethylamine-N-Oxide (TMAO)-Induced Atherosclerosis by Regulating TMAO Synthesis and Bile Acid Metabolism via Remodeling of the Gut Microbiota

    PubMed Central

    Chen, Ming-liang; Yi, Long; Zhang, Yong; Zhou, Xi; Ran, Li; Yang, Jining; Zhu, Jun-dong; Zhang, Qian-yong

    2016-01-01

    ABSTRACT The gut microbiota is found to be strongly associated with atherosclerosis (AS). Resveratrol (RSV) is a natural phytoalexin with anti-AS effects; however, its mechanisms of action remain unclear. Therefore, we sought to determine whether the anti-AS effects of RSV were related to changes in the gut microbiota. We found that RSV attenuated trimethylamine-N-oxide (TMAO)-induced AS in ApoE−/− mice. Meanwhile, RSV decreased TMAO levels by inhibiting commensal microbial trimethylamine (TMA) production via gut microbiota remodeling in mice. Moreover, RSV increased levels of the genera Lactobacillus and Bifidobacterium, which increased the bile salt hydrolase activity, thereby enhancing bile acid (BA) deconjugation and fecal excretion in C57BL/6J and ApoE−/− mice. This was associated with a decrease in ileal BA content, repression of the enterohepatic farnesoid X receptor (FXR)-fibroblast growth factor 15 (FGF15) axis, and increased cholesterol 7a-hydroxylase (CYP7A1) expression and hepatic BA neosynthesis. An FXR antagonist had the same effect on FGF15 and CYP7A1 expression as RSV, while an FXR agonist abolished RSV-induced alterations in FGF15 and CYP7A1 expression. In mice treated with antibiotics, RSV neither decreased TMAO levels nor increased hepatic BA synthesis. Additionally, RSV-induced inhibition of TMAO-caused AS was also markedly abolished by antibiotics. In conclusion, RSV attenuated TMAO-induced AS by decreasing TMAO levels and increasing hepatic BA neosynthesis via gut microbiota remodeling, and the BA neosynthesis was partially mediated through the enterohepatic FXR-FGF15 axis. PMID:27048804

  12. Distinct Phosphorylation Clusters Determine the Signaling Outcome of Free Fatty Acid Receptor 4/G Protein-Coupled Receptor 120.

    PubMed

    Prihandoko, Rudi; Alvarez-Curto, Elisa; Hudson, Brian D; Butcher, Adrian J; Ulven, Trond; Miller, Ashley M; Tobin, Andrew B; Milligan, Graeme

    2016-05-01

    It is established that long-chain free fatty acids includingω-3 fatty acids mediate an array of biologic responses through members of the free fatty acid (FFA) receptor family, which includes FFA4. However, the signaling mechanisms and modes of regulation of this receptor class remain unclear. Here, we employed mass spectrometry to determine that phosphorylation of mouse (m)FFAR4 occurs at five serine and threonine residues clustered in two separable regions of the C-terminal tail, designated cluster 1 (Thr(347), Thr(349), and Ser(350)) and cluster 2 (Ser(357)and Ser(361)). Mutation of these phosphoacceptor sites to alanine completely prevented phosphorylation of mFFA4 but did not limit receptor coupling to extracellular signal regulated protein kinase 1 and 2 (ERK1/2) activation. Rather, an inhibitor of Gq/11proteins completely prevented receptor signaling to ERK1/2. By contrast, the recruitment of arrestin 3, receptor internalization, and activation of Akt were regulated by mFFA4 phosphorylation. The analysis of mFFA4 phosphorylation-dependent signaling was extended further by selective mutations of the phosphoacceptor sites. Mutations within cluster 2 did not affect agonist activation of Akt but instead significantly compromised receptor internalization and arrestin 3 recruitment. Distinctly, mutation of the phosphoacceptor sites within cluster 1 had no effect on receptor internalization and had a less extensive effect on arrestin 3 recruitment but significantly uncoupled the receptor from Akt activation. These unique observations define differential effects on signaling mediated by phosphorylation at distinct locations. This hallmark feature supports the possibility that the signaling outcome of mFFA4 activation can be determined by the pattern of phosphorylation (phosphorylation barcode) at the C terminus of the receptor.

  13. Dominant negative retinoic acid receptor initiates tumor formation in mice

    PubMed Central

    Kupumbati, Tara S; Cattoretti, Giorgio; Marzan, Christine; Farias, Eduardo F; Taneja, Reshma; Mira-y-Lopez, Rafael

    2006-01-01

    Background Retinoic acid suppresses cell growth and promotes cell differentiation, and pharmacological retinoic acid receptor (RAR) activation is anti-tumorigenic. This begs the question of whether chronic physiological RAR activation by endogenous retinoids is likewise anti-tumorigenic. Results To address this question, we generated transgenic mice in which expression of a ligand binding defective dominant negative RARα (RARαG303E) was under the control of the mouse mammary tumor virus (MMTV) promoter. The transgene was expressed in the lymphoid compartment and in the mammary epithelium. Observation of aging mice revealed that transgenic mice, unlike their wild type littermates, developed B cell lymphomas at high penetrance, with a median latency of 40 weeks. MMTV-RARαG303E lymphomas were high grade Pax-5+, surface H+L Ig negative, CD69+ and BCL6- and cytologically and phenotypically resembled human adult high grade (Burkitt's or lymphoblastic) lymphomas. We postulated that mammary tumors might arise after a long latency period as seen in other transgenic models of breast cancer. We tested this idea by transplanting transgenic epithelium into the cleared fat pads of wild type hosts, thus bypassing lymphomagenesis. At 17 months post-transplantation, a metastatic mammary adenocarcinoma developed in one of four transplanted glands whereas no tumors developed in sixteen of sixteen endogenous glands with wild type epithelium. Conclusion These findings suggest that physiological RAR activity may normally suppress B lymphocyte and mammary epithelial cell growth and that global RAR inactivation is sufficient to initiate a stochastic process of tumor development requiring multiple transforming events. Our work makes available to the research community a new animal resource that should prove useful as an experimental model of aggressive sporadic lymphoma in immunologically uncompromised hosts. We anticipate that it may also prove useful as a model of breast cancer. PMID

  14. Expression of retinoic acid receptors in human endometrial carcinoma.

    PubMed

    Tanabe, Kojiro; Utsunomiya, Hiroki; Tamura, Mitsutoshi; Niikura, Hitoshi; Takano, Tadao; Yoshinaga, Kohsuke; Nagase, Satoru; Suzuki, Takashi; Ito, Kiyoshi; Matsumoto, Mitsuyo; Hayashi, Shin-ichi; Yaegashi, Nobuo

    2008-02-01

    The retinoids (vitamin A and its biologically active derivatives) are essential for the health and survival of the individual. Several studies have reported a strong rationale for the use of retinoids in cancer treatment and chemoprevention. It has been discovered that expression of retinoic acid receptor (RAR) beta is frequently silenced in epithelial carcinogenesis, which has led to the hypothesis that RAR beta could act as a tumor suppressor. However, the status of RAR beta in human endometrial carcinoma has not been examined. In the present study, we initially studied the effects of retinoic acid on cell proliferation and the expression of RAR alpha, RAR beta, and RAR gamma using AM580 (a RAR-specific agonist) in the Ishikawa endometrial cancer cell line. We also examined the expression of RAR in human eutopic endometrium (30 cases), endometrial hyperplasia (28 cases), and endometrial carcinoma (103 cases) using immunohistochemistry. Finally, we correlated these findings with the clinicopathological parameters. In vitro, cell growth was inhibited and RAR beta and RAR gamma mRNA was significantly induced by AM580, compared with vehicle controls, whereas RAR alpha mRNA was significantly attenuated by AM580, compared with vehicle. RAR beta was detected predominantly in endometrial hyperplasia, compared with endometrial carcinoma. No statistically significant correlation was obtained between the expression of any other RAR subtypes and clinicopathological parameters in human endometrial carcinoma. The results of our study demonstrate that AM580 inhibits cell growth and induces RAR beta mRNA expression in the Ishikawa cell line, and the expression level of RAR beta in endometrial carcinoma is significantly lower than that in endometrial hyperplasia. AM580 might therefore be considered as a potential treatment for endometrial carcinoma.

  15. 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

  16. Characterization of DNA Binding and Retinoic Acid Binding Properties of Retinoic Acid Receptor

    NASA Astrophysics Data System (ADS)

    Yang, Na; Schule, Roland; Mangelsdorf, David J.; Evans, Ronald M.

    1991-05-01

    High-level expression of the full-length human retinoic acid receptor (RAR) α and the DNA binding domain of the RAR in Escherichia coli was achieved by using a T7 RNA polymerase-directed expression system. After induction, full-length RAR protein was produced at an estimated level of 20% of the total bacterial proteins. Both intact RAR molecules and the DNA binding domain bind to the cognate DNA response element with high specificity in the absence of retinoic acid. However, this binding is enhanced to a great extent upon the addition of eukaryotic cell extracts. The factor responsible for this enhancement is heat-sensitive and forms a complex with RAR that binds to DNA and exhibits a distinct migration pattern in the gel-mobility-shift assay. The interaction site of the factor with RAR is localized in the 70-amino acid DNA binding region of RAR. The hormone binding ability of the RARα protein was assayed by a charcoal absorption assay and the RAR protein was found to bind to retinoic acid with a K_d of 2.1 x 10-10 M.

  17. Among the twenty classical L-amino acids, only glutamate directly activates metabotropic glutamate receptors.

    PubMed

    Frauli, Mélanie; Neuville, Pascal; Vol, Claire; Pin, Jean-Philippe; Prézeau, Laurent

    2006-02-01

    Under pathophysiological conditions, cellular amino acids can be profusely released from cells into the cerebral interstitial space. Because several class-C G protein coupled receptors (GPCRs) display a broad natural ligand spectrum, being sensitive to more than one endogenous ligand, we wondered whether the related metabotropic glutamate (mGlu) receptors could be modulated by various types of L-amino acids, allowing them to sense large increase in extracellular amino acid concentration. Here, the agonist, antagonist and allosteric effects of the twenty classical L-amino acids were evaluated on the eight mGlu receptor subtypes. We show that, in addition to glutamate (Glu), cysteine, aspartate and asparagine also lead to the activation of mGlu3, 4 and 5. Interestingly, our data demonstrate that the effect of these three amino acids did not result from a direct activation of the receptors, but from an indirect action involving Glu-transporters/exchangers. These data first demonstrate that mGlu receptors, unlike other class-C GPCRs, display an extremely high selectivity towards one ligand. Moreover, our results also show that Glu transport systems allow mGlu receptors to sense large increase in the extracellular concentration of some amino acids. Such a system will certainly lead to a large increase in some mGlu receptor activity under pathological conditions, such as seizure, ischemia or other brain injuries. PMID:16310227

  18. Interaction between retinoid acid receptor-related orphan receptor alpha (RORA) and neuropeptide S receptor 1 (NPSR1) in asthma.

    PubMed

    Acevedo, Nathalie; Sääf, Annika; Söderhäll, Cilla; Melén, Erik; Mandelin, Jami; Pietras, Christina Orsmark; Ezer, Sini; Karisola, Piia; Vendelin, Johanna; Gennäs, Gustav Boije af; Yli-Kauhaluoma, Jari; Alenius, Harri; von Mutius, Erika; Doekes, Gert; Braun-Fahrländer, Charlotte; Riedler, Josef; van Hage, Marianne; D'Amato, Mauro; Scheynius, Annika; Pershagen, Göran; Kere, Juha; Pulkkinen, Ville

    2013-01-01

    Retinoid acid receptor-related Orphan Receptor Alpha (RORA) was recently identified as a susceptibility gene for asthma in a genome-wide association study. To investigate the impact of RORA on asthma susceptibility, we performed a genetic association study between RORA single nucleotide polymorphisms (SNPs) in the vicinity of the asthma-associated SNP (rs11071559) and asthma-related traits. Because the regulatory region of a previously implicated asthma susceptibility gene, Neuropeptide S receptor 1 (NPSR1), has predicted elements for RORA binding, we hypothesized that RORA may interact biologically and genetically with NPSR1. 37 RORA SNPs and eight NPSR1 SNPs were genotyped in the Swedish birth cohort BAMSE (2033 children) and the European cross-sectional PARSIFAL study (1120 children). Seven RORA SNPs confined into a 49 kb region were significantly associated with physician-diagnosed childhood asthma. The most significant association with rs7164773 (T/C) was driven by the CC genotype in asthma cases (OR = 2.0, 95%CI 1.36-2.93, p = 0.0003 in BAMSE; and 1.61, 1.18-2.19, p = 0.002 in the combined BAMSE-PARSIFAL datasets, respectively), and strikingly, the risk effect was dependent on the Gln344Arg mutation in NPSR1. In cell models, stimulation of NPSR1 activated a pathway including RORA and other circadian clock genes. Over-expression of RORA decreased NPSR1 promoter activity further suggesting a regulatory loop between these genes. In addition, Rora mRNA expression was lower in the lung tissue of Npsr1 deficient mice compared to wildtype littermates during the early hours of the light period. We conclude that RORA SNPs are associated with childhood asthma and show epistasis with NPSR1, and the interaction between RORA and NPSR1 may be of biological relevance. Combinations of common susceptibility alleles and less common functional polymorphisms may modify the joint risk effects on asthma susceptibility. PMID:23565190

  19. Therapeutic role of bile acids and nuclear receptor agonists in fibrosing cholangiopathies.

    PubMed

    Trauner, Michael; Halilbasic, Emina; Kazemi-Shirazi, Lili; Kienbacher, Christian; Staufer, Katharina; Traussnigg, Stefan; Hofer, Harald

    2014-01-01

    Chronic inflammatory bile duct diseases such as primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC) result in progressive fibrosis of the biliary tract and ultimately cirrhosis of the liver. Since the etiology and pathogenesis of these fibrosing cholangiopathies are still poorly understood, therapeutic options are rather limited at present. Ursodeoxycholic acid (UDCA) is the paradigm therapeutic bile acid and established standard treatment for PBC, but its role for medical therapy of PSC is still under debate. Promising novel bile acid-based therapeutic options include 24-norursodeoxycholic acid, a side chain-shortened C23 homologue of UDCA, and bile acid receptor/farnesoid X receptor agonists (e.g., obeticholic acid) which currently undergo clinical development for fibrosing cholangiopathies such as PBC and PSC. Other nuclear receptors such as vitamin D receptor and fatty acid-activated peroxisome proliferator-activated receptors are also of considerable interest. This review article is a summary of an overview talk given at Falk Symposium 191 on Advances in Pathogenesis and Treatment of Liver Diseases held in London, October 3-4, 2013, and summarizes the recent progress with novel therapeutic bile acids and bile acid derivatives as novel therapies for fibrosing cholangiopathies such as PBC and PSC.

  20. 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

    2006-04-01

    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.

  1. Inhibition of apical sodium-dependent bile acid transporter as a novel treatment for diabetes.

    PubMed

    Chen, Lihong; Yao, Xiaozhou; Young, Andrew; McNulty, Judi; Anderson, Don; Liu, Yaping; Nystrom, Christopher; Croom, Dallas; Ross, Sean; Collins, Jon; Rajpal, Deepak; Hamlet, Kimberly; Smith, Chari; Gedulin, Bronislava

    2012-01-01

    Bile acids are recognized as metabolic modulators. The present study was aimed at evaluating the effects of a potent Asbt inhibitor (264W94), which blocks intestinal absorption of bile acids, on glucose homeostasis in Zucker Diabetic Fatty (ZDF) rats. Oral administration of 264W94 for two wk increased fecal bile acid concentrations and elevated non-fasting plasma total Glp-1. Treatment of 264W94 significantly decreased HbA1c and glucose, and prevented the drop of insulin levels typical of ZDF rats in a dose-dependent manner. An oral glucose tolerance test revealed up to two-fold increase in plasma total Glp-1 and three-fold increase in insulin in 264W94 treated ZDF rats at doses sufficient to achieve glycemic control. Tissue mRNA analysis indicated a decrease in farnesoid X receptor (Fxr) activation in small intestines and the liver but co-administration of a Fxr agonist (GW4064) did not attenuate 264W94 induced glucose lowering effects. In summary, our results demonstrate that inhibition of Asbt increases bile acids in the distal intestine, promotes Glp-1 release and may offer a new therapeutic strategy for type 2 diabetes mellitus.

  2. 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. PMID:27408924

  3. 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

  4. Aromatic L-amino acids activate the calcium-sensing receptor.

    PubMed

    Conigrave, Arthur D; Mun, Hee-Chang; Lok, Hiu-Chuen

    2007-06-01

    The calcium-sensing receptor (CaR) is recognized as a member of class 3 of the G-protein coupled receptor superfamily. Members of this subgroup, which have large N-terminal extracellular domains, include receptors that respond specifically to the amino acid glutamate; receptors that respond to the glutamate analogue, gamma-amino butyric acid; and several receptors that act as broad-spectrum amino acid sensors. The CaR is one of these broad-spectrum amino acid sensors that, along with several other members of the subgroup, also responds to extracellular Ca2+. In this mini-review, we consider evidence that the CaR is a sensor of aromatic amino acids, that it has broad-spectrum amino acid sensing properties, that it provides an amino acid binding site in its extracellular N-terminal Venus Fly Trap domain, and that amino acids have a physiological impact on systems in which the CaR is expressed.

  5. Nuclear retinoic acid receptors: conductors of the retinoic acid symphony during development.

    PubMed

    Samarut, Eric; Rochette-Egly, Cécile

    2012-01-30

    The vitamin A derivative, retinoic acid (RA), is essential for embryonic development through the activation of cognate nuclear receptors, RARs, which work as ligand dependent regulators of transcription. In vitro studies revealed how RARs control gene expression at the molecular level and now it appears that it is fine-tuned by a phosphorylation code. In addition, several genetic approaches provided valuable insights on the functions of RARs during development and on the influence of other actors such as the enzymes involved in RA synthesis and degradation and other signaling pathways. It appears that RARs are the conductors of the RA signaling symphony through controlling the dynamics and the coordination of the different players and development steps.

  6. 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

    2012-06-01

    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.

  7. Estimating The Reliability of the Lawrence Livermore National Laboratory (LLNL) Flash X-ray (FXR) Machine

    SciTech Connect

    Ong, M M; Kihara, R; Zentler, J M; Kreitzer, B R; DeHope, W J

    2007-06-27

    At Lawrence Livermore National Laboratory (LLNL), our flash X-ray accelerator (FXR) is used on multi-million dollar hydrodynamic experiments. Because of the importance of the radiographs, FXR must be ultra-reliable. Flash linear accelerators that can generate a 3 kA beam at 18 MeV are very complex. They have thousands, if not millions, of critical components that could prevent the machine from performing correctly. For the last five years, we have quantified and are tracking component failures. From this data, we have determined that the reliability of the high-voltage gas-switches that initiate the pulses, which drive the accelerator cells, dominates the statistics. The failure mode is a single-switch pre-fire that reduces the energy of the beam and degrades the X-ray spot-size. The unfortunate result is a lower resolution radiograph. FXR is a production machine that allows only a modest number of pulses for testing. Therefore, reliability switch testing that requires thousands of shots is performed on our test stand. Study of representative switches has produced pre-fire statistical information and probability distribution curves. This information is applied to FXR to develop test procedures and determine individual switch reliability using a minimal number of accelerator pulses.

  8. 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

  9. 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.

  10. Expression and localization of the omega-3 fatty acid receptor GPR120 in human term placenta.

    PubMed

    Lager, S; Ramirez, V I; Gaccioli, F; Jansson, T; Powell, T L

    2014-07-01

    Fatty acids can function as signaling molecules, acting through receptors in the cytosol or on the cell surface. G-Protein Receptor (GPR)120 is a membrane-bound receptor mediating anti-inflammatory and insulin-sensitizing effects of the omega-3 fatty acid docohexaenoic acid (DHA). GPR120 dysfunction is associated with obesity in humans. Cellular localization of GPR120 and the influence of maternal obesity on GPR120 protein expression in the placenta are unknown. Herein we demonstrate that GPR120 is predominantly expressed in the microvillous membrane (MVM) of human placenta and that the expression level of this receptor in MVM is not altered by maternal body mass index (BMI).

  11. CMP‑N‑acetylneuraminic acid synthetase interacts with fragile X related protein 1.

    PubMed

    Ma, Yun; Tian, Shuai; Wang, Zongbao; Wang, Changbo; Chen, Xiaowei; Li, Wei; Yang, Yang; He, Shuya

    2016-08-01

    Fragile X mental retardation protein (FMRP), fragile X related 1 protein (FXR1P) and FXR2P are the members of the FMR protein family. These proteins contain two KH domains and a RGG box, which are characteristic of RNA binding proteins. The absence of FMRP, causes fragile X syndrome (FXS), the leading cause of hereditary mental retardation. FXR1P is expressed throughout the body and important for normal muscle development, and its absence causes cardiac abnormality. To investigate the functions of FXR1P, a screen was performed to identify FXR1P‑interacting proteins and determine the biological effect of the interaction. The current study identified CMP‑N‑acetylneuraminic acid synthetase (CMAS) as an interacting protein using the yeast two‑hybrid system, and the interaction between FXR1P and CMAS was validated in yeast using a β‑galactosidase assay and growth studies with selective media. Furthermore, co‑immunoprecipitation was used to analyze the FXR1P/CMAS association and immunofluorescence microscopy was performed to detect expression and intracellular localization of the proteins. The results of the current study indicated that FXR1P and CMAS interact, and colocalize in the cytoplasm and the nucleus of HEK293T and HeLa cells. Accordingly, a fragile X related 1 (FXR1) gene overexpression vector was constructed to investigate the effect of FXR1 overexpression on the level of monosialotetrahexosylganglioside 1 (GM1). The results of the current study suggested that FXR1P is a tissue‑specific regulator of GM1 levels in SH‑SY5Y cells, but not in HEK293T cells. Taken together, the results initially indicate that FXR1P interacts with CMAS, and that FXR1P may enhance the activation of sialic acid via interaction with CMAS, and increase GM1 levels to affect the development of the nervous system, thus providing evidence for further research into the pathogenesis of FXS.

  12. 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.

  13. 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 ...

  14. 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...

  15. Amino acid sequence of the alpha subunit of human leukocyte adhesion receptor Mo1 (complement receptor type 3)

    PubMed Central

    1988-01-01

    Mo1 (complement receptor type 3, CR3; CD11b/CD18) is an adhesion- promoting human leukocyte surface membrane heterodimer (alpha subunit 155 kD [CD11b] noncovalently linked to a beta subunit of 95 kD [CD18]). The complete amino acid sequence deduced from cDNA of the human alpha subunit is reported. The protein consists of 1,136 amino acids with a long amino-terminal extracytoplasmic domain, a 26-amino acid hydrophobic transmembrane segment, and a 19-carboxyl-terminal cytoplasmic domain. The extracytoplasmic region has three putative Ca2+- binding domains with good homology and one with weak homology to the "lock washer" Ca2+-binding consensus sequence. These metal-binding domains explain the divalent cation-dependent functions mediated by Mo1. The alpha subunit is highly homologous to the alpha subunit of leukocyte p150,95 and to a lesser extent, to the alpha subunit of other "integrin" receptors such as fibronectin, vitronectin, and platelet IIb/IIIa receptors in humans and position-specific antigen-2 (PS2) in Drosophila. Mo1 alpha, like p150, contains a unique 187-amino acid stretch NH2-terminal to the metal-binding domains. This region could be involved in some of the specific functions mediated by these leukocyte glycoproteins. PMID:2454931

  16. The inimitable kynurenic acid: the roles of different ionotropic receptors in the action of kynurenic acid at a spinal level.

    PubMed

    Tuboly, Gabor; Tar, Lilla; Bohar, Zsuzsanna; Safrany-Fark, Arpad; Petrovszki, Zita; Kekesi, Gabriella; Vecsei, Laszlo; Pardutz, Arpad; Horvath, Gyongyi

    2015-03-01

    Kynurenic acid (KYNA) is a neuroactive metabolite that interacts with NMDA, AMPA/kainate and alpha 7 nicotinic receptors. The goal of this study was to clarify the roles of these receptors in the action of KYNA at a spinal level by using highly specific receptor antagonists alone or in triple combinations. Chronic osteoarthritis-like joint pain was induced with monosodium-iodoacetate in male Wistar rats. Mechanical allodynia and motor function were quantified. In the first series we determined the dose-response and time course effects of intrathecally administered KYNA (10-100 μg), D-(-)-2-amino-5-phosphonopentanoic acid (AP5; an NMDA receptor antagonist; 10-200 μg), methyllycaconitine (MLA; an alpha 7 nicotinic receptor antagonist; 100-200 μg) and 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzoquinoxaline-7-sulfonamide (NBQX; an AMPA/kainate receptor antagonist; 1-20 μg). In the second series, four different triple combinations of MLA, AP5 and NBQX were investigated. Intrathecal administration of KYNA caused a dose-dependent motor impairment and antinociception. The highly specific NMDA receptor antagonist AP5 caused a motor impairment and antinociception with lower potency. High doses of NBQX resulted in significant antinociception with a slight motor impairment, while only the highest dose of MLA gave rise to significant antinociception with a slight motor impairment. After the coadministration of these ligands as combinations, no potentiation was observed. It may be supposed that the effects of KYNA are primarily due to the inhibition of NMDA receptors at both glycine and phencyclidine (PCP) binding sites, and not to the interactions at the different ionotropic receptors, but the mechanisms behind its high bio-efficiency are still unknown.

  17. Metabolic effects of intestinal absorption and enterohepatic cycling of bile acids

    PubMed Central

    Ferrebee, Courtney B.; Dawson, Paul A.

    2015-01-01

    The classical functions of bile acids include acting as detergents to facilitate the digestion and absorption of nutrients in the gut. In addition, bile acids also act as signaling molecules to regulate glucose homeostasis, lipid metabolism and energy expenditure. The signaling potential of bile acids in compartments such as the systemic circulation is regulated in part by an efficient enterohepatic circulation that functions to conserve and channel the pool of bile acids within the intestinal and hepatobiliary compartments. Changes in hepatobiliary and intestinal bile acid transport can alter the composition, size, and distribution of the bile acid pool. These alterations in turn can have significant effects on bile acid signaling and their downstream metabolic targets. This review discusses recent advances in our understanding of the inter-relationship between the enterohepatic cycling of bile acids and the metabolic consequences of signaling via bile acid-activated receptors, such as farnesoid X nuclear receptor (FXR) and the G-protein-coupled bile acid receptor (TGR5). PMID:26579438

  18. 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.

  19. Stacking interaction and its role in kynurenic acid binding to glutamate ionotropic receptors.

    PubMed

    Zhuravlev, Alexander V; Zakharov, Gennady A; Shchegolev, Boris F; Savvateeva-Popova, Elena V

    2012-05-01

    Stacking interaction is known to play an important role in protein folding, enzyme-substrate and ligand-receptor complex formation. It has been shown to make a contribution into the aromatic antagonists binding with glutamate ionotropic receptors (iGluRs), in particular, the complex of NMDA receptor NR1 subunit with the kynurenic acid (KYNA) derivatives. The specificity of KYNA binding to the glutamate receptors subtypes might partially result from the differences in stacking interaction. We have calculated the optimal geometry and binding energy of KYNA dimers with the four types of aromatic amino acid residues in Rattus and Drosophila ionotropic iGluR subunits. All ab initio quantum chemical calculations were performed taking into account electron correlations at MP2 and MP4 perturbation theory levels. We have also investigated the potential energy surfaces (PES) of stacking and hydrogen bonds (HBs) within the receptor binding site and calculated the free energy of the ligand-receptor complex formation. The energy of stacking interaction depends both on the size of aromatic moieties and the electrostatic effects. The distribution of charges was shown to determine the geometry of polar aromatic ring dimers. Presumably, stacking interaction is important at the first stage of ligand binding when HBs are weak. The freedom of ligand movements and rotation within receptor site provides the precise tuning of the HBs pattern, while the incorrect stacking binding prohibits the ligand-receptor complex formation. PMID:21833825

  20. 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

  1. 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.

  2. Upregulation of retinoic acid receptor-beta by the epidermal growth factor-receptor inhibitor PD153035 is not mediated by blockade of ErbB pathways.

    PubMed

    Grunt, Thomas W; Tomek, Katharina; Wagner, Renate; Puckmair, Klaudia; Kainz, Birgit; Rünzler, Dominik; Gaiger, Alexander; Köhler, Gottfried; Zielinski, Christoph C

    2007-06-01

    Inhibiting epidermal growth factor-receptor (ErbB-1) represents a powerful anticancer strategy. Activation of retinoid pathways is also in development for cancer treatment. Retinoic acid receptor-beta-the tumor suppressor and main retinoid mediator--is silenced in many tumors. The ErbB-1 inhibitor PD153035 cooperates with retinoic acid during growth inhibition and induces retinoic acid receptor-beta suggesting that ErbB-1 controls retinoic acid receptor-beta. However, here we demonstrate that ErbB pathways are not involved in PD153035-mediated retinoic acid receptor-beta-upregulation. PD153035 inhibits ErbB-1-phosphorylation, whereas its derivative EBE-A22 is inactive. Yet both inhibit cell growth and upregulate retinoic acid receptor-beta in ErbB-1-overexpressing (MDA-MB-468), moderately expressing (OVCAR-3), ErbB-1-negative (MDA-MB-453) or ErbB-negative cells (CEM, Jurkat). Both bind DNA, whereas the closely related ErbB-1 inhibitors AG1478 and ZD1839, which are inactive on retinoic acid receptor-beta, do not significantly bind DNA. None of the other ErbB-1/ErbB-2 inhibitors tested (RG-14620, LFM-A12, AG879, AG825) affect retinoic acid receptor-beta. PD153035 decreases methylation of the retinoic acid receptor-beta2 promoter. In OVCAR-3, it stimulates dislodgement of histone deacetylase 1 from the promoter and acetylation of histones H3 and H4. Consequently, PD153035 facilitates recruitment of RNA polymerase II to the promoter and stimulates transcriptional activity. Moreover, PD153035 increases the retinoic acid receptor-beta mRNA half-life. No other retinoid receptor, nor estrogen receptor-alpha, nor RASSF1A is upregulated by PD153035. Thus PD153035 induces retinoic acid receptor-beta by ErbB-independent transcriptional and post-transcriptional mechanisms. This report highlights a triple action for an ErbB-1 inhibitor (ErbB-1 inhibition, DNA intercalation, retinoic acid receptor-beta-induction). Such multitargeting drugs bear great potential for cancer

  3. Phytanic acid and pristanic acid, branched-chain fatty acids associated with Refsum disease and other inherited peroxisomal disorders, mediate intracellular Ca2+ signaling through activation of free fatty acid receptor GPR40.

    PubMed

    Kruska, Nicol; Reiser, Georg

    2011-08-01

    The accumulation of the two branched-chain fatty acids phytanic acid and pristanic acid is known to play an important role in several diseases with peroxisomal impairment, like Refsum disease, Zellweger syndrome and α-methylacyl-CoA racemase deficiency. Recent studies elucidated that the toxic activity of phytanic acid and pristanic acid is mediated by multiple mitochondrial dysfunctions, generation of reactive oxygen species and Ca2+ deregulation via the InsP3-Ca2+ signaling pathway in glial cells. However, the exact signaling mechanism through which both fatty acids mediate toxicity is still under debate. Here, we studied the ability of phytanic acid and pristanic acid to activate the free fatty acid receptor GPR40, a G-protein-coupled receptor, which was described to be involved in the Ca2+ signaling of fatty acids. We treated HEK 293 cells expressing the GPR40 receptor with phytanic acid or pristanic acid. This resulted in a significant increase in the intracellular Ca2+ level, similar to the effect seen after treatment with the synthetic GPR40 agonist GW9508. Furthermore, we demonstrate that the GPR40 activation might be due to an interaction of the carboxylate moiety of fatty acids with the receptor. Our findings indicate that the phytanic acid- and pristanic acid-mediated Ca2+ deregulation can involve the activation of GPR40. Therefore, we suppose that activation of GPR40 might be part of the signaling cascade of the toxicity of phytanic and pristanic acids.

  4. Farnesoid X receptor associates with β-catenin and inhibits its activity in hepatocellular carcinoma

    PubMed Central

    Liu, Xijun; Zhang, Xingwang; Ji, Lingling; Gu, Jianxin; Zhou, Meiling; Chen, She

    2015-01-01

    The association between the temporal activation of Wnt/β-catenin pathway and the spontaneous hepatocellular carcinoma (HCC) development in Farnesoid X receptor (FXR) knockout mice is not well understood. We found that Huh7 cells depleted with FXR by RNAi showed enhanced cell growth, migration and invasion in vitro and accelerated tumor xenografts formation in nude mice. And these phenotypes were attenuated by simultaneous knockdown of β-catenin with RNAi. Furthermore, we identified that FXR could bind with β-Catenin through AF1 domain, and disrupt the assembly of the core β-Catenin/TCF4 complex. Activation of FXR attenuated the DNA-binding activity of β-Catenin/TCF4, and subsequently, its targeting gene-cyclin D1 expression. Importantly, FXR expression was markedly reduced in human HCC, an event which correlated with aberrant activation of β-Catenin. These data identified FXR as a negative regulator of HCC development through direct suppression of Wnt/β-catenin pathway. PMID:25650661

  5. Bovine adenovirus serotype 3 utilizes sialic acid as a cellular receptor for virus entry.

    PubMed

    Li, Xiaoxin; Bangari, Dinesh S; Sharma, Anurag; Mittal, Suresh K

    2009-09-30

    Bovine adenovirus serotype 3 (BAd3) and porcine adenovirus serotype 3 (PAd3) entry into the host cells is independent of Coxsackievirus adenovirus receptor and integrins. The role of sialic acid in BAd3 and PAd3 entry was investigated. Removal of sialic acid by neuraminidase, or blocking sialic acid by wheat germ agglutinin lectin significantly inhibited BAd3, but not PAd3, transduction of Madin-Darby bovine kidney cells. Maackia amurensis agglutinin or Sambucus nigra (elder) agglutinin treatment efficiently blocked BAd3 transduction suggesting that BAd3 utilized alpha(2,3)-linked and alpha(2,6)-linked sialic acid as a cell receptor. BAd3 transduction of MDBK cells was sensitive to sodium periodate, bromelain, or trypsin treatment indicating that the receptor sialoconjugate was a glycoprotein rather than a ganglioside. To determine sialic acid-containing cell membrane proteins that bind to BAd3, virus overlay protein binding assay (VOPBA) was performed and showed that sialylated cell membrane proteins in size of approximately 97 and 34 kDa bind to BAd3. The results suggest that sialic acid serves as a primary receptor for BAd3.

  6. [Discovery of potential nicotinic acid receptor agonists from Chinese herbal medicines based on molecular simulation].

    PubMed

    Jiang, Lu-Di; He, Yu-Su; Zhang, Yan-Ling

    2014-12-01

    Nicotinic acid could increase high density lipoprotein and reduce serum total cholesterol, low density lipoprotein cholesterol and triglycerides in human bodies, thus is frequently applied in treating low high-density lipoprotein cholesterol and hypertriglyceridemia in clinic. However, according to the findings, nicotinic acid could also cause adverse effects, such as skin flush, beside its curative effects. In this study, bioisosterism, fragment-based search and Lipinski's Rule of Five were used to preliminarily screen out potential TCM ingredients that may have similar pharmacological effects with nicotinic acid from Traditional Chinese medicine database (TCMD). Afterwards, homology modeling and flexible docking were used to further screen out potential nicotinic acid receptor agonists. As a result, eleven candidate compounds were derived from eight commonly used traditional Chinese medicines. Specifically, all of the candidate compounds' interaction with nicotinic acid receptor was similar to nicotinic acid, and their docking scores were all higher than that of nicotinic acid, but their druggability remained to be further studied. Some of the eight source traditional Chinese medicines were used to lower lipid according to literature studies, implying that they may show effect through above means. In summary, this study provides basis and reference for extracting new nicotinic acid receptor agonists from traditional Chinese medicines and improving the medication status of hyperlipidemia.

  7. A Mollusk Retinoic Acid Receptor (RAR) Ortholog Sheds Light on the Evolution of Ligand Binding

    PubMed Central

    Gutierrez-Mazariegos, Juliana; Nadendla, Eswar Kumar; Lima, Daniela; Pierzchalski, Keely; Jones, Jace W.; Kane, Maureen; Nishikawa, Jun-Ichi; Hiromori, Youhei; Nakanishi, Tsuyoshi; Santos, Miguel M.; Castro, L. Filipe C.; Bourguet, William

    2014-01-01

    Nuclear receptors are transcription factors that regulate networks of target genes in response to small molecules. There is a strong bias in our knowledge of these receptors because they were mainly characterized in classical model organisms, mostly vertebrates. Therefore, the evolutionary origins of specific ligand-receptor couples still remain elusive. Here we present the identification and characterization of a retinoic acid receptor (RAR) from the mollusk Nucella lapillus (NlRAR). We show that this receptor specifically binds to DNA response elements organized in direct repeats as a heterodimer with retinoid X receptor. Surprisingly, we also find that NlRAR does not bind all-trans retinoic acid or any other retinoid we tested. Furthermore, NlRAR is unable to activate the transcription of reporter genes in response to stimulation by retinoids and to recruit coactivators in the presence of these compounds. Three-dimensional modeling of the ligand-binding domain of NlRAR reveals an overall structure that is similar to vertebrate RARs. However, in the ligand-binding pocket (LBP) of the mollusk receptor, the alteration of several residues interacting with the ligand has apparently led to an overall decrease in the strength of the interaction with the ligand. Accordingly, mutations of NlRAR at key positions within the LBP generate receptors that are responsive to retinoids. Altogether our data suggest that, in mollusks, RAR has lost its affinity for all-trans retinoic acid, highlighting the evolutionary plasticity of its LBP. When put in an evolutionary context, our results reveal new structural and functional features of nuclear receptors validated by millions of years of evolution that were impossible to reveal in model organisms. PMID:25116705

  8. Imaging Cancer Cells Expressing the Folate Receptor with Carbon Dots Produced from Folic Acid.

    PubMed

    Bhunia, Susanta Kumar; Maity, Amit Ranjan; Nandi, Sukhendu; Stepensky, David; Jelinek, Raz

    2016-04-01

    Development of new imaging tools for cancer cells in vitro and in vitro is important for advancing cancer research, elucidating drug effects upon cancer cells, and studying cellular processes. We showed that fluorescent carbon dots (C-dots) synthesized from folic acid can serve as an effective vehicle for imaging cancer cells expressing the folate receptor on their surface. The C-dots, synthesized through a simple one-step process from folic acid as the carbon source, exhibited selectivity towards cancer cells displaying the folate receptor, making such cells easily distinguishable in fluorescence microscopy imaging. Biophysical measurements and competition experiments both confirmed the specific targeting and enhanced uptake of C-dots by the folate receptor-expressing cells. The folic acid-derived C-dots were not cytotoxic, and their use in bioimaging applications could aid biological studies of cancer cells, identification of agonists/antagonists, and cancer diagnostics.

  9. TRIP6 Enhances Lysophosphatidic Acid-induced Cell Migration by Interacting with the Lysophosphatidic Acid 2 Receptor*

    PubMed Central

    Xu, Jun; Lai, Yun-Ju; Lin, Weei-Chin; Lin, Fang-Tsyr

    2014-01-01

    Lysophosphatidic acid (LPA) induces actin rearrangement, focal adhesion assembly, and cell migration through the activation of small G protein Rho and its downstream effectors. These diverse cellular responses are mediated by its associated G protein-coupled receptors. However, the mechanisms and specificity by which these LPA receptors mediate LPA actions are still poorly understood. Here we show that LPA stimulation promotes the interaction of the LPA2 receptor with a focal adhesion molecule, TRIP6 (thyroid receptor interacting protein 6)/ZRP-1 (zyxin-related protein 1). TRIP6 directly binds to the carboxyl-terminal tail of the LPA2 receptor through its LIM domains. LPA-dependent recruitment of TRIP6 to the plasma membrane promotes its targeting to focal adhesions and co-localization with actin stress fibers. In addition, TRIP6 associates with the components of focal complexes including paxillin, focal adhesion kinase, c-Src, and p130cas in an agonist-dependent manner. Overexpression of TRIP6 augments LPA-induced cell migration; in contrast, suppression of endogenous TRIP6 expression by a TRIP6-specific small interfering RNA reduces it in SKOV3 ovarian cancer cells. Strikingly, the association with TRIP6 is specific to the LPA2 receptor but not LPA1 or LPA3 receptor, indicating a specific role for TRIP6 in regulating LPA2 receptor-mediated signaling. Taken together, our results suggest that TRIP6 functions at a point of convergence between the activated LPA2 receptor and downstream signals involved in cell adhesion and migration. PMID:14688263

  10. 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

  11. The bile acid membrane receptor TGR5 as an emerging target in metabolism and inflammation.

    PubMed

    Pols, Thijs W H; Noriega, Lilia G; Nomura, Mitsunori; Auwerx, Johan; Schoonjans, Kristina

    2011-06-01

    Bile acids (BAs) are amphipathic molecules that facilitate the uptake of lipids, and their levels fluctuate in the intestine as well as in the blood circulation depending on food intake. Besides their role in dietary lipid absorption, bile acids function as signaling molecules capable to activate specific receptors. These BA receptors are not only important in the regulation of bile acid synthesis and their metabolism, but also regulate glucose homeostasis, lipid metabolism, and energy expenditure. These processes are important in diabetes and other facets of the metabolic syndrome, which represents a considerable increasing health burden. In addition to the function of the nuclear receptor FXRα in regulating local effects in the organs of the enterohepatic axis, increasing evidence points to a crucial role of the G-protein coupled receptor (GPCR) TGR5 in mediating systemic actions of BAs. Here we discuss the current knowledge on BA receptors, with a strong focus on the cell membrane receptor TGR5, which emerges as a valuable target for intervention in metabolic diseases. PMID:21145931

  12. Free fatty acids and protein kinase C activation induce GPR120 (free fatty acid receptor 4) phosphorylation.

    PubMed

    Sánchez-Reyes, Omar B; Romero-Ávila, M Teresa; Castillo-Badillo, Jean A; Takei, Yoshinori; Hirasawa, Akira; Tsujimoto, Gozoh; Villalobos-Molina, Rafael; García-Sáinz, J Adolfo

    2014-01-15

    GPR120, free fatty acid receptor 4, is a recently deorphanized G protein-coupled receptor that seems to play cardinal roles in the regulation of metabolism and in the pathophysiology of inflammatory and metabolic disorders. In the present work a GPR120-Venus fusion protein was expressed in HEK293 Flp-In T-REx cells and its function (increase in intracellular calcium) and phosphorylation were studied. It was observed that the fusion protein migrated in sodium dodecyl sulfate-polyacrylamide gels as a band with a mass of ≈70-75kDa, although other bands of higher apparent weight (>130kDa) were also detected. Cell stimulation with docosahexaenoic acid or α-linolenic acid induced concentration-dependent increases in intracellular calcium and GPR120 phosphorylation. Activation of protein kinase C with phorbol esters also induced a marked receptor phosphorylation but did not alter the ability of 1µM docosahexaenoic acid to increase the intracellular calcium concentration. Phorbol ester-induced GPR120 phosphorylation, but not that induced with docosahexaenoic acid, was blocked by protein kinase C inhibitors (bis-indolyl-maleimide I and Gö 6976) suggesting that conventional kinase isoforms mediate this action. The absence of effect of protein kinase C inhibitors on agonist-induced GPR120 phosphorylation indicates that this kinase does not play a major role in agonist-induced receptor phosphorylation. Docosahexaenoic acid action was associated with marked GPR120 internalization whereas that induced with phorbol esters was smaller at early times. PMID:24239485

  13. Statistical Mechanics Model for the Interaction between the Neurotransmitter γ-Aminobutyric acid and GABAA Receptors

    NASA Astrophysics Data System (ADS)

    Zafar, Sufi; Saxena, Nina C.; Conrad, Kevin A.; Hussain, Arif

    2004-07-01

    Interactions between the neurotransmitter γ-aminobutyric acid (GABA) and GABAA receptor ion channels play an important role in the central nervous system. A statistical mechanics model is proposed for the interaction between GABA and GABAA receptors. The model provides good fits to the electrophysiology data as well as an estimation of receptor activation energies, and predicts the temperature dependence consistent with measurements. In addition, the model provides insights into single channel conductance measurements. This model is also applicable to other ligand-gated ion channels with similar pentameric structures.

  14. Site-specific incorporation of keto amino acids into functional G protein-coupled receptors using unnatural amino acid mutagenesis.

    PubMed

    Ye, Shixin; Köhrer, Caroline; Huber, Thomas; Kazmi, Manija; Sachdev, Pallavi; Yan, Elsa C Y; Bhagat, Aditi; RajBhandary, Uttam L; Sakmar, Thomas P

    2008-01-18

    G protein-coupled receptors (GPCRs) are ubiquitous heptahelical transmembrane proteins involved in a wide variety of signaling pathways. The work described here on application of unnatural amino acid mutagenesis to two GPCRs, the chemokine receptor CCR5 (a major co-receptor for the human immunodeficiency virus) and rhodopsin (the visual photoreceptor), adds a new dimension to studies of GPCRs. We incorporated the unnatural amino acids p-acetyl-L-phenylalanine (Acp) and p-benzoyl-L-phenylalanine (Bzp) into CCR5 at high efficiency in mammalian cells to produce functional receptors harboring reactive keto groups at three specific positions. We obtained functional mutant CCR5, at levels up to approximately 50% of wild type as judged by immunoblotting, cell surface expression, and ligand-dependent calcium flux. Rhodopsin containing Acp at three different sites was also purified in high yield (0.5-2 microg/10(7) cells) and reacted with fluorescein hydrazide in vitro to produce fluorescently labeled rhodopsin. The incorporation of reactive keto groups such as Acp or Bzp into GPCRs allows their reaction with different reagents to introduce a variety of spectroscopic and other probes. Bzp also provides the possibility of photo-cross-linking to identify precise sites of protein-protein interactions, including GPCR binding to G proteins and arrestins, and for understanding the molecular basis of ligand recognition by chemokine receptors. PMID:17993461

  15. A differential fluorescent receptor for nucleic acid analysis.

    PubMed

    Bengtson, Hillary N; Kolpashchikov, Dmitry M

    2014-01-24

    Differential receptors use an array of sensors to recognize analytes. Each sensor in the array can recognize not one, but several analytes with different rates, so a single analyte triggers a response of several sensors in the array. The receptor thus produces a pattern of signals that is unique for each analyte, thereby enabling identification of a specific analyte by producing a "fingerprint" pattern. We applied this approach for the analysis of DNA sequences of Mycobacterium tuberculosis strains that differ by single nucleotide substitutions in the 81-bp hot-spot region that imparts rifampin resistance. The technology takes advantage of the new multicomponent, selfassembling sensor, which produces a fluorescent signal in the presence of specific DNA sequences. A differential fluorescent receptor (DFR) contained an array of three such sensors and differentiated at least eight DNA sequences. The approach requires only one molecular-beacon-like fluorescent reporter, which can be used by all three sensors. The DFR developed in this study represents a cost-efficient alternative to molecular diagnostic technologies that use fluorescent hybridization probes.

  16. Clustering Nuclear Receptors in Liver Regeneration Identifies Candidate Modulators of Hepatocyte Proliferation and Hepatocarcinoma

    PubMed Central

    Graziano, Giusi; D'Orazio, Andria; Cariello, Marica; Massafra, Vittoria; Salvatore, Lorena; Martelli, Nicola; Murzilli, Stefania; Sasso, Giuseppe Lo; Mariani-Costantini, Renato; Moschetta, Antonio

    2014-01-01

    Background & Aims Liver regeneration (LR) is a valuable model for studying mechanisms modulating hepatocyte proliferation. Nuclear receptors (NRs) are key players in the control of cellular functions, being ideal modulators of hepatic proliferation and carcinogenesis. Methods & Results We used a previously validated RT-qPCR platform to profile modifications in the expression of all 49 members of the NR superfamily in mouse liver during LR. Twenty-nine NR transcripts were significantly modified in their expression during LR, including fatty acid (peroxisome proliferator-activated receptors, PPARs) and oxysterol (liver X receptors, Lxrs) sensors, circadian masters RevErbα and RevErbβ, glucocorticoid receptor (Gr) and constitutive androxane receptor (Car). In order to detect the NRs that better characterize proliferative status vs. proliferating liver, we used the novel Random Forest (RF) analysis to selected a trio of down-regulated NRs (thyroid receptor alpha, Trα; farsenoid X receptor beta, Fxrβ; Pparδ) as best discriminators of the proliferating status. To validate our approach, we further studied PPARδ role in modulating hepatic proliferation. We first confirmed the suppression of PPARδ both in LR and human hepatocellular carcinoma at protein level, and then demonstrated that PPARδ agonist GW501516 reduces the proliferative potential of hepatoma cells. Conclusions Our data suggest that NR transcriptome is modulated in proliferating liver and is a source of biomarkers and bona fide pharmacological targets for the management of liver disease affecting hepatocyte proliferation. PMID:25116592

  17. Regulation of Expression of Citrate Synthase by the Retinoic Acid Receptor-Related Orphan Receptor α (RORα)

    PubMed Central

    Crumbley, Christine; Wang, Yongjun; Banerjee, Subhashis; Burris, Thomas P.

    2012-01-01

    The retinoic acid receptor-related orphan receptor α (RORα) is a member of the nuclear receptor superfamily of transcription factors that plays an important role in regulation of the circadian rhythm and metabolism. Mice lacking a functional RORα display a range of metabolic abnormalities including decreased serum cholesterol and plasma triglycerides. Citrate synthase (CS) is a key enzyme of the citric acid cycle that provides energy for cellular function. Additionally, CS plays a critical role in providing citrate derived acetyl-CoA for lipogenesis and cholesterologenesis. Here, we identified a functional RORα response element (RORE) in the promoter of the CS gene. ChIP analysis demonstrates RORα occupancy of the CS promoter and a putative RORE binds to RORα effectively in an electrophoretic mobility shift assay and confers RORα responsiveness to a reporter gene in a cotransfection assay. We also observed a decrease in CS gene expression and CS enzymatic activity in the staggerer mouse, which has a mutation of in the Rora gene resulting in nonfunctional RORα protein. Furthermore, we found that SR1001 a RORα inverse agonist eliminated the circadian pattern of expression of CS mRNA in mice. These data suggest that CS is a direct RORα target gene and one mechanism by which RORα regulates lipid metabolism is via regulation of CS expression. PMID:22485150

  18. Amiloride inhibition of gamma-aminobutyric acid(A) receptors depends upon the alpha subunit subtype.

    PubMed

    Fisher, Janet L

    2002-06-01

    gamma-Aminobutyric acid(A) (GABA(A)) receptors (GABARs) are responsible for most fast inhibitory neurotransmission in the mammalian brain. The GABARs contain several allosteric modulatory sites, many of which are useful clinically. The activity of most of these modulators depends upon the subunit composition of the receptor. The diuretic amiloride was previously reported to inhibit GABARs in frog sensory neurons. We measured its effects on recombinant GABARs to determine its mechanism of action at mammalian receptors and to examine the effect of subunit composition. Amiloride acted primarily as a competitive antagonist, reducing the sensitivity of the receptor to GABA without affecting the maximal current amplitude. Receptors containing an alpha6 subunit were about 10-fold more sensitive to amiloride than those containing other alpha subunits. In contrast, the identity of the beta or gamma subtype had little effect on amiloride sensitivity. Although several other modulators have specific effects at alpha6-containing receptors, amiloride is the first inhibitor to be reported with no additional dependence on the identity of the beta or gamma subunit. Therefore, it probably represents a unique modulatory site on the GABAR, which could be useful for developing drugs targeting these receptors. The selective activity of amiloride could also be helpful for isolating the contribution of receptors composed of alpha6 subtypes in heterogeneous native GABAR populations.

  19. Structural basis for molecular recognition of folic acid by folate receptors.

    PubMed

    Chen, Chen; Ke, Jiyuan; Zhou, X Edward; Yi, Wei; Brunzelle, Joseph S; Li, Jun; Yong, Eu-Leong; Xu, H Eric; Melcher, Karsten

    2013-08-22

    Folate receptors (FRα, FRβ and FRγ) are cysteine-rich cell-surface glycoproteins that bind folate with high affinity to mediate cellular uptake of folate. Although expressed at very low levels in most tissues, folate receptors, especially FRα, are expressed at high levels in numerous cancers to meet the folate demand of rapidly dividing cells under low folate conditions. The folate dependency of many tumours has been therapeutically and diagnostically exploited by administration of anti-FRα antibodies, high-affinity antifolates, folate-based imaging agents and folate-conjugated drugs and toxins. To understand how folate binds its receptors, we determined the crystal structure of human FRα in complex with folic acid at 2.8 Å resolution. FRα has a globular structure stabilized by eight disulphide bonds and contains a deep open folate-binding pocket comprised of residues that are conserved in all receptor subtypes. The folate pteroate moiety is buried inside the receptor, whereas its glutamate moiety is solvent-exposed and sticks out of the pocket entrance, allowing it to be conjugated to drugs without adversely affecting FRα binding. The extensive interactions between the receptor and ligand readily explain the high folate-binding affinity of folate receptors and provide a template for designing more specific drugs targeting the folate receptor system.

  20. 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

  1. Characterization of cDNAs encoding the chick retinoic acid receptor gamma 2 and preferential distribution of retinoic acid receptor gamma transcripts during chick skin development.

    PubMed

    Michaille, J J; Blanchet, S; Kanzler, B; Garnier, J M; Dhouailly, D

    1994-12-01

    Retinoic acid receptors alpha, beta and gamma (RAR alpha, beta and gamma) are ligand-inductible transcriptional activators which belong to the steroid/thyroid hormone receptor superfamily. At least two major isoforms (1 and 2) of each RAR arise by differential use of two promoters and alternative splicing. In mouse, the three RAR genes are expressed in stage- and tissue-specific patterns during embryonic development. In order to understand the role of the different RARs in chick, RAR gamma 2 cDNAs were isolated from an 8.5-day (stage 35 of Hamburger and Hamilton) chick embryo skin library. The deduced chick RAR gamma 2 amino acid sequence displays uncommon features such as 21 specific amino acid replacements, 12 of them being clustered in the amino-terminal region (domains A2 and B), and a truncated acidic carboxy-terminal region (F domain). However, the pattern of RAR gamma expression in chick embryo resembles that reported in mouse, particularly in skin where RAR gamma expression occurs in both the dermal and epidermal layers at the beginning of feather formation, and is subsequently restricted to the differentiating epidermal cells. Northern blot analysis suggests that different RAR gamma isoforms could be successively required during chick development.

  2. Minireview: The Effects of Species Ortholog and SNP Variation on Receptors for Free Fatty Acids

    PubMed Central

    Hudson, Brian D.; Murdoch, Hannah

    2013-01-01

    Although it is widely assumed that species orthologs of hormone-responsive G protein-coupled receptors will be activated by the same endogenously produced ligand(s), variation in potency, particularly in cases in which more than 1 receptor responds to the same hormone, can result in challenges in defining the contribution of individual receptors in different species. This can create considerably greater issues when using synthetic chemical ligands and, in some cases, may result in a complete lack of efficacy of such a ligand when used in animal models of pathophysiology. In man, the concept that distinct responses of individuals to medicines may reflect differences in the ability of such drugs to bind to or activate single nucleotide polymorphism variants of receptors is more established as a concept but, in many cases, clear links between such variants that are associated with disease phenotypes and substantial differences in receptor ligand pharmacology have been more difficult to obtain. Herein we consider each of these issues for the group of free fatty acid receptors, FFA1-FFA4, defined to be activated by free fatty acids of varying chain length, which, based on their production by 1 tissue or location and action in distinct locations, have been suggested to possess characteristics of hormones. PMID:23686113

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

    PubMed Central

    Brighton, Cheryl A.; Rievaj, Juraj; Kuhre, Rune E.; Glass, Leslie L.; Schoonjans, Kristina; Holst, Jens J.

    2015-01-01

    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 Ca2+. In primary intestinal cultures, TDCA was a more potent GLP-1 secretagogue than taurocholate (TCA) and TLCA, correlating with a stronger Ca2+ 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. PMID:26280129

  4. 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.

  5. 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. PMID:26597701

  6. Ubiquitin/proteasome pathway regulates levels of retinoic acid receptor gamma and retinoid X receptor alpha in human keratinocytes.

    PubMed

    Boudjelal, M; Wang, Z; Voorhees, J J; Fisher, G J

    2000-04-15

    Repeated exposure of human skin to solar UV radiation leads to premature aging (photoaging) and skin cancer. UV-induced skin damage can be ameliorated by all-trans retinoic acid treatment. The actions of retinoic acid in skin keratinocytes are mediated primarily by nuclear retinoic acid receptor gamma (RARgamma) and retinoid X receptor alpha (RXRalpha). We found that exposure of cultured primary human keratinocytes to UV irradiation (30 mJ/cm2) substantially reduced (50-90%) RARgamma and RXRalpha mRNA and protein within 8 h. The rates of disappearance of RARgamma and RXRalpha proteins after UV exposure or treatment with the protein synthesis inhibitor cycloheximide were similar. UV irradiation did not increase the rate of breakdown of RARgamma or RXRalpha but rather reduced their rate of synthesis. The addition of proteasome inhibitors MG132 and LLvL, but not the lysosomal inhibitor E64, prevented loss of RARgamma and RXRalpha proteins after exposure of keratinocytes to either UV radiation or cycloheximide. Soluble extracts from nonirradiated or UV-irradiated keratinocytes possessed similar levels of proteasome activity that degraded RARgamma and RXRalpha proteins in vitro. Furthermore, RARgamma and RXRalpha were polyubiquitinated in intact cells. RXRalpha was found to contain two proline, glutamate/aspartate, serine, and threonine (PEST) motifs, which confer rapid turnover of many short-lived regulatory proteins that are degraded by the ubiquitin/proteasome pathway. However, the PEST motifs in RXRalpha did not function to regulate its stability, because deletion of the PEST motifs individually or together did not alter ubiquitination or proteasome-mediated degradation of RXRalpha. These results demonstrate that loss of RARgamma and RXRalpha proteins after UV irradiation results from degradation via the ubiquitin/proteasome pathway. Taken together, the data here indicate that ubiquitin/proteasome-mediated breakdown is an important mechanism regulating the levels of

  7. 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. PMID:25200370

  8. 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.

  9. Myeloid differentiation and retinoblastoma phosphorylation changes in HL-60 cells induced by retinoic acid receptor- and retinoid X receptor-selective retinoic acid analogs.

    PubMed

    Brooks, S C; Kazmer, S; Levin, A A; Yen, A

    1996-01-01

    The ability of subtypes of retinoic acid receptors (RARs) and retinoid X receptors (RXRs) singly and in combination to elicit myeloid differentiation, G1/0-specific growth arrest, and retinoblastoma (RB) tumor suppressor protein dephosphorylation was determined in the human myeloblastic leukemia cell line HL-60 using subtype-selective retinoic acid (RA) analogs. RA analogs that selectively bind only to RARs (Am580 and/or TTNPB) or to RXRs (Ro 25-6603, SR11237, and/or SR11234) did not elicit the above-mentioned three cellular responses. In contrast, simultaneous treatment with both an RAR-selective ligand (Am580 or TTNPB) and an RXR-selective ligand (Ro 25-6603, SR11237, or SR11234) induced all three cellular processes. An RAR alpha-selective ligand used with an RXR-selective ligand generated the same responses as did all-trans RA or 9-cis RA, which affect both families of receptors, suggesting an important role for RAR alpha among RAR subtypes in eliciting cellular response. Consistent with this finding, the RAR alpha antagonist, Ro 41-5253, reduced the level of the cellular responses elicited by treatment with an RAR alpha-selective ligand plus RXR-selective ligand. The coupling of the shift of RB to its hypophosphorylated form with G1/0 arrest and differentiation in response to ligands is consistent with a possible role of RB as a downstream target or effector of RAR alpha and RXR in combination.

  10. Γ-aminobutyric acid receptors affect the progression and migration of tumor cells.

    PubMed

    Zhang, Xiaoxue; Du, Zuoyi; Liu, Jun; He, Jianxing

    2014-12-01

    Γ-aminobutyric acid (GABA) is a multifunctional molecule found in the nervous system and non-neuronal tissues. GABA receptors combine with GABA molecules and transmit signal stimuli into cells. In addition to traditional neurotransmission and regulation of secretion, GABA and GABA receptors are involved in cell differentiation and proliferation throughout peripheral organs, as well as in tumorigenesis. The exact mechanism of the GABAergic system in regulating tumor development is unclear, but many studies have revealed that GABA receptors exert critical regulative effects on tumor cell proliferation and migration. In this review, the molecular structure, distribution and biological function of GABA receptors associated with tumorigenesis are described. Recent advances in the elucidation of mechanisms underlying GABAergic signaling control over tumor growth are also discussed.

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

    PubMed Central

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

    2015-01-01

    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. PMID:25584517

  12. Specificity of the antibody receptor site to D-lysergamide: model of a physiological receptor for lysergic acid diethylamide.

    PubMed

    Van Vunakis, H; Farrow, J T; Gjika, H B; Levine, L

    1971-07-01

    Antibodies to D-lysergic acid have been produced in rabbits and guinea pigs and a radioimmunoassay for the hapten was developed. The specificity of this lysergamide-antilysergamide reaction was determined by competitive binding with unlabeled lysergic acid diethylamide (LSD), psychotomimetic drugs, neurotransmitters, and other compounds with diverse structures. LSD and several related ergot alkaloids were potent competitors, three to seven times more potent than lysergic acid itself. The N,N-dimethyl derivatives of several compounds, including tryptamine, 5-hydroxytryptamine, 4-hydroxytryptamine, 5-methoxytryptamine, tyramine, and mescaline, were only about ten times less effective than lysergic acid, even though these compounds lack some of the ring systems of lysergic acid. The pattern of inhibition by related compounds with various substituents suggests that the antibody receptor site recognizes structural features resembling the LSD molecule. In particular, the aromatic nucleus and the dimethylated ethylamine side chain in phenylethylamine and tryptamine derivatives may assume in solution a conformation resembling ring A and the methylated nitrogen in ring C of LSD. Among the tryptamine derivatives, a large percentage of the most potent competitors are also psychotomimetic compounds.

  13. 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.

  14. Phosphorylation and Internalization of Lysophosphatidic Acid Receptors LPA1, LPA2, and LPA3

    PubMed Central

    Alcántara-Hernández, Rocío; Hernández-Méndez, Aurelio; Campos-Martínez, Gisselle A.; Meizoso-Huesca, Aldo; García-Sáinz, J. Adolfo

    2015-01-01

    Results The lysophosphatidic acid receptors LPA1, LPA2, and LPA3 were individually expressed in C9 cells and their signaling and regulation were studied. Agonist-activation increases intracellular calcium concentration in a concentration-dependent fashion. Phorbol myristate acetate markedly inhibited LPA1- and LPA3-mediated effect, whereas that mediated by LPA2 was only partially diminished; the actions of the phorbol ester were inhibited by bisindolylmaleimide I and by overnight incubation with the protein kinase C activator, which leads to down regulation of this protein kinase. Homologous desensitization was also observed for the three LPA receptors studied, with that of LPA2 receptors being consistently of lesser magnitude; neither inhibition nor down-regulation of protein kinase C exerted any effect on homologous desensitization. Activation of LPA1–3 receptors induced ERK 1/2 phosphorylation; this effect was markedly attenuated by inhibition of epidermal growth factor receptor tyrosine kinase activity, suggesting growth factor receptor transactivation in this effect. Lysophosphatidic acid and phorbol myristate acetate were able to induce LPA1–3 phosphorylation, in time- and concentration-dependent fashions. It was also clearly observed that agonists and protein kinase C activation induced internalization of these receptors. Phosphorylation of the LPA2 subtype required larger concentrations of these agents and its internalization was less intense than that of the other subtypes. Conclusion Our data show that these three LPA receptors are phosphoproteins whose phosphorylation state is modulated by agonist-stimulation and protein kinase C-activation and that differences in regulation and cellular localization exist, among the subtypes. PMID:26473723

  15. Receptor-mediated uptake of low density lipoprotein stimulates bile acid synthesis by cultured rat hepatocytes

    SciTech Connect

    Junker, L.H.; Davis, R.A. )

    1989-12-01

    The cellular mechanisms responsible for the lipoprotein-mediated stimulation of bile acid synthesis in cultured rat hepatocytes were investigated. Adding 280 micrograms/ml of cholesterol in the form of human or rat low density lipoprotein (LDL) to the culture medium increased bile acid synthesis by 1.8- and 1.6-fold, respectively. As a result of the uptake of LDL, the synthesis of (14C)cholesterol from (2-14C)acetate was decreased and cellular cholesteryl ester mass was increased. Further studies demonstrated that rat apoE-free LDL and apoE-rich high density lipoprotein (HDL) both stimulated bile acid synthesis 1.5-fold, as well as inhibited the formation of (14C)cholesterol from (2-14C)acetate. Reductive methylation of LDL blocked the inhibition of cholesterol synthesis, as well as the stimulation of bile acid synthesis, suggesting that these processes require receptor-mediated uptake. To identify the receptors responsible, competitive binding studies using 125I-labeled apoE-free LDL and 125I-labeled apoE-rich HDL were performed. Both apoE-free LDL and apoE-rich HDL displayed an equal ability to compete for binding of the other, suggesting that a receptor or a group of receptors that recognizes both apolipoproteins is involved. Additional studies show that hepatocytes from cholestyramine-treated rats displayed 2.2- and 3.4-fold increases in the binding of apoE-free LDL and apoE-rich HDL, respectively. These data show for the first time that receptor-mediated uptake of LDL by the liver is intimately linked to processes activating bile acid synthesis.

  16. Farnesoid X receptor inhibits tamoxifen-resistant MCF-7 breast cancer cell growth through downregulation of HER2 expression.

    PubMed

    Giordano, C; Catalano, S; Panza, S; Vizza, D; Barone, I; Bonofiglio, D; Gelsomino, L; Rizza, P; Fuqua, S A W; Andò, S

    2011-09-29

    Tamoxifen (Tam) treatment is a first-line endocrine therapy for estrogen receptor-α-positive breast cancer patients. Unfortunately, resistance frequently occurs and is often related with overexpression of the membrane tyrosine kinase receptor HER2. This is the rationale behind combined treatments with endocrine therapy and novel inhibitors that reduce HER2 expression and signaling and thus inhibit Tam-resistant breast cancer cell growth. In this study, we show that activation of farnesoid X receptor (FXR), by the primary bile acid chenodeoxycholic acid (CDCA) or the synthetic agonist GW4064, inhibited growth of Tam-resistant breast cancer cells (termed MCF-7 TR1), which was used as an in vitro model of acquired Tam resistance. Our results demonstrate that CDCA treatment significantly reduced both anchorage-dependent and anchorage-independent epidermal growth factor (EGF)-induced growth in MCF-7 TR1 cells. Furthermore, results from western blot analysis and real-time reverse transcription-PCR revealed that CDCA treatment reduced HER2 expression and inhibited EGF-mediated HER2 and p42/44 mitogen-activated protein kinase (MAPK) phosphorylation in these Tam-resistant breast cancer cells. Transient transfection experiments, using a vector containing the human HER2 promoter region, showed that CDCA treatment downregulated basal HER2 promoter activity. This occurred through an inhibition of nuclear factor-κB transcription factor binding to its specific responsive element located in the HER2 promoter region as revealed by mutagenesis studies, electrophoretic mobility shift assay and chromatin immunoprecipitation analysis. Collectively, these data suggest that FXR ligand-dependent activity, blocking HER2/MAPK signaling, may overcome anti-estrogen resistance in human breast cancer cells and could represent a new therapeutic tool to treat breast cancer patients that develop resistance.

  17. Intracrine prostaglandin E(2) signalling regulates hypoxia-inducible factor-1α expression through retinoic acid receptor-β.

    PubMed

    Fernández-Martínez, Ana B; Jiménez, María I Arenas; Manzano, Victoria Moreno; Lucio-Cazaña, Francisco J

    2012-12-01

    We have previously found in human renal proximal tubular HK-2 cells that hypoxia- and all-trans retinoic acid-induced hypoxia-inducible factor-1α up-regulation is accompanied by retinoic acid receptor-β up-regulation. Here we first investigated whether hypoxia-inducible factor-1α expression is dependent on retinoic acid receptor-β and our results confirmed it since (i) hypoxia-inducible factor-1α-inducing agents hypoxia, hypoxia-mimetic agent desferrioxamine, all-trans retinoic acid and interleukin-1β increased retinoic acid receptor-β expression, (ii) hypoxia-inducible factor-1α up-regulation was prevented by retinoic acid receptor-β antagonist LE-135 or siRNA retinoic acid receptor-β and (iii) there was direct binding of retinoic acid receptor-β to the retinoic acid response element in hypoxia-inducible factor-1α promoter upon treatment with all-trans retinoic acid and 16,16-dimethyl-prostaglandin E(2). Since intracellular prostaglandin E(2) mediates hypoxia-inducible factor-1α up-regulation in normoxia in HK-2 cells, we next investigated and confirmed, its role in the up-regulation of retinoic acid receptor-β in normoxia by hypoxia-inducible factor-1α-inducing agents all-trans retinoic acid, interleukin-1β and 16,16-dimethyl-prostaglandin E(2) by inhibiting cyclooxygenases, prostaglandin influx transporter or EP receptors. Interestingly, the hypoxia-induced increase in retinoic acid receptor-β expression and accumulation of hypoxia-inducible factor-1α was also blocked by the inhibitors tested. This is the first time, to our knowledge, that retinoic acid receptor-β signalling is involved in the control of the expression of transcription factor hypoxia-inducible factor-1α in both normoxia and hypoxia and that retinoic acid receptor-β expression is found to be strictly regulated by intracellular prostaglandin E(2). Given the relevance of hypoxia-inducible factor-1α in the kidney in terms of tumorigenesis, progressive renal failure, production

  18. 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. PMID:25881750

  19. 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.

  20. The NHR-8 nuclear receptor regulates cholesterol and bile acid homeostasis in C. elegans.

    PubMed

    Magner, Daniel B; Wollam, Joshua; Shen, Yidong; Hoppe, Caroline; Li, Dongling; Latza, Christian; Rottiers, Veerle; Hutter, Harald; Antebi, Adam

    2013-08-01

    Hormone-gated nuclear receptors (NRs) are conserved transcriptional regulators of metabolism, reproduction, and homeostasis. Here we show that C. elegans NHR-8 NR, a homolog of vertebrate liver X and vitamin D receptors, regulates nematode cholesterol balance, fatty acid desaturation, apolipoprotein production, and bile acid metabolism. Loss of nhr-8 results in a deficiency in bile acid-like steroids, called the dafachronic acids, which regulate the related DAF-12/NR, thus controlling entry into the long-lived dauer stage through cholesterol availability. Cholesterol supplementation rescues various nhr-8 phenotypes, including developmental arrest, unsaturated fatty acid deficiency, reduced fertility, and shortened life span. Notably, nhr-8 also interacts with daf-16/FOXO to regulate steady-state cholesterol levels and is synthetically lethal in combination with insulin signaling mutants that promote unregulated growth. Our studies provide important insights into nuclear receptor control of cholesterol balance and metabolism and their impact on development, reproduction, and aging in the context of larger endocrine networks.

  1. Endocytosis of hyaluronic acid by rat liver endothelial cells. Evidence for receptor recycling.

    PubMed Central

    McGary, C T; Raja, R H; Weigel, P H

    1989-01-01

    Hyaluronic acid (HA) is cleared from the blood by liver endothelial cells through receptor-mediated endocytosis [Eriksson, Fraser, Laurent, Pertoft & Smedsrod (1983) Exp. Cell Res. 144, 223-238]. We have measured the capacity of cultured rat liver endothelial cells to endocytose and degrade 125I-HA (Mr approximately 44,000) at 37 degrees C. Endocytosis was linear for 3 h and then reached a plateau. The rate of endocytosis was concentration-dependent and reached a maximum of 250 molecules/s per cell. Endocytosis of 125I-HA was inhibited more than 92% by a 150-fold excess of non-radiolabelled HA. HA, chondroitin sulphate and heparin effectively competed for endocytosis of 125I-HA, whereas glucuronic acid, N-acetylglucosamine, DNA, RNA, polygalacturonic acid and dextran did not compete. In the absence of cycloheximide, endothelial cells processed 13 times more 125I-HA in 6 h than their total (cell-surface and intracellular) specific HA-binding capacity. This result was not due to degradation and rapid replacement of receptors, because, even in the presence of cycloheximide, these cells processed 6 times more HA than their total receptor content in 6 h. Also, in the presence of cycloheximide, no decrease in 125I-HA-binding capacity was seen in cells processing or not processing HA for 6 h, indicating that receptors are not degraded after the endocytosis of HA. During endocytosis of HA at 37 degrees C, at least 65% of the intracellular HA receptors became occupied with HA within 30 min. This indicates that the intracellular HA receptors (75% of the total) function during continuous endocytosis. Hyperosmolarity inhibits endocytosis and receptor recycling in the asialoglycoprotein and low-density-lipoprotein receptor systems by disrupting the coated-pit pathway [Heuser & Anderson (1987) J. Cell Biol. 105, 230a; Oka & Weigel (1988) J. Cell. Biochem. 36, 169-183]. Hyperosmolarity inhibited 125I-HA endocytosis in liver endothelial cells by more than 90%, suggesting use of a

  2. 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. PMID:27190515

  3. 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

  4. 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

  5. Impaired generation of 12-hydroxylated bile acids links hepatic insulin signaling with dyslipidemia.

    PubMed

    Haeusler, Rebecca A; Pratt-Hyatt, Matthew; Welch, Carrie L; Klaassen, Curtis D; Accili, Domenico

    2012-01-01

    The association of type 2 diabetes with elevated plasma triglyceride (TG) and very low-density lipoproteins (VLDL), and intrahepatic lipid accumulation represents a pathophysiological enigma and an unmet therapeutic challenge. Here, we uncover a link between insulin action through FoxO1, bile acid (BA) composition, and altered lipid homeostasis that brings new insight to this longstanding conundrum. FoxO1 ablation brings about two signature lipid abnormalities of diabetes and the metabolic syndrome, elevated liver and plasma TG. These changes are associated with deficiency of 12α-hydroxylated BAs and their synthetic enzyme, Cyp8b1, that hinders the TG-lowering effects of the BA receptor, Fxr. Accordingly, pharmacological activation of Fxr with GW4064 overcomes the BA imbalance, restoring hepatic and plasma TG levels of FoxO1-deficient mice to normal levels. We propose that generation of 12α-hydroxylated products of BA metabolism represents a signaling mechanism linking hepatic lipid abnormalities with type 2 diabetes, and a treatment target for this condition.

  6. Molecular recognition of amino acids with some fluorescent ditopic pyrylium- and pyridinium-based crown ether receptors

    NASA Astrophysics Data System (ADS)

    Moghimi, A.; Maddah, B.; Yari, A.; Shamsipur, M.; Boostani, M.; Fall Rastegar, M.; Ghaderi, A. R.

    2005-10-01

    The molecular recognition of L-amino acids such as asparagine, glutamine, lysine and arginine with some crownpyryliums, CP's, and a crownpyridinium compound, as receptors, were examined in methanol. 1H NMR spectroscopy was used to examine the structural stability of the receptors in the presence of the amino acids. The fluorimetric titration of the receptors by specified amino acids, other than arginine, was followed within a few minutes and the stoichiometry and stability of the resulting amino acid complexes were evaluated. The data analysis clearly demonstrated the critical role of the terminal amino group to carboxylic acid distance of amino acids for their proper fixation on the receptor molecules. Ion pairing for the two oppositely charged carboxylate anion and pyrylium (or pyridinium) cation, as well as the hydrogen bonding between crown ethers' oxygens and ammonium hydrogens are expected as the main interaction sources in the host-guest complexations.

  7. Peripheral tackykinin and excitatory amino acid receptors mediate hyperalgesia induced by Phoneutria nigriventer venom.

    PubMed

    Zanchet, Eliane Maria; Cury, Yara

    2003-04-25

    The generation of hyperalgesia by Phoneutria nigriventer venom was investigated in rats using the paw pressure test, through the intraplantar injection of the venom. Hyperalgesia was significantly inhibited by N-[2-(4-chlorophenyl) ethyl]-1,3,4,5-tetrahydro-7,8-dihydroxy-2H-2-benzazepine-2-carbothioamide (capsazepine), a vanilloid receptor antagonist, by the local administration of pGlu-Ala-Asp-Pro-Asn-Lys-Phe-Tyr-Pro (spiro-gamma-lactam) Leu-Trp-NH(2) (GR82334) or of Phenyl-CO-Ala-Ala-D-Trp-Phe-D-Pro-Pro-Nle-NH(2) (GR94800), inhibitors of tachykinin NK(1) and NK(2) receptors, respectively, or by the local injection of dizocilpine (MK 801), (+/-)-2-amino-5-phosphonopentanoic acid ((+/-)-AP-5), or 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), antagonists of NMDA and non-NMDA excitatory amino acid receptors. The correlation between hyperalgesia and the inflammatory response induced by the venom was also investigated. The venom-induced edematogenic response was not modified by the pharmacological treatments. These results suggest that hyperalgesia induced by P. nigriventer venom is mediated by stimulation of capsaicin-sensitive neurons, with activation of peripheral tachykinin NK(1) and NK(2) receptors and of both the NMDA and AMPA receptors. Distinct mechanisms are involved in the development of hyperalgesia and edema induced by the venom.

  8. 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.

  9. G-CSF receptor-binding cyclic peptides designed with artificial amino-acid linkers

    SciTech Connect

    Shibata, Kenji . E-mail: kshibata@kyowa.co.jp; Maruyama-Takahashi, Kumiko; Yamasaki, Motoo; Hirayama, Noriaki . E-mail: hirayama@is.icc.u-tokai.ac.jp

    2006-03-10

    Designing small molecules that mimic the receptor-binding local surface structure of large proteins such as cytokines or growth factors is fascinating and challenging. In this study, we designed cyclic peptides that reproduce the receptor-binding loop structures of G-CSF. We found it is important to select a suitable linker to join two or more discontinuous sequences and both termini of the peptide corresponding to the receptor-binding loop. Structural simulations based on the crystallographic structure of KW-2228, a stable and potent analog of human G-CSF, led us to choose 4-aminobenzoic acid (Abz) as a part of the linker. A combination of 4-Abz with {beta}-alanine or glycine, and disulfide bridges between cysteins or homocysteins, gave a structure suitable for receptor binding. In this structure, the side-chains of several amino acids important for the interactions with the receptor are protruding from one side of the peptide ring. This artificial peptide showed G-CSF antagonistic activity in a cell proliferation assay.

  10. 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.

  11. Structure-dependent effects of pyridine derivatives on mechanisms of intestinal fatty acid uptake: regulation of nicotinic acid receptor and fatty acid transporter expression.

    PubMed

    Riedel, Annett; Lang, Roman; Rohm, Barbara; Rubach, Malte; Hofmann, Thomas; Somoza, Veronika

    2014-07-01

    Pyridines are widely distributed in foods. Nicotinic acid (NA), a carboxylated pyridine derivative, inhibits lipolysis in adipocytes by activation of the orphan NA receptor (HM74A) and is applied to treat hyperlipidemia. However, knowledge on the impact of pyridine derivatives on intestinal lipid metabolism is scarce. This study was performed to identify the structural determinants of pyridines for their effects on fatty acid uptake in enterocyte-like Caco-2 cells and to elucidate the mechanisms of action. The impact of 17 pyridine derivatives on fatty acid uptake was tested. Multiple regression analysis revealed the presence of a methyl group to be the structural determinant at 0.1 mM, whereas at 1 mM, the presence of a carboxylic group and the N-methylation presented further structural characteristics to affect the fatty acid uptake. NA, showing a stimulating effect on FA uptake, and N-methyl-4-phenylpyridinium (MPP), inhibiting FA uptake, were selected for mechanistic studies. Gene expression of the fatty acid transporters CD36, FATP2 and FATP4, and the lipid metabolism regulating transcription factors peroxisome proliferator-activated receptor (PPAR) α and PPARγ was up-regulated upon NA treatment. Caco-2 cells were demonstrated to express the low-affinity NA receptor HM74 of which the gene expression was up-regulated upon NA treatment. We hypothesize that the NA-induced fatty acid uptake might result from NA receptor activation and related intracellular signaling cascades. In contrast, MPP increased transepithelial electrical resistance. We therefore conclude that NA and MPP, both sharing the pyridine motif core, exhibit their contrary effects on intestinal FA uptake by activation of different mechanisms.

  12. Structure-dependent effects of pyridine derivatives on mechanisms of intestinal fatty acid uptake: regulation of nicotinic acid receptor and fatty acid transporter expression.

    PubMed

    Riedel, Annett; Lang, Roman; Rohm, Barbara; Rubach, Malte; Hofmann, Thomas; Somoza, Veronika

    2014-07-01

    Pyridines are widely distributed in foods. Nicotinic acid (NA), a carboxylated pyridine derivative, inhibits lipolysis in adipocytes by activation of the orphan NA receptor (HM74A) and is applied to treat hyperlipidemia. However, knowledge on the impact of pyridine derivatives on intestinal lipid metabolism is scarce. This study was performed to identify the structural determinants of pyridines for their effects on fatty acid uptake in enterocyte-like Caco-2 cells and to elucidate the mechanisms of action. The impact of 17 pyridine derivatives on fatty acid uptake was tested. Multiple regression analysis revealed the presence of a methyl group to be the structural determinant at 0.1 mM, whereas at 1 mM, the presence of a carboxylic group and the N-methylation presented further structural characteristics to affect the fatty acid uptake. NA, showing a stimulating effect on FA uptake, and N-methyl-4-phenylpyridinium (MPP), inhibiting FA uptake, were selected for mechanistic studies. Gene expression of the fatty acid transporters CD36, FATP2 and FATP4, and the lipid metabolism regulating transcription factors peroxisome proliferator-activated receptor (PPAR) α and PPARγ was up-regulated upon NA treatment. Caco-2 cells were demonstrated to express the low-affinity NA receptor HM74 of which the gene expression was up-regulated upon NA treatment. We hypothesize that the NA-induced fatty acid uptake might result from NA receptor activation and related intracellular signaling cascades. In contrast, MPP increased transepithelial electrical resistance. We therefore conclude that NA and MPP, both sharing the pyridine motif core, exhibit their contrary effects on intestinal FA uptake by activation of different mechanisms. PMID:24767308

  13. RNA-Binding Protein FXR1 Regulates p21 and TERC RNA to Bypass p53-Mediated Cellular Senescence in OSCC.

    PubMed

    Majumder, Mrinmoyee; House, Reniqua; Palanisamy, Nallasivam; Qie, Shuo; Day, Terrence A; Neskey, David; Diehl, J Alan; Palanisamy, Viswanathan

    2016-09-01

    RNA-binding proteins (RBP) regulate numerous aspects of co- and post-transcriptional gene expression in cancer cells. Here, we demonstrate that RBP, fragile X-related protein 1 (FXR1), plays an essential role in cellular senescence by utilizing mRNA turnover pathway. We report that overexpressed FXR1 in head and neck squamous cell carcinoma targets (G-quadruplex (G4) RNA structure within) both mRNA encoding p21 (Cyclin-Dependent Kinase Inhibitor 1A (CDKN1A, Cip1) and the non-coding RNA Telomerase RNA Component (TERC), and regulates their turnover to avoid senescence. Silencing of FXR1 in cancer cells triggers the activation of Cyclin-Dependent Kinase Inhibitors, p53, increases DNA damage, and ultimately, cellular senescence. Overexpressed FXR1 binds and destabilizes p21 mRNA, subsequently reduces p21 protein expression in oral cancer cells. In addition, FXR1 also binds and stabilizes TERC RNA and suppresses the cellular senescence possibly through telomerase activity. Finally, we report that FXR1-regulated senescence is irreversible and FXR1-depleted cells fail to form colonies to re-enter cellular proliferation. Collectively, FXR1 displays a novel mechanism of controlling the expression of p21 through p53-dependent manner to bypass cellular senescence in oral cancer cells. PMID:27606879

  14. RNA-Binding Protein FXR1 Regulates p21 and TERC RNA to Bypass p53-Mediated Cellular Senescence in OSCC

    PubMed Central

    Majumder, Mrinmoyee; House, Reniqua; Palanisamy, Nallasivam; Qie, Shuo; Day, Terrence A.; Neskey, David; Diehl, J. Alan

    2016-01-01

    RNA-binding proteins (RBP) regulate numerous aspects of co- and post-transcriptional gene expression in cancer cells. Here, we demonstrate that RBP, fragile X-related protein 1 (FXR1), plays an essential role in cellular senescence by utilizing mRNA turnover pathway. We report that overexpressed FXR1 in head and neck squamous cell carcinoma targets (G-quadruplex (G4) RNA structure within) both mRNA encoding p21 (Cyclin-Dependent Kinase Inhibitor 1A (CDKN1A, Cip1) and the non-coding RNA Telomerase RNA Component (TERC), and regulates their turnover to avoid senescence. Silencing of FXR1 in cancer cells triggers the activation of Cyclin-Dependent Kinase Inhibitors, p53, increases DNA damage, and ultimately, cellular senescence. Overexpressed FXR1 binds and destabilizes p21 mRNA, subsequently reduces p21 protein expression in oral cancer cells. In addition, FXR1 also binds and stabilizes TERC RNA and suppresses the cellular senescence possibly through telomerase activity. Finally, we report that FXR1-regulated senescence is irreversible and FXR1-depleted cells fail to form colonies to re-enter cellular proliferation. Collectively, FXR1 displays a novel mechanism of controlling the expression of p21 through p53-dependent manner to bypass cellular senescence in oral cancer cells. PMID:27606879

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

    PubMed

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

    2003-09-01

    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.

  16. G-Protein-Coupled Lysophosphatidic Acid Receptors and Their Regulation of AKT Signaling

    PubMed Central

    Riaz, Anjum; Huang, Ying; Johansson, Staffan

    2016-01-01

    A hallmark of G-protein-coupled receptors (GPCRs) is their ability to recognize and respond to chemically diverse ligands. Lysophospholipids constitute a relatively recent addition to these ligands and carry out their biological functions by activating G-proteins coupled to a large family of cell-surface receptors. This review aims to highlight salient features of cell signaling by one class of these receptors, known as lysophosphatidic acid (LPA) receptors, in the context of phosphatidylinositol 3-kinase (PI3K)–AKT pathway activation. LPA moieties efficiently activate AKT phosphorylation and activation in a multitude of cell types. The interplay between LPA, its receptors, the associated Gαi/o subunits, PI3K and AKT contributes to the regulation of cell survival, migration, proliferation and confers chemotherapy-resistance in certain cancers. However, detailed information on the regulation of PI3K–AKT signals induced by LPA receptors is missing from the literature. Here, some urgent issues for investigation are highlighted. PMID:26861299

  17. G-Protein-Coupled Lysophosphatidic Acid Receptors and Their Regulation of AKT Signaling.

    PubMed

    Riaz, Anjum; Huang, Ying; Johansson, Staffan

    2016-01-01

    A hallmark of G-protein-coupled receptors (GPCRs) is their ability to recognize and respond to chemically diverse ligands. Lysophospholipids constitute a relatively recent addition to these ligands and carry out their biological functions by activating G-proteins coupled to a large family of cell-surface receptors. This review aims to highlight salient features of cell signaling by one class of these receptors, known as lysophosphatidic acid (LPA) receptors, in the context of phosphatidylinositol 3-kinase (PI3K)-AKT pathway activation. LPA moieties efficiently activate AKT phosphorylation and activation in a multitude of cell types. The interplay between LPA, its receptors, the associated Gαi/o subunits, PI3K and AKT contributes to the regulation of cell survival, migration, proliferation and confers chemotherapy-resistance in certain cancers. However, detailed information on the regulation of PI3K-AKT signals induced by LPA receptors is missing from the literature. Here, some urgent issues for investigation are highlighted. PMID:26861299

  18. 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.

  19. Two isoforms of Xenopus retinoic acid receptor gamma 2 (B) exhibit differential expression and sensitivity to retinoic acid during embryogenesis.

    PubMed

    Crawford, M J; Liversage, R A; Varmuza, S L

    1995-01-01

    We report the isolation of two retinoic acid receptor isoforms (RAR gamma), which differ only in the 5'untranslated and putative N-terminus A regions. The two isoforms appear to serve as early markers for the presumptive neural axis; however, their expression patterns differ. RAR-gamma 2.1 is first expressed at gastrulation at the dorsal lip and subsequently along the presumptive neural axis. RAR- gamma 2.2 represents the full-length sequence of a receptor cDNA already partially characterized and present as a maternal transcript [Ellinger-Ziegelbauer and Dreyer (1991); Genes Dev 5:94-104, (1993): Mech Dev 41:31-46; Pfeffer and DeRobertis, (1994) Mech Dev: 45:147-153]. Unlike RAR-gamma 2.2, the 2.1 variant is not expressed either in pre-somitic mesoderm or notochord. RAR-gamma 2.1 is strongly expressed in branchial arches and to a lesser extent in the neural floor plate. The two isoforms also exhibit differential sensitivity to retinoic acid. Constitutive expression of RAR gamma 2.2 following neurulation appears to be depressed by treatment with retinoic acid, but domains of highest expression, namely, the head and tail, remain relatively unaffected, as do patterns of expression prior to late neurulation. By contrast, RAR-gamma 2.1 is not transcribed in retinoid-inhibited structures. Using microinjection techniques, we show that changes of RAR-gamma 2.1 expression in presumptive head structures occur as an early and local consequence of retinoic acid administration. Since RAR-gamma 2.1 expression is inhibited by retinoic acid, we tested to see if other treatments that perturb axis formation had any effect. Surprisingly, UV irradiation did not suppress that its inhibition by retinoic acid is not due solely to inhibition of anterior neural development. These experiments demonstrate a new subdivision of isoforms that undergo differential expression during development and that exhibit differential sensitivity to retinoic acid and to UV. This sensitivity and the presence

  20. Rapid attenuation of receptor-induced diacylglycerol and phosphatidic acid by phospholipase D-mediated transphosphatidylation: formation of bisphosphatidic acid.

    PubMed Central

    van Blitterswijk, W J; Hilkmann, H

    1993-01-01

    Generation and attenuation of lipid second messengers are key processes in cellular signalling. Receptor-mediated increase in 1,2-diacylglycerol (DG) levels is attenuated by DG kinase and DG lipase. We here report a novel mechanism of DG attenuation by phospholipase D (PLD), which also precludes the production of another (putative) second messenger, phosphatidic acid (PA). In the presence of an alcohol, PLD converts phosphatidylcholine (PC) into a phosphatidylalcohol (by transphosphatidylation) rather than into PA. We found in bradykinin-stimulated human fibroblasts that PLD mediates transphosphatidylation from PC (donor) to the endogenous 'alcohol' DG (acceptor), yielding bis(1,2-diacylglycero)-3-sn-phosphate (bisphosphatidic acid; bisPA). This uncommon phospholipid is thus a condensation product of the phospholipase C (PLC) and PLD signalling pathways, where PLC produces DG and PLD couples this DG to a phosphatidyl moiety. Long-term phorbol ester treatment blocks bradykinin-induced activation of PLD and consequent bisPA formation, thereby unveiling rapid formation of DG. BisPA formation is rapid (15 s) and transient (peaks at 2-10 min) and is also induced by other stimuli capable of raising DG and activating PLD simultaneously, e.g. endothelin, lysophosphatidic acid, fetal calf serum, phorbol ester, dioctanoylglycerol or bacterial PLC. This novel metabolic route counteracts rapid accumulation of receptor-induced DG and PA, and assigns for the first time a physiological role to the transphosphatidylation activity of PLD, that is signal attenuation. Images PMID:8392931

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

    PubMed Central

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

    2015-01-01

    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. PMID:25839160

  2. 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.

  3. Unsurmountable antagonism of brain 5-hydroxytryptamine2 receptors by (+)-lysergic acid diethylamide and bromo-lysergic acid diethylamide.

    PubMed

    Burris, K D; Sanders-Bush, E

    1992-11-01

    Lysergic acid diethylamide (LSD) and its structural analogue 2-bromo-lysergic acid diethylamide (BOL) act as unsurmountable antagonists of serotonin-elicited contractions in smooth muscle preparations. Two different models, allosteric and kinetic, have been invoked to explain these findings. The present studies investigate the mechanism of antagonism of brain 5-hydroxytryptamine (5HT)2 receptors, utilizing cells transfected with 5HT2 receptor cDNA cloned from rat brain. A proximal cellular response, phosphoinositide hydrolysis, was examined in order to minimize possible postreceptor effects. Even though LSD behaved as a partial agonist and BOL as a pure antagonist, both drugs blocked the effect of serotonin in an unsurmountable manner, i.e., increasing concentrations of serotonin could not overcome the blocking effect of LSD or BOL. Radioligand binding studies showed that preincubation of membranes with either LSD or BOL reduced the density of [3H]ketanserin binding sites, suggesting that the drugs bind tightly to the 5HT2 receptor and are not displaced during the binding assay. Two additional experiments supported this hypothesis. First, the off-rate of [3H] LSD was slow (20 min), relative to that of [3H]ketanserin (approximately 4 min). Second, when the length of incubation with [3H]ketanserin was increased to 60 min, the LSD-induced decrease in Bmax was essentially eliminated. The possibility that LSD and BOL decrease [3H]ketanserin binding by interacting with an allosteric site was rejected, because neither drug altered the rate of dissociation of [3H]ketanserin. The most parsimonious interpretation of these results is that unsurmountable antagonism reflects prolonged occupancy of the receptor by slowly reversible antagonists.

  4. Upregulation of UGT2B4 Expression by 3′-Phosphoadenosine-5′-Phosphosulfate Synthase Knockdown: Implications for Coordinated Control of Bile Acid Conjugation

    PubMed Central

    Barrett, Kathleen G.; Fang, Hailin; Cukovic, Daniela; Dombkowski, Alan A.; Kocarek, Thomas A.

    2015-01-01

    During cholestasis, the bile acid–conjugating enzymes, SULT2A1 and UGT2B4, work in concert to prevent the accumulation of toxic bile acids. To understand the impact of sulfotransferase deficiency on human hepatic gene expression, we knocked down 3′-phosphoadenosine-5′-phosphosulfate synthases (PAPSS) 1 and 2, which catalyze synthesis of the obligate sulfotransferase cofactor, in HepG2 cells. PAPSS knockdown caused no change in SULT2A1 expression; however, UGT2B4 expression increased markedly (∼41-fold increase in UGT2B4 mRNA content). Knockdown of SULT2A1 in HepG2 cells also increased UGT2B4 expression. To investigate the underlying mechanism, we transfected PAPSS-deficient HepG2 cells with a luciferase reporter plasmid containing ∼2 Kb of the UGT2B4 5′-flanking region, which included a response element for the bile acid–sensing nuclear receptor, farnesoid X receptor (FXR). FXR activation or overexpression increased UGT2B4 promoter activity; however, knocking down FXR or mutating or deleting the FXR response element did not significantly decrease UGT2B4 promoter activity. Further evaluation of the UGT2B4 5′-flanking region indicated the presence of distal regulatory elements between nucleotides −10090 and −10037 that negatively and positively regulated UGT2B4 transcription. Pulse-chase analysis showed that increased UGT2B4 expression in PAPSS-deficient cells was attributable to both increased mRNA synthesis and stability. Transfection analysis demonstrated that the UGT2B4 3′-untranslated region decreased luciferase reporter expression less in PAPSS-deficient cells than in control cells. These data indicate that knocking down PAPSS increases UGT2B4 transcription and mRNA stability as a compensatory response to the loss of SULT2A1 activity, presumably to maintain bile acid–conjugating activity. PMID:25948711

  5. LE135, a retinoid acid receptor antagonist, produces pain through direct activation of TRP channels

    PubMed Central

    Yin, Shijin; Luo, Jialie; Qian, Aihua; Yu, Weihua; Hu, Hongzhen

    2014-01-01

    Background and PurposeRetinoids, through their activation of retinoic acid receptors (RARs) and retinoid X receptors, regulate diverse cellular processes, and pharmacological intervention in their actions has been successful in the treatment of skin disorders and cancers. Despite the many beneficial effects, administration of retinoids causes irritating side effects with unknown mechanisms. Here, we demonstrate that LE135 [4-(7,8,9,10-tetrahydro-5,7,7,10,10-pentamethyl-5H-benzo[e]naphtho[2,3-b][1,4]diazepin-13-yl)benzoic acid], a selective antagonist of RARβ, is a potent activator of the capsaicin (TRPV1) and wasabi (TRPA1) receptors, two critical pain-initiating cation channels. Experimental ApproachWe performed to investigate the excitatory effects of LE135 on TRPV1 and TRPA1 channels expressed in HEK293T cells and in dorsal root ganglia neurons with calcium imaging and patch-clamp recordings. We also used site-directed mutagenesis of the channels to determine the structural basis of LE135-induced activation of TRPV1 and TRPA1 channels and behavioural testing to examine if pharmacological inhibition and genetic deletion of the channels affected LE135-evoked pain-related behaviours. Key ResultsLE135 activated both the capsaicin receptor (TRPV1) and the allyl isothiocyanate receptor (TRPA1) heterologously expressed in HEK293T cells and endogenously expressed by sensory nociceptors. Mutations disrupting the capsaicin-binding site attenuated LE135 activation of TRPV1 channels and a single mutation (K170R) eliminated TRPA1 activity evoked by LE135. Intraplantar injection of LE135 evoked pain-related behaviours. Both TRPV1 and TRPA1 channels were involved in LE135-elicited pain-related responses, as shown by pharmacological and genetic ablation studies. Conclusions and ImplicationsThis blocker of retinoid acid signalling also exerted non-genomic effects through activating the pain-initiating TRPV1 and TRPA1 channels. PMID:24308840

  6. The fate of P2Y-related orphan receptors: GPR80/99 and GPR91 are receptors of dicarboxylic acids.

    PubMed

    Gonzalez, Nathalie Suarez; Communi, Didier; Hannedouche, Sébastien; Boeynaems, Jean-Marie

    2004-12-01

    Several orphan G protein-coupled receptors are structurally close to the family of P2Y nucleotide receptors: GPR80/99 and GPR91 are close to P2Y(1/2/4/6/11) receptors, whereas GPR87, H963 and GPR34 are close to P2Y(12/13/14). Over the years, several laboratories have attempted without success to identify the ligands of those receptors. In early 2004, two papers have been published: One claiming that GPR80/99 is an AMP receptor, called P2Y(15), and the other one showing that GPR80/99 is a receptor for alpha-ketoglutarate, while GPR91 is a succinate receptor. The accompanying paper by Qi et al. entirely supports that GPR80/99 is an alpha-ketoglutarate receptor and not an AMP receptor. The closeness of dicarboxylic acid and P2Y nucleotide receptors might be linked to the negative charges of both types of ligands and the involvement of conserved Arg residues in their neutralization. PMID:18404396

  7. Retinoic acid receptor alpha mediates growth inhibition by retinoids in human colon carcinoma HT29 cells.

    PubMed

    Nicke, B; Kaiser, A; Wiedenmann, B; Riecken, E O; Rosewicz, S

    1999-08-11

    Although retinoids have been suggested to inhibit chemically induced colon carcinogenesis, the molecular mechanisms underlying retinoid-mediated growth regulation in colon carcinoma cells are unknown. Therefore, we investigated the biological effects of retinoids on growth in HT29 colon carcinoma cells. All-trans retinoic acid (ATRA) treatment of HT29 cells resulted in a profound inhibition of anchorage-independent growth without biochemical or morphological evidence for induction of differentiation. Treatment with the selective RARalpha agonist Ro 40-6055 completely mimicked the effects of ATRA on growth and transactivation of a betaRAREx2-luciferase reporter construct, while RARbeta- and gamma-specific analogues were ineffective. Furthermore, ATRA-regulated growth and transactivation could be completely blocked by a RARalpha-selective receptor antagonist. Thus, ATRA potently inhibits anchorage-independent growth in HT29 cells and this effect is mainly if not exclusively mediated by the retinoic acid receptor alpha.

  8. New World and Old World Alphaviruses Have Evolved to Exploit Different Components of Stress Granules, FXR and G3BP Proteins, for Assembly of Viral Replication Complexes

    PubMed Central

    Kim, Dal Young; Reynaud, Josephine M.; Rasalouskaya, Aliaksandra; Akhrymuk, Ivan; Mobley, James A.; Frolov, Ilya; Frolova, Elena I.

    2016-01-01

    The positive-strand RNA viruses initiate their amplification in the cell from a single genome delivered by virion. This single RNA molecule needs to become involved in replication process before it is recognized and degraded by cellular machinery. In this study, we show that distantly related New World and Old World alphaviruses have independently evolved to utilize different cellular stress granule-related proteins for assembly of complexes, which recruit viral genomic RNA and facilitate formation of viral replication complexes (vRCs). Venezuelan equine encephalitis virus (VEEV) utilizes all members of the Fragile X syndrome (FXR) family, while chikungunya and Sindbis viruses exploit both members of the G3BP family. Despite being in different families, these proteins share common characteristics, which determine their role in alphavirus replication, namely, the abilities for RNA-binding and for self-assembly into large structures. Both FXR and G3BP proteins interact with virus-specific, repeating amino acid sequences located in the C-termini of hypervariable, intrinsically disordered domains (HVDs) of viral nonstructural protein nsP3. We demonstrate that these host factors orchestrate assembly of vRCs and play key roles in RNA and virus replication. Only knockout of all of the homologs results in either pronounced or complete inhibition of replication of different alphaviruses. The use of multiple homologous proteins with redundant functions mediates highly efficient recruitment of viral RNA into the replication process. This independently evolved acquisition of different families of cellular proteins by the disordered protein fragment to support alphavirus replication suggests that other RNA viruses may utilize a similar mechanism of host factor recruitment for vRC assembly. The use of different host factors by alphavirus species may be one of the important determinants of their pathogenesis. PMID:27509095

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

    PubMed

    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 (E max = 26 ± 2.0%) on 5-HT1A receptor. In conclusion, our findings showed antidepressant-like effect of D1 and possible involvement of 5-HT1A receptor.

  10. Specific interaction of aurintricarboxylic acid with the human immunodeficiency virus/CD4 cell receptor

    SciTech Connect

    Schols, D.; Baba, M.; Pauwels, R.; Desmyter, J.; De Clercq, E. )

    1989-05-01

    The triphenylmethane derivative aurintricarboxylic acid (ATA), but not aurin, selectively prevented the binding of OKT4A/Leu-3a monoclonal antibody (mAb) and, to a lesser extent, OKT4 mAb to the CD4 cell receptor for human immunodeficiency virus type 1 (HIV-1). The effect was seen within 1 min at an ATA concentration of 10 {mu}M in various T4{sup +} cells (MT-4, U-937, peripheral blood lymphocytes, and monocytes). It was dose-dependent and reversible. ATA prevented the attachment of radiolabeled HIV-1 particles to MT-4 cells, which could be expected as the result of its specific binding to the HIV/CD4 receptor. Other HIV inhibitors such as suramin, fuchsin acid, azidothymidine, dextran sulfate, heparin, and pentosan polysulfate did not affect OKT4A/Leu-3a mAb binding to the CD4 receptor, although the sulfated polysaccharides suppressed HIV-1 adsorption to the cells at concentrations required for complete protection against HIV-1 cytopathogenicity. Thus, ATA is a selective marker molecule for the CD4 receptor. ATA also interfered with the staining of membrane-associated HIV-1 glycoprotein gp120 by a mAb against it. These unusual properties of a small molecule of nonimmunological origin may have important implications for the study of CD4/HIV/AIDS pathogenesis and possibly treatment.

  11. The myeloperoxidase product hypochlorous acid generates irreversible high-density lipoprotein receptor inhibitors

    PubMed Central

    Binder, Veronika; Ljubojevic, Senka; Haybaeck, Johannes; Holzer, Michael; El-Gamal, Dalia; Schicho, Rudolf; Pieske, Burkert; Heinemann, Akos; Marsche, Gunther

    2014-01-01

    Objective Elevated levels of advanced oxidation protein products (AOPPs) have been described in several chronic inflammatory diseases, like chronic renal insufficiency, rheumatoid arthritis and atherosclerosis. Recent findings revealed that AOPPs are inhibitors of the major high-density lipoprotein (HDL) receptor, scavenger receptor class B, type 1 (SR-BI). Here we investigated what oxidation induced structural alterations convert plasma albumin into an HDL-receptor inhibitor. Approach and Results Exposure of albumin to the physiological oxidant, hypochlorous acid, generated high affinity SR-BI ligands. Protection of albumin lysine-residues prior exposure to hypochlorous acid as well as regeneration of N-chloramines after oxidation of albumin completely prevented binding of oxidized albumin to SR-BI, indicating that modification of albumin lysine-residues is required to generate SR-BI ligands. Of particular interest, N-chloramines within oxidized albumin promoted irreversible binding to SR-BI, resulting in permanent receptor blockade. We observed that the SR-BI inhibitory activity of albumin isolated from chronic kidney disease patients correlated with the content of the myeloperoxidase-specific oxidation product 3-chlorotyrosine and was associated with alterations in the composition of HDL. Conclusion Given that several potential atheroprotective activities of HDL are mediated by SR-BI, the present results raise the possibility that oxidized plasma albumin, through permanent SR-BI blockade, contributes to the pathophysiology of cardiovascular disease. PMID:23493288

  12. The acidic domains of the Toc159 chloroplast preprotein receptor family are intrinsically disordered protein domains

    PubMed Central

    2009-01-01

    Background The Toc159 family of proteins serve as receptors for chloroplast-destined preproteins. They directly bind to transit peptides, and exhibit preprotein substrate selectivity conferred by an unknown mechanism. The Toc159 receptors each include three domains: C-terminal membrane, central GTPase, and N-terminal acidic (A-) domains. Although the function(s) of the A-domain remains largely unknown, the amino acid sequences are most variable within these domains, suggesting they may contribute to the functional specificity of the receptors. Results The physicochemical properties of the A-domains are characteristic of intrinsically disordered proteins (IDPs). Using CD spectroscopy we show that the A-domains of two Arabidopsis Toc159 family members (atToc132 and atToc159) are disordered at physiological pH and temperature and undergo conformational changes at temperature and pH extremes that are characteristic of IDPs. Conclusions Identification of the A-domains as IDPs will be important for determining their precise function(s), and suggests a role in protein-protein interactions, which may explain how these proteins serve as receptors for such a wide variety of preprotein substrates. PMID:20042108

  13. 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

  14. Retinoic acid and retinoid receptors: potential chemopreventive and therapeutic role in cervical cancer.

    PubMed

    Abu, Jafaru; Batuwangala, Madu; Herbert, Karl; Symonds, Paul

    2005-09-01

    Retinoids are natural and synthetic derivatives of vitamin A, which can be obtained from animal products (milk, liver, beef, fish oils, and eggs) and vegetables (carrots, mangos, sweet potatoes, and spinach). Retinoids regulate various important cellular functions in the body through specific nuclear retinoic-acid receptors and retinoid-X receptors, which are encoded by separate genes. Retinoic-acid receptors specifically bind tretinoin and alitretinoin, whereas retinoid-X receptors bind only alitretinoin. Retinoids have long been established as crucial for several essential life processes-healthy growth, vision, maintenance of tissues, reproduction, metabolism, tissue differentiation (normal, premalignant cells, and malignant cells), haemopoiesis, bone development, spermatogenesis, embryogenesis, and overall survival. Therefore, deficiency of vitamin A can lead to various unwanted biological effects. Several experimental and epidemiological studies have shown the antiproliferative activity of retinoids and their potential use in cancer treatment and chemoprevention. Emerging clinical trials have shown the chemotherapeutic and chemopreventive potential of retinoids in cancerous and precancerous conditions of the uterine cervix. In this review, we explore the potential chemopreventive and therapeutic roles of retinoids in preinvasive and invasive cervical neoplasia.

  15. Synthesis and evaluation of a (68)Ga labeled folic acid derivative for targeting folate receptors.

    PubMed

    Jain, Akanksha; Mathur, Anupam; Pandey, Usha; Bhatt, Jyotsna; Mukherjee, Archana; Ram, Ramu; Sarma, Haladhar Dev; Dash, Ashutosh

    2016-10-01

    Present work evaluates the potential of a newly synthesized (68)Ga-NOTA-folic acid conjugate for PET imaging of tumors over-expressing folate receptors (FRs). NOTA-folic acid conjugate was synthesized and characterized. It was radiolabeled with (68)Ga in ≥ 95% radiolabeling yields. In vitro cell binding studies showed a maximum cell uptake of 1.7±0.4% per million KB cells which was completely blocked on addition of cold folic acid showing specificity towards the FRs. However, further studies in tumor xenografts are warranted in order to assess the potential of (68)Ga-folic acid complex for imaging tumors over-expressing FRs. PMID:27501138

  16. 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. PMID:26503258

  17. Autoradiographic localization and characterization of [125I]lysergic acid diethylamide binding to serotonin receptors in Aplysia.

    PubMed

    Kadan, M J; Hartig, P R

    1988-03-01

    The sensitive serotonergic radioligand 2-[125I]lysergic acid diethylamide was used to study the distribution and pharmacological binding properties of serotonin receptors in Aplysia californica. The high specific activity of this radioligand allowed us to develop a methodology for the investigation of receptor binding properties and receptor distribution in a single ganglion. [125I]Lysergic acid diethylamide labels a population of high-affinity serotonergic sites (Kd = 0.41 nM) in Aplysia ganglia whose regional distribution matches that expected from previous electrophysiological and immunohistochemical studies. The properties of [125I]lysergic acid diethylamide binding sites in Aplysia are in general agreement with previous studies on [3H]lysergic acid diethylamide binding in this system but these sites differ from the serotonergic receptor subtypes described in the mammalian brain. Guanine nucleotides were shown to modulate agonist but not antagonist affinity for the [125I]lysergic acid diethylamide binding site in Aplysia, suggesting that this site is coupled to a G-protein. Images of serotonin receptor distribution in the Aplysia nervous system were obtained from autoradiograms of [125I]lysergic acid diethylamide binding. Serotonin receptors in ganglia tissue sections are located primarily within the neuropil. In addition, a subset of neuronal soma are specifically labeled by [125I]lysergic acid diethylamide. These studies indicate that [125I]lysergic acid diethylamide binds to sites in the Aplysia nervous system which display a regional distribution, pharmacological binding properties and evidence of coupling to a G-protein consistent with labeling of a subset of functional serotonin receptors. In addition, the techniques used in this investigation provide a general approach for rapidly characterizing the pharmacological properties and anatomical distribution of receptor binding sites in single invertebrate ganglia. Individual neurons containing these receptor

  18. Inhibition of Leydig tumor growth by farnesoid X receptor activation: the in vitro and in vivo basis for a novel therapeutic strategy.

    PubMed

    Catalano, Stefania; Panza, Salvatore; Malivindi, Rocco; Giordano, Cinzia; Barone, Ines; Bossi, Gianluca; Lanzino, Marilena; Sirianni, Rosa; Mauro, Loredana; Sisci, Diego; Bonofiglio, Daniela; Andò, Sebastiano

    2013-05-15

    Leydig cell tumors (LCTs) are the most common tumors of the gonadal stroma and represent about 3% of all testicular neoplasms. In most cases, LCTs are benign; however, if the tumor is malignant, no effective treatments are currently available. We have recently reported that farnesoid X receptor (FXR) is expressed in R2C Leydig tumor cells, and it reduces the estrogen-dependent cell proliferation by negatively regulating aromatase expression. Here, we demonstrated that treatment with GW4064, a specific FXR agonist, markedly reduced Leydig tumor growth in vivo by inhibiting proliferation and inducing apoptosis. Indeed, the tumors from GW4064-treated mice exhibited a decrease in the expression of the proliferation marker Ki-67 and aromatase along with an increase in the apoptotic nuclei. FXR activation induced an enhanced poly(ADP-ribose) polymerase cleavage, a marked DNA fragmentation and a strong increase in TUNEL-positive R2C cells also in vitro. Moreover, in both in vivo and in vitro models, FXR ligands upregulated mRNA and protein levels of p53 and of its downstream effector p21(WAF1/Cip1) . Functional experiments showed that FXR ligands upregulated p53 promoter activity and this occurred through an increased binding of FXR/nuclear factor-kB (NF-kB) complex to the NF-kB site located within p53 promoter region as revealed by electrophoretic mobility shift assay and chromatin immunoprecipitation analysis. Taken together, results from our study show, for the first time, that treatment with FXR ligands induces Leydig tumor regression in vivo, suggesting that activation of FXR may represent a promising therapeutic strategy for LCTs.

  19. 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. PMID:25652827

  20. Abscisic Acid Analogues That Act as Universal or Selective Antagonists of Phytohormone Receptors.

    PubMed

    Rajagopalan, Nandhakishore; Nelson, Ken M; Douglas, Amy F; Jheengut, Vishal; Alarcon, Idralyn Q; McKenna, Sean A; Surpin, Marci; Loewen, Michele C; Abrams, Suzanne R

    2016-09-13

    The plant hormone abscisic acid (ABA) plays many important roles in controlling plant development and physiology, from flowering to senescence. ABA is now known to exert its effects through a family of soluble ABA receptors, which in Arabidopsis thaliana has 13 members divided into three clades. Homologues of these receptors are present in other plants, also in relatively large numbers. Investigation of the roles of each homologue in mediating the diverse physiological roles of ABA is hampered by this genetic redundancy. We report herein the in vitro screening of a targeted ABA-like analogue library and identification of novel antagonist hits, including the analogue PBI686 that had been developed previously as a probe for identifying ABA-binding proteins. Further in vitro characterization of PBI686 and development of second-generation leads yielded both receptor-selective and universal antagonist hits. In planta assays in different species have demonstrated that these antagonist leads can overcome various ABA-induced physiological changes. While the general antagonists open up a hitherto unexplored avenue for controlling plant growth through inhibition of ABA-regulated physiological processes, the receptor-selective antagonist can be developed into chemical probes to explore the physiological roles of individual receptors.

  1. Nuclear receptors for retinoic acid and thyroid hormone regulate transcription of keratin genes.

    PubMed Central

    Tomic, M; Jiang, C K; Epstein, H S; Freedberg, I M; Samuels, H H; Blumenberg, M

    1990-01-01

    In the epidermis, retinoids regulate the expression of keratins, the intermediate filament proteins of epithelial cells. We have cloned the 5' regulatory regions of four human epidermal keratin genes, K#5, K#6, K#10, and K#14, and engineered constructs in which these regions drive the expression of the CAT reporter gene. By co-transfecting the constructs into epithelial cells along with the vectors expressing nuclear receptors for retinoic acid (RA) and thyroid hormone, we have demonstrated that the receptors can suppress the promoters of keratin genes. The suppression is ligand dependent; it is evident both in established cell lines and in primary cultures of epithelial cells. The three RA receptors have similar effects on keratin gene transcription. Our data indicate that the nuclear receptors for RA and thyroid hormone regulate keratin synthesis by binding to negative recognition elements in the upstream DNA sequences of the keratin genes. RA thus has a twofold effect on epidermal keratin expression: qualitatively, it regulates the regulators that effect the switch from basal cell-specific keratins to differentiation-specific ones; and quantitatively, it determines the level of keratin synthesis within the cell by direct interaction of its receptors with the keratin gene promoters. Images PMID:1712634

  2. Abscisic Acid Analogues That Act as Universal or Selective Antagonists of Phytohormone Receptors.

    PubMed

    Rajagopalan, Nandhakishore; Nelson, Ken M; Douglas, Amy F; Jheengut, Vishal; Alarcon, Idralyn Q; McKenna, Sean A; Surpin, Marci; Loewen, Michele C; Abrams, Suzanne R

    2016-09-13

    The plant hormone abscisic acid (ABA) plays many important roles in controlling plant development and physiology, from flowering to senescence. ABA is now known to exert its effects through a family of soluble ABA receptors, which in Arabidopsis thaliana has 13 members divided into three clades. Homologues of these receptors are present in other plants, also in relatively large numbers. Investigation of the roles of each homologue in mediating the diverse physiological roles of ABA is hampered by this genetic redundancy. We report herein the in vitro screening of a targeted ABA-like analogue library and identification of novel antagonist hits, including the analogue PBI686 that had been developed previously as a probe for identifying ABA-binding proteins. Further in vitro characterization of PBI686 and development of second-generation leads yielded both receptor-selective and universal antagonist hits. In planta assays in different species have demonstrated that these antagonist leads can overcome various ABA-induced physiological changes. While the general antagonists open up a hitherto unexplored avenue for controlling plant growth through inhibition of ABA-regulated physiological processes, the receptor-selective antagonist can be developed into chemical probes to explore the physiological roles of individual receptors. PMID:27523384

  3. 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.

  4. Aldose Reductase acts as a Selective Derepressor of PPARγ and Retinoic Acid Receptor

    PubMed Central

    Thiagarajan, Devi; Ananthakrishnan, Radha; Zhang, Jinghua; O’Shea, Karen M.; Quadri, Nosirudeen; Li, Qing; Sas, Kelli; Jing, Xiao; Rosario, Rosa; Pennathur, Subramaniam; Schmidt, Ann Marie; Ramasamy, Ravichandran

    2016-01-01

    Summary Histone deacetylase 3 (HDAC3), a chromatin modifying enzyme, requires association with the deacetylase containing domain (DAD) of the nuclear receptor co-repressors NCOR1 and SMRT for its stability and activity. Here we show that aldose reductase (AR), the rate-limiting enzyme of the polyol pathway, competes with HDAC3 to bind the NCOR1/SMRT DAD. Increased AR expression leads to HDAC3 degradation followed by increased PPARγ signaling resulting in lipid accumulation in the heart. AR also downregulates expression of nuclear corepressor complex cofactors including Gps2 and Tblr1, thus affecting activity of the nuclear corepressor complex itself. Though AR reduces HDAC3-corepressor complex formation, it specifically de-represses the retinoic acid receptor (RAR), but not other nuclear receptors such as the thyroid receptor (TR) and liver X receptor (LXR). In summary, this work defines a distinct role for AR in lipid and retinoid metabolism through HDAC3 regulation and consequent de-repression of PPARγ and RAR. PMID:27052179

  5. 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

  6. 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.

  7. Benzodiazepine receptor agonists affect both binding and gating of recombinant alpha1beta2gamma2 gamma-aminobutyric acid-A receptors.

    PubMed

    Mercik, Katarzyna; Piast, Michał; Mozrzymas, Jerzy W

    2007-05-28

    Benzodiazepines are known to act by enhancing the effect of gamma-aminobutyric acid-A receptor agonists. Positive modulation by benzodiazepines is typically ascribed to upregulation of agonist binding affinity but their effect on gamma-aminobutyric acid-A receptor gating remain unclear. In this work, we have used the ultrafast application system to examine the impact of flurazepam and zolpidem on recombinant alpha1beta2gamma2 gamma-aminobutyric acid-A receptors. As expected, both drugs strongly enhanced currents evoked by low [gamma-aminobutyric acid]. These compounds, however, also affected currents elicited by saturating agonist concentration. In particular, flurazepam and zolpidem reduced current amplitudes and slowed down the recovery process in paired-pulse experiments. Moreover, flurazepam accelerated the current rise time and zolpidem enhanced the rate and extent of desensitization. We propose that flurazepam and zolpidem modulate gamma-aminobutyric acid-A receptors by strong enhancement of agonist binding with a superimposed limited effect on the receptor gating.

  8. 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

  9. Design, synthesis and biological activity of phenoxyacetic acid derivatives as novel free fatty acid receptor 1 agonists.

    PubMed

    Li, Zheng; Wang, Xuekun; Xu, Xue; Yang, Jianyong; Xia, Wenting; Zhou, Xianhao; Huang, Wenlong; Qian, Hai

    2015-11-15

    The free fatty acid receptor 1 (FFA1) is a novel antidiabetic target for the treatment of type 2 diabetes based on particular mechanism in amplifying glucose-stimulated insulin secretion. We have previously identified a series of phenoxyacetic acid derivatives. Herein, we describe the further chemical modification of this series directed by ligand efficiency and ligand lipophilicity efficiency. All of these efforts lead to the discovery of the promising candidate 16, an excellent FFA1 agonist with robust agonistic activity (43.6 nM), desired LE and LLE values. Moreover, compound 16 revealed a great potential for improving the hyperglycemia levels in both normal and type 2 diabetic mice without the risk of hypoglycemia even at the high dose of 40 mg/kg. PMID:26482570

  10. Modulation of GABA receptors expressed in Xenopus oocytes by 13-L-hydroxylinoleic acid and food additives.

    PubMed

    Aoshima, H; Tenpaku, Y

    1997-12-01

    To study the effects of 13-L-hydroxylinoleic acid (LOH) and food additives on gamma-aminobutyric acid (GABA) receptors, ionotropic GABA receptors were expressed in Xenopus oocytes by injecting mRNAs prepared from rat whole brain. LOH, which was prepared by reduction of 13-L-hydroperoxylinoleic acid (LOOH), inhibited the response of GABA receptors in the presence of high concentrations of GABA. LOH also inhibited nicotinic acetylcholine, glycine, and kainate receptors, while it had little effect on NMDA receptors expressed in Xenopus oocytes. However, LOH potentiated the response of GABA receptors as well as LOOH in the presence of low concentrations of GABA, possibly increasing the affinity of GABA for the receptors, while linoleic acid did not. Since some modification of the compounds seemed to change their effects on GABA receptors, the responses of GABA receptors elicited by 10 microM GABA were measured in the presence of compounds with various kinds of functional groups or the structural isomers of pentanol. Potentiation of GABA receptors depended strongly on the species of functional groups and also depended on the structure of the isomers. Then effects of various kinds of food additives on GABA receptors were also examined; perfumes such as alcohols or esters potentiated the responses strongly, while hexylamine, nicotinamide, or caffeine inhibited the responses, mainly in a competitive manner, and vanillin inhibited the responses noncompetitively. These results suggest the possibility that production of LOOH and LOH, or intake of much of some food additives, modulates the neural transmission in the brain, especially through ionotropic GABA receptors and changes the frame of the human mind, as alcohol or tobacco does.

  11. PTH1 Receptor Is Involved in Mediating Cellular Response to Long-Chain Polyunsaturated Fatty Acids

    PubMed Central

    Chachisvilis, Mirianas

    2012-01-01

    The molecular pathways by which long chain polyunsaturated fatty acids (LCPUFA) influence skeletal health remain elusive. Both LCPUFA and parathyroid hormone type 1 receptor (PTH1R) are known to be involved in bone metabolism while any direct link between the two is yet to be established. Here we report that LCPUFA are capable of direct, PTH1R dependent activation of extracellular ligand-regulated kinases (ERK). From a wide range of fatty acids studied, varying in chain length, saturation, and position of double bonds, eicosapentaenoic (EPA) and docosahexaenoic fatty acids (DHA) caused the highest ERK phosphorylation. Moreover, EPA potentiated the effect of parathyroid hormone (PTH(1–34)) in a superagonistic manner. EPA or DHA dependent ERK phosphorylation was inhibited by the PTH1R antagonist and by knockdown of PTH1R. Inhibition of PTH1R downstream signaling molecules, protein kinases A (PKA) and C (PKC), reduced EPA and DHA dependent ERK phosphorylation indicating that fatty acids predominantly activate G-protein pathway and not the β-arrestin pathway. Using picosecond time-resolved fluorescence microscopy and a genetically engineered PTH1R sensor (PTH-CC), we detected conformational responses to EPA similar to those caused by PTH(1–34). PTH1R antagonist blocked the EPA induced conformational response of the PTH-CC. Competitive binding studies using fluorescence anisotropy technique showed that EPA and DHA competitively bind to and alter the affinity of PTH1 receptor to PTH(1–34) leading to a superagonistic response. Finally, we showed that EPA stimulates protein kinase B (Akt) phosphorylation in a PTH1R-dependent manner and affects the osteoblast survival pathway, by inhibiting glucocorticoid-induced cell death. Our findings demonstrate for the first time that LCPUFAs, EPA and DHA, can activate PTH1R receptor at nanomolar concentrations and consequently provide a putative molecular mechanism for the action of fatty acids in bone. PMID:23300710

  12. Histamine (H3) receptors modulate the excitatory amino acid receptor response of the vestibular afferents.

    PubMed

    Chávez, Hortencia; Vega, Rosario; Soto, Enrique

    2005-12-01

    Although the effectiveness of histamine-related drugs in the treatment of peripheral and central vestibular disorders may be explained by their action on the vestibular nuclei, it has also been shown that antivertigo effects can take place at the peripheral level. In this work, we examined the actions of H3 histaminergic agonists and antagonists on the afferent neuron electrical discharge in the isolated inner ear of the axolotl. Our results indicate that H3 antagonists such as thioperamide, clobenpropit, and betahistine (BH) decreased the electrical discharge of afferent neurons by interfering with the postsynaptic response to excitatory amino acid agonists. These results lend further support to the idea that the antivertigo action of histamine-related drugs may be caused, at least in part, by a decrease in the sensory input from the vestibular endorgans. The present data show that the inhibitory action of the afferent neurons discharge previously described for BH is not restricted to this molecule but is also shared by other H3 antagonists.

  13. A Comparison of the Roles of Peroxisome Proliferator-Activated Receptor and Retinoic Acid Receptor on CYP26 Regulation

    PubMed Central

    Tay, Suzanne; Dickmann, Leslie; Dixit, Vaishali

    2010-01-01

    The cytochrome P450 26 family is believed to be responsible for all-trans-retinoic acid (atRA) metabolism and elimination in the human fetus and adults. CYP26A1 and CYP26B1 mRNA is expressed in a tissue-specific manner, and mice in which the CPY26 isoform has been knocked out show distinct malformations and lethality. The aim of this study was to determine differences in CYP26A1 and CYP26B1 regulation and expression. Analysis of CYP26A1 and CYP26B1 expression in a panel of 57 human livers showed CYP26A1 to be the major CYP26 isoform present in the liver, and its expression to be subject to large interindividual variability between donors. CYP26A1 and retinoic acid receptor (RAR) β were found to be greatly inducible by atRA in HepG2 cells, whereas CYP26B1, RARα, and RARγ were induced to a much lesser extent. Based on treatments with RAR isoform-selective ligands, RARα is the major isoform responsible for CYP26A1 and RARβ induction in HepG2 cells. Classic cytochrome P450 inducers did not affect CYP26 transcription, whereas the peroxisome proliferator-activated receptor (PPAR) γ agonists pioglitazone and rosiglitazone up-regulated CYP26B1 transcription by as much as 209- ± 80-fold and CYP26A1 by 10-fold. RARβ was also up-regulated by pioglitazone and rosiglitazone. CYP26B1 induction by PPARγ agonists was abolished by the irreversible PPARγ antagonist 2-chloro-5-nitrobenzanilide (GW9662), whereas RARβ and CYP26A1 induction was unaffected by GW9662. Overall, the results of this study suggest that CYP26B1 and CYP26A1 are regulated by different nuclear receptors, resulting in tissue-specific expression patterns. The fact that drugs can alter the expression of CYP26 enzymes may have toxicological and therapeutic importance. PMID:19884280

  14. 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.…

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

    PubMed

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

    2016-09-01

    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

  16. 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.

  17. 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

  18. Amino acid sequence coevolution in the insect bursicon ligand-receptor system.

    PubMed

    Hughes, Austin L

    2012-06-01

    The pattern of amino acid residue replacement in the components of the bursicon signaling system (involving the BURSα/BURSβ heterodimer and its receptor BURSrec) was reconstructed across a phylogeny of 17 insect species, in order to test for the co-occurrence of replacements at sets of individual sites. Sets of three or more branches with perfectly concordant changes occurred to a greater extent than expected by chance, given the observed level of amino acid change. The latter sites (SPC sites) were found to have distinctive characteristics: (1) the mean number of changes was significantly lower at SPC sites than that at other sites with multiple changes; (2) SPC sites had a significantly greater tendency toward parallel amino acid changes than other sites with multiple changes, but no greater tendency toward convergent changes; and (3) parallel changes tended to involve relatively similar amino acids, as indicated by relatively low mean chemical distances. The results implicated functional constraint, permitting only a limited subset of amino acids in a given site, as a major factor in causing both parallel amino acid replacement and coordinated amino acid changes in different sites of the same protein and of interacting proteins in this system.

  19. Role of the retinoic acid receptor-α in HIV-associated nephropathy.

    PubMed

    Ratnam, Krishna K; Feng, Xiaobei; Chuang, Peter Y; Verma, Vikram; Lu, Ting-Chi; Wang, Jinshan; Jin, Yuanmeng; Farias, Eduardo F; Napoli, Joseph L; Chen, Nan; Kaufman, Lewis; Takano, Tomoko; D'Agati, Vivette D; Klotman, Paul E; He, John C

    2011-03-01

    All-trans retinoic acid protects against the development of HIV-associated nephropathy (HIVAN) in HIV-1 transgenic mice (Tg26). In vitro, all-trans retinoic acid inhibits HIV-induced podocyte proliferation and restores podocyte differentiation markers by activating its receptor-α (RARα). Here, we report that Am580, a water-soluble RARα-specific agonist, attenuated proteinuria, glomerosclerosis, and podocyte proliferation, and restored podocyte differentiation markers in kidneys of Tg26 mice. Furthermore, RARα-/- Tg26 mice developed more severe kidney and podocyte injury than did RARα+/- Tg26 mice. Am580 failed to ameliorate kidney injury in RARα-/- Tg26 mice, confirming our hypothesis that Am580 acts through RARα. Although the expression of RARα-target genes was suppressed in the kidneys of Tg26 mice and of patients with HIVAN, the expression of RARα in the kidney was not different between patients with HIVAN and minimal change disease. However, the tissue levels of retinoic acid were reduced in the kidney cortex and isolated glomeruli of Tg26 mice. Consistent with this, the expression of two key enzymes in the retinoic acid synthetic pathway, retinol dehydrogenase type 1 and 9, and the overall enzymatic activity for retinoic acid synthesis were significantly reduced in the glomeruli of Tg26 mice. Thus, a defect in the endogenous synthesis of retinoic acid contributes to loss of the protection by retinoic acid in HIVAN. Hence, RARα agonists may be potential agents for the treatment of HIVAN.

  20. Mechanism of bile acid-regulated glucose and lipid metabolism in duodenal-jejunal bypass

    PubMed Central

    Chai, Jie; Zou, Lei; Li, Xirui; Han, Dali; Wang, Shan; Hu, Sanyuan; Guan, Jie

    2015-01-01

    Bile acid plays an important role in regulating blood glucose, lipid and energy metabolism. The present study was implemented to determine the effect of duodenal-jejunal bypass (DJB) on FXR, TGR-5expression in terminal ileum and its bile acid-related mechanism on glucose and lipid metabolism. Immunohistochemistry was used to detect relative gene or protein expression in liver and intestine. Firstly, we found that expression of FXR in liver and terminal ileum of DJB group was significantly higher than that in S-DJB group (P<0.05). In addition, DJB dramatically increased the activation of TGR-5 in the liver of rats. Furthermore, PEPCK, G6Pase, FBPase 1 and GLP-1 were up-regulated by DJB. In conclusion, these results showed that bile acid ameliorated glucose and lipid metabolism through bile acid-FXR and bile acid- TGR-5 signaling pathway. PMID:26884847

  1. Salicylic acid receptors activate jasmonic acid signalling through a non-canonical pathway to promote effector-triggered immunity

    PubMed Central

    Liu, Lijing; Sonbol, Fathi-Mohamed; Huot, Bethany; Gu, Yangnan; Withers, John; Mwimba, Musoki; Yao, Jian; He, Sheng Yang; Dong, Xinnian

    2016-01-01

    It is an apparent conundrum how plants evolved effector-triggered immunity (ETI), involving programmed cell death (PCD), as a major defence mechanism against biotrophic pathogens, because ETI-associated PCD could leave them vulnerable to necrotrophic pathogens that thrive on dead host cells. Interestingly, during ETI, the normally antagonistic defence hormones, salicylic acid (SA) and jasmonic acid (JA) associated with defence against biotrophs and necrotrophs respectively, both accumulate to high levels. In this study, we made the surprising finding that JA is a positive regulator of RPS2-mediated ETI. Early induction of JA-responsive genes and de novo JA synthesis following SA accumulation is activated through the SA receptors NPR3 and NPR4, instead of the JA receptor COI1. We provide evidence that NPR3 and NPR4 may mediate this effect by promoting degradation of the JA transcriptional repressor JAZs. This unique interplay between SA and JA offers a possible explanation of how plants can mount defence against a biotrophic pathogen without becoming vulnerable to necrotrophic pathogens. PMID:27725643

  2. 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

  3. Bile Acid Diarrhea: Prevalence, Pathogenesis, and Therapy

    PubMed Central

    Camilleri, Michael

    2015-01-01

    Bile acid diarrhea (BAD) is usually seen in patients with ileal Crohn’s disease or ileal resection. However, 25% to 50% of patients with functional diarrhea or diarrhea-predominant irritable bowel syndrome (IBS-D) also have evidence of BAD. It is estimated that 1% of the population may have BAD. The causes of BAD include a deficiency in fibroblast growth factor 19 (FGF-19), a hormone produced in enterocytes that regulates hepatic bile acid (BA) synthesis. Other potential causes include genetic variations that affect the proteins involved in BA enterohepatic circulation and synthesis or in the TGR5 receptor that mediates the actions of BA in colonic secretion and motility. BAs enhance mucosal permeability, induce water and electrolyte secretion, and accelerate colonic transit partly by stimulating propulsive high-amplitude colonic contractions. There is an increased proportion of primary BAs in the stool of patients with IBS-D, and some changes in the fecal microbiome have been described. There are several methods of diagnosing BAD, such as 75selenium homotaurocholic acid test retention, serum C4, FGF-19, and fecal BA measurement; presently, therapeutic trials with BA sequestrants are most commonly used for diagnosis. Management involves the use of BA sequestrants including cholestyramine, colestipol, and colesevelam. FXR agonists such as obeticholic acid constitute a promising new approach to treating BAD. PMID:25918262

  4. Novel Retinoic Acid Receptor Alpha Agonists for Treatment of Kidney Disease

    PubMed Central

    Liu, Ruijie; Li, Zhengzhe; Chen, Yibang; Evans, Todd; Chuang, Peter; Das, Bhaskar; He, John Cijiang

    2011-01-01

    Development of pharmacologic agents that protect podocytes from injury is a critical strategy for the treatment of kidney glomerular diseases. Retinoic acid reduces proteinuria and glomerulosclerosis in multiple animal models of kidney diseases. However, clinical studies are limited because of significant side effects of retinoic acid. Animal studies suggest that all trans retinoic acid (ATRA) attenuates proteinuria by protecting podocytes from injury. The physiological actions of ATRA are mediated by binding to all three isoforms of the nuclear retinoic acid receptors (RARs): RARα, RARβ, and RARγ. We have previously shown that ATRA exerts its renal protective effects mainly through the agonism of RARα. Here, we designed and synthesized a novel boron-containing derivative of the RARα-specific agonist Am580. This new derivative, BD4, binds to RARα receptor specifically and is predicted to have less toxicity based on its structure. We confirmed experimentally that BD4 binds to RARα with a higher affinity and exhibits less cellular toxicity than Am580 and ATRA. BD4 induces the expression of podocyte differentiation markers (synaptopodin, nephrin, and WT-1) in cultured podocytes. Finally, we confirmed that BD4 reduces proteinuria and improves kidney injury in HIV-1 transgenic mice, a model for HIV-associated nephropathy (HIVAN). Mice treated with BD4 did not develop any obvious toxicity or side effect. Our data suggest that BD4 is a novel RARα agonist, which could be used as a potential therapy for patients with kidney disease such as HIVAN. PMID:22125642

  5. Novel retinoic acid receptor alpha agonists for treatment of kidney disease.

    PubMed

    Zhong, Yifei; Wu, Yingwei; Liu, Ruijie; Li, Zhengzhe; Chen, Yibang; Evans, Todd; Chuang, Peter; Das, Bhaskar; He, John Cijiang

    2011-01-01

    Development of pharmacologic agents that protect podocytes from injury is a critical strategy for the treatment of kidney glomerular diseases. Retinoic acid reduces proteinuria and glomerulosclerosis in multiple animal models of kidney diseases. However, clinical studies are limited because of significant side effects of retinoic acid. Animal studies suggest that all trans retinoic acid (ATRA) attenuates proteinuria by protecting podocytes from injury. The physiological actions of ATRA are mediated by binding to all three isoforms of the nuclear retinoic acid receptors (RARs): RARα, RARβ, and RARγ. We have previously shown that ATRA exerts its renal protective effects mainly through the agonism of RARα. Here, we designed and synthesized a novel boron-containing derivative of the RARα-specific agonist Am580. This new derivative, BD4, binds to RARα receptor specifically and is predicted to have less toxicity based on its structure. We confirmed experimentally that BD4 binds to RARα with a higher affinity and exhibits less cellular toxicity than Am580 and ATRA. BD4 induces the expression of podocyte differentiation markers (synaptopodin, nephrin, and WT-1) in cultured podocytes. Finally, we confirmed that BD4 reduces proteinuria and improves kidney injury in HIV-1 transgenic mice, a model for HIV-associated nephropathy (HIVAN). Mice treated with BD4 did not develop any obvious toxicity or side effect. Our data suggest that BD4 is a novel RARα agonist, which could be used as a potential therapy for patients with kidney disease such as HIVAN.

  6. Predicting G-protein-coupled receptors families using different physiochemical properties and pseudo amino acid composition.

    PubMed

    Rehman, Zia-Ur; Mirza, Muhammad Tayyeb; Khan, Asifullah; Xhaard, Henri

    2013-01-01

    G-protein-coupled receptors (GPCRs) initiate signaling pathways via trimetric guanine nucleotide-binding proteins. GPCRs are classified based on their ligand-binding properties and molecular phylogenetic analyses. Nonetheless, these later analyses are in most case dependent on multiple sequence alignments, themselves dependent on human intervention and expertise. Alignment-free classifications of GPCR sequences, in addition to being unbiased, present many applications uncovering hidden physicochemical parameters shared among specific groups of receptors, to being used in automated workflows for large-scale molecular modeling applications. Current alignment-free classification methods, however, do not reach a full accuracy. This chapter discusses how GPCRs amino acid sequences can be classified using pseudo amino acid composition and multiscale energy representation of different physiochemical properties of amino acids. A hybrid feature extraction strategy is shown to be suitable to represent GPCRs and to be able to exploit GPCR amino acid sequence discrimination capability in spatial as well as transform domain. Classification strategies such as support vector machine and probabilistic neural network are then discussed in regards to GPCRs classification. The work of GPCR-Hybrid web predictor is also discussed.

  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. FLASH X-RAY (FXR) LINEAR INDUCTION ACCELERATOR (LIA) OPTIMIZATION Upgrade of the OTR Emittance Diagnostic

    SciTech Connect

    Houck, T L; Wargo, P E

    2006-12-01

    Knowing the electron beam parameters at the exit of an accelerator is critical for several reasons. Foremost is to optimize the application of the beam, which is flash radiography in the case of the FXR accelerator. The beam parameters not only determine the theoretical dose, x-ray spectrum, and radiograph resolution (spot size), they are required to calculate the final transport magnetic fields that focus the beam on the bremsstrahlung converter to achieve the theoretical limits. Equally important is the comparison of beam parameters to the design specifications. This comparison indicates the ''health'' of the accelerator, warning the operator when systems are deteriorating or failing. For an accelerator of the size and complexity of FXR, a large suite of diagnostics is normally employed to measure and/or infer beam parameters. These diagnostics are distributed throughout the accelerator and can require a large number of ''shots'' (measurements) to calculate a specific beam parameter. The OTR Emittance Diagnostic, however, has the potential to measure all but one of the beam parameters simultaneous at a specific location. Using measurements from a scan of a few shots, this final parameter can also be determined. Since first deployment, the OTR Emittance Diagnostic has been limited to measuring only one of the seven desired parameters, the beam's divergence. This report describes recent upgrades to the diagnostic that permit full realization of its potential.

  10. Glutamic acid decarboxylase and glutamate receptor changes during tolerance and dependence to benzodiazepines

    PubMed Central

    Izzo, Emanuela; Auta, James; Impagnatiello, Francesco; Pesold, Christine; Guidotti, Alessandro; Costa, Erminio

    2001-01-01

    Protracted administration of diazepam elicits tolerance, whereas discontinuation of treatment results in signs of dependence. Tolerance to the anticonvulsant action of diazepam is present in an early phase (6, 24, and 36 h) but disappears in a late phase (72–96 h) of withdrawal. In contrast, signs of dependence such as decrease in open-arm entries on an elevated plus-maze and increased susceptibility to pentylenetetrazol-induced seizures were apparent 96 h (but not 12, 24, or 48 h) after diazepam withdrawal. During the first 72 h of withdrawal, tolerance is associated with changes in the expression of GABAA (γ-aminobutyric acid type A) receptor subunits (decrease in γ2 and α1; increase in α5) and with an increase of mRNA expression of the most abundant form of glutamic acid decarboxylase (GAD), GAD67. In contrast, dl-α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit mRNA and cognate protein, which are normal during the early phase of diazepam withdrawal, increase by approximately 30% in cortex and hippocampus in association with the appearance of signs of dependence 96 h after diazepam withdrawal. Immunohistochemical studies of GluR1 subunit expression with gold-immunolabeling technique reveal that the increase of GluR1 subunit protein is localized to layer V pyramidal neurons and their apical dendrites in the cortex, and to pyramidal neurons and in their dendritic fields in hippocampus. The results suggest an involvement of GABA-mediated processes in the development and maintenance of tolerance to diazepam, whereas excitatory amino acid-related processes (presumably via AMPA receptors) may be involved in the expression of signs of dependence after withdrawal. PMID:11248104

  11. Glutamic acid decarboxylase and glutamate receptor changes during tolerance and dependence to benzodiazepines.

    PubMed

    Izzo, E; Auta, J; Impagnatiello, F; Pesold, C; Guidotti, A; Costa, E

    2001-03-13

    Protracted administration of diazepam elicits tolerance, whereas discontinuation of treatment results in signs of dependence. Tolerance to the anticonvulsant action of diazepam is present in an early phase (6, 24, and 36 h) but disappears in a late phase (72-96 h) of withdrawal. In contrast, signs of dependence such as decrease in open-arm entries on an elevated plus-maze and increased susceptibility to pentylenetetrazol-induced seizures were apparent 96 h (but not 12, 24, or 48 h) after diazepam withdrawal. During the first 72 h of withdrawal, tolerance is associated with changes in the expression of GABA(A) (gamma-aminobutyric acid type A) receptor subunits (decrease in gamma(2) and alpha(1); increase in alpha(5)) and with an increase of mRNA expression of the most abundant form of glutamic acid decarboxylase (GAD), GAD(67). In contrast, dl-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit mRNA and cognate protein, which are normal during the early phase of diazepam withdrawal, increase by approximately 30% in cortex and hippocampus in association with the appearance of signs of dependence 96 h after diazepam withdrawal. Immunohistochemical studies of GluR1 subunit expression with gold-immunolabeling technique reveal that the increase of GluR1 subunit protein is localized to layer V pyramidal neurons and their apical dendrites in the cortex, and to pyramidal neurons and in their dendritic fields in hippocampus. The results suggest an involvement of GABA-mediated processes in the development and maintenance of tolerance to diazepam, whereas excitatory amino acid-related processes (presumably via AMPA receptors) may be involved in the expression of signs of dependence after withdrawal.

  12. Retinoic Acid Receptor-Mediated Induction of ABCA1 in Macrophages

    PubMed Central

    Costet, Philippe; Lalanne, Florent; Gerbod-Giannone, Marie C.; Molina, Jennifer R.; Fu, Xuan; Lund, Erik G.; Gudas, Lorraine J.; Tall, Alan R.

    2003-01-01

    ABCA1, the mutant molecule in Tangier Disease, mediates efflux of cellular cholesterol to apoA-I and is induced by liver X receptor (LXR)/retinoid X receptor (RXR) transcription factors. Retinoic acid receptor (RAR) activators (all-trans-retinoic acid [ATRA] and TTNPB) were found to increase ATP-binding cassette transporter 1 (ABCA1) mRNA and protein in macrophages. In cellular cotransfection assays, RARγ/RXR activated the human ABCA1 promoter, via the same direct repeat 4 (DR4) promoter element as LXR/RXR. Chromatin immunoprecipitation analysis in macrophages confirmed the binding of RARγ/RXR to the ABCA1 promoter DR4 element in the presence of ATRA, with weaker binding of RARα/RXR, and no binding of RARβ/RXR. However, in macrophages from RARγ−/− mice, TTNPB still induced ABCA1, in association with marked upregulation of RARα, suggesting that high levels of RARα can compensate for the absence of RARγ. Dose-response experiments with ATRA in mouse primary macrophages showed that other LXR target genes were weakly induced (ABCG1 and SREBP-1c) or not induced (apoE and LXRα). The more specific RAR activator TTNPB did not induce SREBP-1c in mouse primary macrophages or liver. These studies indicate a direct role of RARγ/RXR in induction of macrophage ABCA1. PMID:14560020

  13. Functional Analysis of Free Fatty Acid Receptor GPR120 in Human Eosinophils: Implications in Metabolic Homeostasis

    PubMed Central

    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. PMID:25790291

  14. Recognition and sequestration of ω-fatty acids by a cavitand receptor

    PubMed Central

    Mosca, Simone; Ajami, Dariush; Rebek, Julius

    2015-01-01

    One of the largest driving forces for molecular association in aqueous solution is the hydrophobic effect, and many synthetic receptors with hydrophobic interiors have been devised for molecular recognition studies in water. Attempts to create the longer, narrower cavities appropriate for long-chain fatty acids have been thwarted by solvophobic collapse of the synthetic receptors, giving structures that have no internal spaces. The collapse generally involves the stacking of aromatic panels onto themselves. We describe here the synthesis and application of a deep cavitand receptor featuring “prestacked” aromatic panels at the upper rim of the binding pocket. The cavitand remains open and readily sequesters biologically relevant long-chain molecules—unsaturated ω-3, -6, and -9 fatty acids and derivatives such as anandamide—from aqueous media. The cavitand exists in isomeric forms with different stacking geometries and n-alkanes were used to characterize the binding modes and conformational properties. Long alkyl chains are accommodated in inverted J-shaped conformations. An analogous cavitand with electron-rich aromatic walls was prepared and comparative binding experiments indicated the role of intramolecular stacking in the binding properties of these deep container molecules. PMID:26305974

  15. Recognition and sequestration of ω-fatty acids by a cavitand receptor.

    PubMed

    Mosca, Simone; Ajami, Dariush; Rebek, Julius

    2015-09-01

    One of the largest driving forces for molecular association in aqueous solution is the hydrophobic effect, and many synthetic receptors with hydrophobic interiors have been devised for molecular recognition studies in water. Attempts to create the longer, narrower cavities appropriate for long-chain fatty acids have been thwarted by solvophobic collapse of the synthetic receptors, giving structures that have no internal spaces. The collapse generally involves the stacking of aromatic panels onto themselves. We describe here the synthesis and application of a deep cavitand receptor featuring "prestacked" aromatic panels at the upper rim of the binding pocket. The cavitand remains open and readily sequesters biologically relevant long-chain molecules-unsaturated ω-3, -6, and -9 fatty acids and derivatives such as anandamide-from aqueous media. The cavitand exists in isomeric forms with different stacking geometries and n-alkanes were used to characterize the binding modes and conformational properties. Long alkyl chains are accommodated in inverted J-shaped conformations. An analogous cavitand with electron-rich aromatic walls was prepared and comparative binding experiments indicated the role of intramolecular stacking in the binding properties of these deep container molecules.

  16. Comparison of the effects of pelargonic acid vanillylamide and capsaicin on human vanilloid receptors.

    PubMed

    Weiser, Thomas; Roufogalis, Basil; Chrubasik, Sigrun

    2013-07-01

    Pelargonic acid vanillylamide is like capsaicin a natural capsaicinoid from chili peppers and commonly used in food additives to create a hot sensation, even in self-defense pepper sprays and as an alternative to capsaicin in medical products for topical treatment of pain. Although the chemical structures of both compounds are similar, preclinical data suggest that capsaicin is the more potent compound. We therefore performed voltage-clamp recordings using cells transfected with the human vanilloid receptor TRPV1 in order to assess the responses of pelargonic acid vanillylamide and capsaicin at the receptor level. We provide evidence that at the molecular target TRPV1, the concentration-response curves, kinetics of current activation, as well as inhibition by the competitive antagonist capsazepine were not significantly different between the two capsaicinoids. We suggest that the different effects of the two capsaicinoids observed in previous studies may rather be due to different physicochemical or pharmacokinetic properties than to different pharmacological profiles at the receptor level. PMID:22961689

  17. Comparison of the effects of pelargonic acid vanillylamide and capsaicin on human vanilloid receptors.

    PubMed

    Weiser, Thomas; Roufogalis, Basil; Chrubasik, Sigrun

    2013-07-01

    Pelargonic acid vanillylamide is like capsaicin a natural capsaicinoid from chili peppers and commonly used in food additives to create a hot sensation, even in self-defense pepper sprays and as an alternative to capsaicin in medical products for topical treatment of pain. Although the chemical structures of both compounds are similar, preclinical data suggest that capsaicin is the more potent compound. We therefore performed voltage-clamp recordings using cells transfected with the human vanilloid receptor TRPV1 in order to assess the responses of pelargonic acid vanillylamide and capsaicin at the receptor level. We provide evidence that at the molecular target TRPV1, the concentration-response curves, kinetics of current activation, as well as inhibition by the competitive antagonist capsazepine were not significantly different between the two capsaicinoids. We suggest that the different effects of the two capsaicinoids observed in previous studies may rather be due to different physicochemical or pharmacokinetic properties than to different pharmacological profiles at the receptor level.

  18. Development and Characterization of a Potent Free Fatty Acid Receptor 1 (FFA1) Fluorescent Tracer.

    PubMed

    Christiansen, Elisabeth; Hudson, Brian D; Hansen, Anders Højgaard; Milligan, Graeme; Ulven, Trond

    2016-05-26

    The free fatty acid receptor 1 (FFA1/GPR40) is a potential target for treatment of type 2 diabetes. Although several potent agonists have been described, there remains a strong need for suitable tracers to interrogate ligand binding to this receptor. We address this by exploring fluorophore-tethering to known potent FFA1 agonists. This led to the development of 4, a high affinity FFA1 tracer with favorable and polarity-dependent fluorescent properties. A close to ideal overlap between the emission spectrum of the NanoLuciferase receptor tag and the excitation spectrum of 4 enabled the establishment of a homogeneous BRET-based binding assay suitable for both detailed kinetic studies and high throughput competition binding studies. Using 4 as a tracer demonstrated that the compound acts fully competitively with selected synthetic agonists but not with lauric acid and allowed for the characterization of binding affinities of a diverse selection of known FFA1 agonists, indicating that 4 will be a valuable tool for future studies at FFA1. PMID:27074625

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

    PubMed Central

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

    2013-01-01

    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 α2,6linked (human type) sialic acid (SA) influenza virus receptors in tissues is considered to be 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 (SNA) 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. PMID:23391183

  20. Plakophilins 1 and 3 Bind to FXR1 and Thereby Influence the mRNA Stability of Desmosomal Proteins

    PubMed Central

    Fischer-Kešo, Regina; Breuninger, Sonja; Hofmann, Sarah; Henn, Manuela; Röhrig, Theresa; Ströbel, Philipp; Stoecklin, Georg

    2014-01-01

    Plakophilins 1 and 3 (PKP1/3) are members of the arm repeat family of catenin proteins and serve as structural components of desmosomes, which are important for cell-cell-adhesion. In addition, PKP1/3 occur as soluble proteins outside desmosomes, yet their role in the cytoplasm is not known. We found that cytoplasmic PKP1/3 coprecipitated with the RNA-binding proteins FXR1, G3BP, PABPC1, and UPF1, and these PKP1/3 complexes also comprised desmoplakin and PKP2 mRNAs. Moreover, we showed that the interaction of PKP1/3 with G3BP, PABPC1, and UPF1 but not with FXR1 was RNase sensitive. To address the cytoplasmic function of PKP1/3, we performed gain-and-loss-of-function studies. Both PKP1 and PKP3 knockdown cell lines showed reduced protein and mRNA levels for desmoplakin and PKP2. Whereas global rates of translation were unaffected, desmoplakin and PKP2 mRNA were destabilized. Furthermore, binding of PKP1/3 to FXR1 was RNA independent, and both PKP3 and FXR1 stabilized PKP2 mRNA. Our results demonstrate that cytoplasmic PKP1/3 are components of mRNA ribonucleoprotein particles and act as posttranscriptional regulators of gene expression. PMID:25225333

  1. Effects of perfluoroalkyl acids on the function of the thyroid hormone and the aryl hydrocarbon receptor.

    PubMed

    Long, Manhai; Ghisari, Mandana; Bonefeld-Jørgensen, Eva Cecilie

    2013-11-01

    Perfluoroalkyl acids (PFAAs) are perfluorinated compounds that widely exist in the environment and can elicit adverse effects including endocrine disruption in humans and animals. This study investigated the effect of seven PFAAs on the thyroid hormone (TH) system assessing the proliferation of the 3,3',5-triiodo-L-thryonine (T3)-dependent rat pituitary GH3 cells using the T-screen assay and the effect on the aryl hydrocarbon receptor (AhR) transactivation in the AhR-luciferase reporter gene bioassay. A dose-dependent impact on GH3 cells was observed in the range 1×10(-9)-1×10(-4) M: seven PFAAs (perfluorooctane sulfonate (PFOS), perfluorohexane sulfonate (PFHxS), perfluorooctanoic acid, perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluoroundecanoic acid (PFUnA), and perfluorododecanoic acid (PFDoA)) inhibited the GH3 cell growth, and four PFAAs (PFOS, PFHxS, PFNA, and PFUnA) antagonized the T3-induced GH3 cell proliferation. At the highest test concentration, PFHxS showed a further increase of the T3-induced GH3 growth. Among the seven tested PFAAs, only PFDoA and PFDA elicited an activating effect on the AhR. In conclusion, PFAAs possess in vitro endocrine-disrupting potential by interfering with TH and AhR functions, which need to be taken into consideration when assessing the impact on human health. PMID:23539207

  2. Mapping amino acids of the measles virus hemagglutinin responsible for receptor (CD46) downregulation.

    PubMed

    Bartz, R; Brinckmann, U; Dunster, L M; Rima, B; Ter Meulen, V; Schneider-Schaulies, J

    1996-10-01

    We compared the amino acid sequences of groups of receptor (CD46) downregulating and nondownregulating measles virus (MV) hemagglutinins (Hs) and identified seven group-specific differences as candidates for the mediation of the observed differential effects. Using site-directed mutagenesis, we mutated the chosen amino acids of the H of MV-strain WTF (WTF-H), a nondownregulating H, and Introduced the corresponding amino acids of Edmonston-H (Edm-H), a downregulating H. We identified four amino acids, 211G, 243R, 451V, and 481Y, which influenced the downregulative function when introduced into WTF-H. The double mutation 451V and 481Y in WTF-H led to a degree of CD46 downregulation comparable to that of Edm-H. Conversely, introducing amino acids 451E and 481N into Edm-H resulted in a loss of the downregulative function. These results indicate that these amino acids play a decisive role in the H-CD46 interaction. PMID:8862431

  3. Linking γ-aminobutyric acid A receptor to epidermal growth factor receptor pathways activation in human prostate cancer.

    PubMed

    Wu, Weijuan; Yang, Qing; Fung, Kar-Ming; Humphreys, Mitchell R; Brame, Lacy S; Cao, Amy; Fang, Yu-Ting; Shih, Pin-Tsen; Kropp, Bradley P; Lin, Hsueh-Kung

    2014-03-01

    Neuroendocrine (NE) differentiation has been attributed to the progression of castration-resistant prostate cancer (CRPC). Growth factor pathways including the epidermal growth factor receptor (EGFR) signaling have been implicated in the development of NE features and progression to a castration-resistant phenotype. However, upstream molecules that regulate the growth factor pathway remain largely unknown. Using androgen-insensitive bone metastasis PC-3 cells and androgen-sensitive lymph node metastasis LNCaP cells derived from human prostate cancer (PCa) patients, we demonstrated that γ-aminobutyric acid A receptor (GABA(A)R) ligand (GABA) and agonist (isoguvacine) stimulate cell proliferation, enhance EGF family members expression, and activate EGFR and a downstream signaling molecule, Src, in both PC-3 and LNCaP cells. Inclusion of a GABA(A)R antagonist, picrotoxin, or an EGFR tyrosine kinase inhibitor, Gefitinib (ZD1839 or Iressa), blocked isoguvacine and GABA-stimulated cell growth, trans-phospohorylation of EGFR, and tyrosyl phosphorylation of Src in both PCa cell lines. Spatial distributions of GABAAR α₁ and phosphorylated Src (Tyr416) were studied in human prostate tissues by immunohistochemistry. In contrast to extremely low or absence of GABA(A)R α₁-positive immunoreactivity in normal prostate epithelium, elevated GABA(A)R α₁ immunoreactivity was detected in prostate carcinomatous glands. Similarly, immunoreactivity of phospho-Src (Tyr416) was specifically localized and limited to the nucleoli of all invasive prostate carcinoma cells, but negative in normal tissues. Strong GABAAR α₁ immunoreactivity was spatially adjacent to the neoplastic glands where strong phospho-Src (Tyr416)-positive immunoreactivity was demonstrated, but not in adjacent to normal glands. These results suggest that the GABA signaling is linked to the EGFR pathway and may work through autocrine or paracine mechanism to promote CRPC progression.

  4. Bile acids as metabolic regulators

    PubMed Central

    Li, Tiangang; Chiang, John Y. L.

    2015-01-01

    Summary Small molecule ligands that target to TGR5 and FXR have shown promise in treating various metabolic and inflammation-related human diseases. New insights into the mechanisms underlying the bariatric surgery and bile acid sequestrant treatment suggest that targeting the enterohepatic circulation to modulate gut-liver bile acid signaling, incretin production and microbiota represents a new strategy to treat obesity and type-2 diabetes. PMID:25584736

  5. 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

  6. Dissecting the Role of Retinoic Acid Receptor Isoforms in the CD8 Response to Infection

    PubMed Central

    Guo, Yanxia; Lee, Yu-Chi; Brown, Chrysothemis; Zhang, Weijun; Usherwood, Edward; Noelle, Randolph J.

    2015-01-01

    Vitamin A deficiency leads to increased susceptibility to a spectrum of infectious diseases. The studies presented dissect the intrinsic role of each of the retinoic acid receptor (RAR) isoforms in the clonal expansion, differentiation, and survival of pathogen-specific CD8 T cells in vivo. The data show that RARα is required for the expression of gut-homing receptors on CD8+ T cells and survival of CD8+ T cells in vitro. Furthermore, RARα is essential for survival of CD8+ T cells in vivo following Listeria monocytogenes infection. In contrast, RARβ deletion leads to modest deficiency in Ag-specific CD8+ T cell expansion during infection. The defective survival of RARα-deficient CD8+ T cells leads to a deficiency in control of L. monocytogenes expansion in the spleen. To our knowledge, these are the first comparative studies of the role of RAR isoforms in CD8+ T cell immunity. PMID:24610012

  7. Bile acid transporters and regulatory nuclear receptors in the liver and beyond

    PubMed Central

    Halilbasic, Emina; Claudel, Thierry; Trauner, Michael

    2013-01-01

    Summary Bile acid (BA) transporters are critical for maintenance of the enterohepatic BA circulation where BAs exert their multiple physiological functions including stimulation of bile flow, intestinal absorption of lipophilic nutrients, solubilization and excretion of cholesterol, as well as antimicrobial and metabolic effects. Tight regulation of BA transporters via nuclear receptors is necessary to maintain proper BA homeostasis. Hereditary and acquired defects of BA transporters are involved in the pathogenesis of several hepatobiliary disorders including cholestasis, gallstones, fatty liver disease and liver cancer, but also play a role in intestinal and metabolic disorders beyond the liver. Thus, pharmacological modification of BA transporters and their regulatory nuclear receptors opens novel treatment strategies for a wide range of disorders. PMID:22885388

  8. 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

  9. Inhibitory effects of lysophosphatidic acid receptor-5 on cellular functions of sarcoma cells.

    PubMed

    Araki, Mutsumi; Kitayoshi, Misaho; Dong, Yan; Hirane, Miku; Ozaki, Shuhei; Mori, Shiori; Fukushima, Nobuyuki; Honoki, Kanya; Tsujiuchi, Toshifumi

    2014-06-01

    Lysophosphatidic acid (LPA) is a bioactive lipid that interacts with G protein-coupled LPA receptors (LPA receptor-1 (LPA1) to LPA6). Here, we investigated the effects of LPA signaling via LPA5 on cellular functions of sarcoma cells by generating Lpar5 overexpressing and Lpar5 knockdown cells from rat osteosarcoma and malignant fibrous histiocytoma cells, respectively. The cell motility activity of Lpar5 overexpressing cells was significantly lower, while Lpar5 knockdown cells showed high cell motility, compared with respective controls. Gelatin zymography showed that LPA5 suppressed the activation of matrix metalloproteinase-2. LPA5 also inhibited the cell motility activity of endothelial cells, correlating with the expression levels of vascular endothelial growth factor genes. These results suggest that LPA signaling via LPA5 negatively regulates the cellular functions of rat sarcoma cells. PMID:24798396

  10. 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.

  11. 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

  12. 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. PMID:16290140

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

    PubMed

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

    2016-10-01

    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

  14. 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.

  15. 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

  16. Distinct Pathways of ERK1/2 Activation by Hydroxy-Carboxylic Acid Receptor-1

    PubMed Central

    Li, Guo; Wang, Hui-qian; Wang, Li-hui; Chen, Ru-ping; Liu, Jun-ping

    2014-01-01

    Mechanistic investigations have shown that, upon agonist activation, hydroxy-carboxylic acid receptor-1(HCA1) couples to a Gi protein and inhibits adenylate cyclase activity, leading to inhibition of liberation of free fatty acid. However, the underlying molecular mechanisms for HCA1 signaling remain largely unknown. Using CHO-K1 cells stably expressing HCA1, and L6 cells, which endogenously express rat HCA1 receptors, we found that activation of ERK1/2 by HCA1 was rapid, peaking at 5 min, and was significantly blocked by pertussis toxin. Furthermore, time course experiments with different kinase inhibitors demonstrated that HCA1 induced ERK1/2 activation via the extracellular Ca2+, PKC and IGF-I receptor transactivation-dependent pathways. In addition, we observed that pretreated the cells with M119K, an inhibitor of Gβγ subunit-dependent signaling, effectively attenuated the ERK1/2 activation triggered by HCA1, suggesting a critical role for βγ-subunits in HCA1-activated ERK1/2 phosphorylation. Furthermore, the present results also indicated that the arrestin2/3 were not required for ERK1/2 activation. In conclusion, our findings demonstrate that upon binding to agonist, HCA1 receptors initially activate Gi, leading to dissociation of the Gβγ subunit from activated Gi, and subsequently induce ERK1/2 activation via two distinct pathways: one PKC-dependent pathway and the other IGF-IR transactivation-dependent pathway. Our results provide the first in-depth evidence that defines the molecular mechanism of HCA1-mediated ERK1/2 activation. PMID:24671202

  17. Elicidation by a H-2-receptor antagonist of the significance of mucosal histamine mobilization in exciting acid secretion.

    PubMed Central

    Lundell, L

    1975-01-01

    1. The consequence of H-2-receptor blockade for the secretory responses of the gastric mucosa to hormonal or cholinergic stimulation was studied in conscious rats with Heindenhain pouches or Pavlov pouches with the antrum retained or resected. 2. Metiamide almost completely abolished acid secretion induced by pentagastrin without altering significantly the amount of histamine excreted in the urine. Histamine mobilization on pentagastrin infusion determined in vitro, seemed to be larger during H-2-receptor blockade than with pentagastrin alone. 3. CCK-PZ mobilized mucosal histamine to a considerable extent; the secretory response to this hormone was completely abolished by H-2-receptor blockade. 4. Acid secretion in response to 2-deoxy-D-glucose was inhibited by H-2-receptor blockade in the presence or absence of the antrum; however the inhibition was less complete than with hormone-induced secretion. 5. The acid secretory response to 100 mg/kg of 2-deoxy-D-glucose appeared to be less susceptible to H-2-receptor blockade than that of 50-mg/kg of 2-deoxy-D-glucose. 6. Feeding induced a secretory response in the Pavlov pouch which initially was more effectively inhibited by H-2-receptor blockade than the response to 2-deoxy-D-glucose. In the absence of antral gastrin secretion by either stimulus was equally inhibited. 7. Methacholine-induced acid secretion was inhibited by infusion of the H-2-receptor antagonist, an inhibition that was absent when pentagastrin was concomitantly infused. 8. Although acid secretion induced by cholinergic stimuli was readily inhibited by the H-2-receptor antagonist, slight or nor inhibition was noted on pepsin secretion. 9. The role of histamine as a physiological stimulus for the parietal cell is discussed in view of the fact that the secretory effect of natural stimuli, known or demonstrated to mobilize mucosal histamine, is restrained by H-2-receptor blockade. PMID:49418

  18. G-protein-coupled receptors for neurotransmitter amino acids: C-terminal tails, crowded signalosomes.

    PubMed Central

    El Far, Oussama; Betz, Heinrich

    2002-01-01

    G-protein-coupled receptors (GPCRs) represent a superfamily of highly diverse integral membrane proteins that transduce external signals to different subcellular compartments, including nuclei, via trimeric G-proteins. By differential activation of diffusible G(alpha) and membrane-bound G(beta)gamma subunits, GPCRs might act on both cytoplasmic/intracellular and plasma-membrane-bound effector systems. The coupling efficiency and the plasma membrane localization of GPCRs are regulated by a variety of interacting proteins. In this review, we discuss recently disclosed protein interactions found with the cytoplasmic C-terminal tail regions of two types of presynaptic neurotransmitter receptors, the group III metabotropic glutamate receptors and the gamma-aminobutyric acid type-B receptors (GABA(B)Rs). Calmodulin binding to mGluR7 and other group III mGluRs may provide a Ca(2+)-dependent switch for unidirectional (G(alpha)) versus bidirectional (G(alpha) and G(beta)gamma) signalling to downstream effector proteins. In addition, clustering of mGluR7 by PICK1 (protein interacting with C-kinase 1), a polyspecific PDZ (PSD-95/Dlg1/ZO-1) domain containing synaptic organizer protein, sheds light on how higher-order receptor complexes with regulatory enzymes (or 'signalosomes') could be formed. The interaction of GABA(B)Rs with the adaptor protein 14-3-3 and the transcription factor ATF4 (activating transcription factor 4) suggests novel regulatory pathways for G-protein signalling, cytoskeletal reorganization and nuclear gene expression: processes that may all contribute to synaptic plasticity. PMID:12006104

  19. Gintonin enhances performance of mice in rotarod test: Involvement of lysophosphatidic acid receptors and catecholamine release.

    PubMed

    Lee, Byung-Hwan; Kim, Jisu; Lee, Ra Mi; Choi, Sun-Hye; Kim, Hyeon-Joong; Hwang, Sung-Hee; Lee, Myung Koo; Bae, Chun-Sik; Kim, Hyoung-Chun; Rhim, Hyewon; Lim, Kiwon; Nah, Seung-Yeol

    2016-01-26

    Ginseng has a long history of use as a tonic for restoration of vigor. One example of ginseng-derived tonic effect is that it can improve physical stamina under conditions of stress. However, the active ingredient and the underlying molecular mechanism responsible for the ergogenic effect are unknown. Recent studies show that ginseng contains a novel ingredient, gintonin, which consists of a unique class of herbal-medicine lysophosphatidic acids (LPAs). Gintonin activates G protein-coupled LPA receptors to produce a transient [Ca(2+)]i signal, which is coupled to diverse intra- and inter-cellular signal transduction pathways that stimulate hormone or neurotransmitter release. However, relatively little is known about how gintonin-mediated cellular modulation is linked to physical endurance. In the present study, systemic administration of gintonin, but not ginsenosides, in fasted mice increased blood glucose concentrations in a dose-dependent manner. Gintonin treatment elevated blood glucose to a maximum level after 30min. This elevation in blood glucose level could be abrogated by the LPA1/3 receptor antagonist, Ki16425, or the β-adrenergic receptor antagonist, propranolol. Furthermore, gintonin-dependent enhanced performance of fasted mice in rotarod test was likewise abrogated by Ki16425. Gintonin also elevated plasma epinephrine and norepinephrine concentrations. The present study shows that gintonin mediates catecholamine release through activation of the LPA receptor and that activation of the β-adrenergic receptor is coupled to liver glycogenolysis, thereby increasing the supply of glucose and enhancing performance in the rotarod test. Thus, gintonin acts via the LPA-catecholamine-glycogenolysis axis, representing a candidate mechanism that can explain how ginseng treatment enhances physical stamina. PMID:26706688

  20. Arginine of retinoic acid receptor beta which coordinates with the carboxyl group of retinoic acid functions independent of the amino acid residues responsible for retinoic acid receptor subtype ligand specificity.

    PubMed

    Zhang, Zeng Ping; Hutcheson, Juliet M; Poynton, Helen C; Gabriel, Jerome L; Soprano, Kenneth J; Soprano, Dianne Robert

    2003-01-15

    The biological actions of retinoic acid (RA) are mediated by retinoic acid receptors (RARalpha, RARbeta, and RARgamma) and retinoid X receptors (RXRalpha, RXRbeta, and RXRgamma). Consistent with the X-ray crystal structures of RARalpha and RARgamma, site-directed mutagenesis studies have demonstrated the importance of a conserved Arg residue (alphaArg(276), betaArg(269), and gammaArg(278)) for coordination with the carboxyl group of RA. However, mutation of Arg(269) to Ala in RARbeta causes only a 3- to 6-fold increase in the K(d) for RA and EC(50) in RA-dependent transcriptional transactivation assays while the homologous mutation in either RARalpha or RARgamma causes a 110-fold and a 45-fold increase in EC(50) value, respectively. To further investigate the nature of this difference, we prepared mutant RARs to determine the effect of conversion of betaR269A to a mutant which mimics either RARalpha ligand selectivity (betaA225S/R269A) or RARgamma ligand selectivity (betaI263M/R269A/V338A). Our results demonstrate that in RARbeta mutants that acquire either RARalpha or RARgamma ligand specificity the Arg(269) position responsible for coordination with the carboxyl group of retinoids continued to function like that of RARbeta. Furthermore, three mutant receptors (betaA225S/R269A, betaA225S/F279, and alphaF286A) were found to have a greater than wild-type affinity for the RARalpha-selective ligand Am580. Finally, a homology-based computer model of the ligand binding domain (LBD) of RARbeta and the X-ray crystal structures of the LBD of both RARalpha and RARgamma are used to describe potential mechanisms responsible for the increased affinity of some mutants for Am580 and for the difference in the effect of mutation of Arg(269) in RARbeta compared to its homologous Arg in RARalpha and RARgamma.

  1. Recessive and dominant mutations in retinoic acid receptor beta in cases with microphthalmia and diaphragmatic hernia.

    PubMed

    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

    2013-10-01

    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.

  2. 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. PMID:26394664

  3. Retinoic