RESUMEN
Obesity leads to insulin resistance and is a major risk factor for the development of diabetes mellitus in cats. Prevention of obesity and obesity-induced insulin resistance is difficult, and reliable long-term strategies are currently lacking. Retinoid-related orphan receptor gamma (RORγ) was recently identified as an important transcription factor in the development of large insulin-resistant adipocytes in mice and humans. RORγ negatively affects adipocyte differentiation through expression of its target gene matrix metalloproteinase 3 (MMP3) and promotes the development of large insulin-resistant adipocytes. Preliminary studies in mice showed that RORγ can be inhibited by its ligand tetra-hydroxylated bile acid (THBA). In the present study, serum THBA levels were determined in healthy and diabetic cats. Moreover, potential side effects and the effects of THBA supplementation on adipocyte size, mRNA expression of RORγ, MMP3, interleukin 6, tumor necrosis factor α, adiponectin and leptin in feline subcutaneous adipocytes and insulin sensitivity were investigated in healthy normal weight cats. Thirteen healthy and 13 diabetic cats were used for determination of serum THBA level, and six healthy normal-weight cats were included in a feeding trial. Similar THBA levels were determined in serum of healthy and diabetic cats. Supplementation of 5 mg/kg THBA for 8 wk did not cause any negative effect on feeding behavior, general condition and blood parameters of tested cats. It significantly reduced adipocyte size and mRNA expression of MMP3, interleukin 6, and tumor necrosis factor α in adipocytes, while mRNA expression of adiponectin significantly increased and mRNA expression of RORγ and leptin remained unchanged. Administration of THBA did not influence fasting blood glucose levels or the response of cats to acute insulin administration. Based on these results, THBA is palatable and is considered safe for use in cats. It reduces expression of MMP3 and promotes the development of small adipocytes with increased expression of adiponectin and reduced expression of interleukin 6 and tumor necrosis factor α. Further studies are recommended to evaluate the effect of THBA on adipocyte size and insulin sensitivity in obese cats.
Asunto(s)
Enfermedades de los Gatos , Diabetes Mellitus , Resistencia a la Insulina , Obesidad , Enfermedades de los Roedores , Adipocitos/metabolismo , Adiponectina , Animales , Ácidos y Sales Biliares/metabolismo , Enfermedades de los Gatos/metabolismo , Gatos , Diabetes Mellitus/veterinaria , Insulina/metabolismo , Resistencia a la Insulina/fisiología , Interleucina-6/farmacología , Leptina , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/farmacología , Ratones , Obesidad/metabolismo , Obesidad/veterinaria , ARN Mensajero/metabolismo , Enfermedades de los Roedores/metabolismo , Enfermedades de los Roedores/patología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Epidemiological studies have shown an association between maternal overnutrition and increased risk of the progeny for the development of obesity as well as psychiatric disorders. Animal studies have shown results regarding maternal high-fat diet (HFD) and a greater risk of the offspring to develop obesity. However, it still remains unknown whether maternal HFD can program the central reward system in such a way that it will imprint long-term changes that will predispose the offspring to addictive-like behaviors that may lead to obesity. We exposed female dams to either laboratory chow or HFD for a period of 9 weeks: 3 weeks before conception, during gestation and lactation. Offspring born to either control or HFD-exposed dams were examined in behavioral, neurochemical, neuroanatomical, metabolic and positron emission tomography (PET) scan tests. Our results demonstrate that HFD offspring compared with controls consume more alcohol, exhibit increased sensitivity to amphetamine and show greater conditioned place preference to cocaine. In addition, maternal HFD leads to increased preference to sucrose as well as to HFD while leaving the general feeding behavior intact. The hedonic behavioral alterations are accompanied by reduction of striatal dopamine and by increased dopamine 2 receptors in the same brain region as evaluated by post-mortem neurochemical, immunohistochemical as well as PET analyses. Taken together, our data suggest that maternal overnutrition predisposes the offspring to develop hedonic-like behaviors to both drugs of abuse as well as palatable foods and that these types of behaviors may share common neuronal underlying mechanisms that can lead to obesity.
Asunto(s)
Conducta Adictiva/fisiopatología , Conducta Adictiva/psicología , Dieta Alta en Grasa , Preferencias Alimentarias/fisiología , Preferencias Alimentarias/psicología , Drogas Ilícitas , Obesidad/fisiopatología , Complicaciones del Embarazo/fisiopatología , Efectos Tardíos de la Exposición Prenatal , Gusto/fisiología , Alcoholismo/fisiopatología , Alcoholismo/psicología , Trastornos Relacionados con Anfetaminas/fisiopatología , Trastornos Relacionados con Anfetaminas/psicología , Animales , Peso Corporal/fisiología , Trastornos Relacionados con Cocaína/fisiopatología , Trastornos Relacionados con Cocaína/psicología , Cuerpo Estriado/fisiopatología , Dopamina/metabolismo , Conducta Alimentaria/fisiología , Femenino , Impronta Psicológica/fisiología , Masculino , Ratones Endogámicos C57BL , Embarazo , Receptores de Dopamina D2/metabolismo , Factores de Riesgo , Sacarosa/administración & dosificaciónRESUMEN
BACKGROUND: Predicting an individual's success in a non-surgical weight loss approach is a demanding need since obesity is becoming an epidemic burden. A possible predictive marker is myostatin, a member of the transforming growth factor b superfamily, which has been shown to be an important regulator of muscle homeostasis. METHODS: In the present study, we analyzed myostatin as a marker to predict weight loss of patients that participated in a 2 phased weight reduction program, comprising a weight loss period of 12 weeks and a weight stabilization period of 40 weeks. Therefore, 62 obese individuals with a mean BMI of 40.6 kg/m(2) were included. Plasma myostatin was measured with ELISA at the beginning (T0), after weight loss (T1) and at the end of the program (T2). RESULTS: Although significant weight loss of -23.9±14.9 kg was achieved, myostatin did not change significantly during the program (T0>T1: p=0.46; T1>T2: p=0.70; T0>T2: p=0.57). Myostatin at baseline did neither negatively correlate with the achieved weight loss in the weight reduction phase (T0>T1: r=0.27, p=0.16) nor with weight loss during the whole program (T0>T2: r=0.20, p=0.29). Only a minor correlation with myostatin levels after weight loss with weight regain during maintenance period was detected. (T1>T2: r=-0.37, p=0.05). CONCLUSION: Plasma myostatin might be suitable in predicting weight regain after marked weight loss, but no association with weight loss was observed in patients undergoing a non-surgical weight loss program. Therefore, myostatin does not seem to be a predictor for success in non-surgical weight loss approaches.
Asunto(s)
Miostatina/sangre , Obesidad/sangre , Obesidad/terapia , Pérdida de Peso , Adulto , Anciano , Biomarcadores/sangre , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
OBJECTIVE: The A-allele of the fat mass and obesity-associated (FTO) gene variant rs9939609 has been associated with increased body weight, whereas no effect on weight loss during weight reduction programs has been observed. We questioned whether the AA-genotype interferes with weight stabilization after weight loss. DESIGN: We conducted a monocentric, longitudinal study involving obese individuals. The FTO gene variant rs9939609 was genotyped in participants attending a weight reduction program that was divided into two phases: a weight reduction period with formula diet (12 weeks) and a weight maintenance phase (40 weeks). Body weight, body mass index (BMI), blood pressure and concentrations of blood glucose, total cholesterol, low-density lipoprotein, high-density lipoprotein and triglycerides were determined in week 0 (T(0)), after 12 weeks (T(1)) and at the end in week 52 (T(2)). SUBJECTS: A total of 193 obese subjects aged between 18 and 72 years (129 female, 64 male; initial body weight: 122.4±22.3 kg, initial BMI: 41.8±6.7 kg m(-2)) were included. RESULTS: Genotyping revealed 32.1% TT-, 39.4% AT- and 28.5% AA-genotype carriers. At T (0), carriers of the AA-genotype had significantly higher body weight (P=0.04) and BMI (P=0.005) than carriers of the TT-genotype. Of the 193 participants, 68 discontinued and 125 completed the program. Dropout rate was not influenced by genotype (P=0.33). Completers with AA-genotype showed significantly lower additional weight loss during the weight maintenance phase than TT-genotype carriers (P=0.02). Furthermore, among participants facing weight regain during weight maintenance (n=52), more subjects were carrying the AA-genotype (P=0.006). No influence of genotype on weight reduction under formula diet was observed (P=0.32). CONCLUSION: In this program, the AA-genotype of rs9939609 was associated with a higher initial body weight and did influence success of weight stabilization. Thus, emphasizing the maintenance phase during a weight reduction program might result in better success for AA-genotype carriers.
Asunto(s)
Peso Corporal/genética , Obesidad/genética , Proteínas/genética , Aumento de Peso/genética , Pérdida de Peso/genética , Adolescente , Adulto , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato , Glucemia/metabolismo , Índice de Masa Corporal , Estudios Transversales , Femenino , Predisposición Genética a la Enfermedad , Variación Genética , Genotipo , Alemania/epidemiología , Humanos , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Obesidad/sangre , Obesidad/epidemiología , Proyectos Piloto , Triglicéridos/sangreRESUMEN
This prospective study was performed in order to establish whether improvement of endothelial function after weight reduction can be explained by a decrease of elevated asymmetric dimethyl arginine (ADMA), an inhibitor of endogenous NO-synthase (eNOS). Therefore, 21 obese subjects (BMI: 41.1±6.4 kg/m(2)) were studied at baseline and after 12 weeks of weight reduction with a very low calorie diet. Biochemical and clinical parameters of endothelial function were assessed before and after weight loss. Biochemical parameters were determined by measurement of ADMA and soluble intercellular adhesion molecule (sICAM). Clinical parameters were assessed by pulse wave analysis (PWA). Weight intervention resulted in a 21.4±6.8 kg reduction of body weight from 119.7±12.8 kg at study start to 98.3±11.6 kg at study end (p<0.001). Accordingly, biochemical markers improved under weight reduction (ADMA from 0.47±0.07 mmol/l to 0.42±0.08 mmol/l; p=0.002; ICAM from 276±42 ng/ml to 236±29 ng/ml; p<0.001). Further, clinical parameters of functional endothelial function improved with an increase of deltaRI after salbutamol inhalation from -1% before to -9% after weight reduction (p=0.02). Interestingly, improvement of endothelial function correlated with improved HOMA index only (r=-0.60, p=0.04) but not with reduced ADMA levels, improved hypertension or reduced body weight. In conclusion, weight reduction with a very low calorie diet improves endothelial function measured by pulse wave velocity. The missing correlation with ADMA suggests possible further mechanisms underlying this observed effect, for example, improvement of insulin resistance.
Asunto(s)
Arginina/análogos & derivados , Células Endoteliales/fisiología , Obesidad/metabolismo , Pérdida de Peso , Adolescente , Adulto , Anciano , Arginina/metabolismo , Índice de Masa Corporal , Restricción Calórica/estadística & datos numéricos , Moléculas de Adhesión Celular/metabolismo , Células Endoteliales/efectos de los fármacos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Obesidad/dietoterapia , Obesidad/fisiopatología , Adulto JovenRESUMEN
AIMS/HYPOTHESIS: Extracellular matrix reorganisation is a crucial step of adipocyte differentiation and is controlled by the matrix metalloproteinase-tissue inhibitor of matrix metalloproteinase (TIMP) enzyme system. We therefore sought to define the role of TIMP1 in adipogenesis and to elucidate whether upregulation of TIMP1 in obesity has direct effects on adipocyte formation. METHODS: TIMP1 protein levels and mRNA were measured in lean and obese mice with a focus on levels in adipose tissue. We also analysed the effect of recombinant murine TIMP1 on adipogenesis, adipocyte size and metabolic control in vitro and in vivo. RESULTS: TIMP1 levels were increased in the serum and adipose tissue of obese mouse models. Recombinant murine TIMP1 inhibited adipocyte differentiation in 3T3-L1 as well as in subcutaneous primary pre-adipocytes. Conversely, neutralising TIMP1 with a specific antibody enhanced adipocyte differentiation. In vivo, injection of recombinant TIMP1 in mice challenged with a high-fat diet led to enlarged adipocytes. TIMP1-treated mice developed an impaired metabolic profile with increased circulating NEFA levels, hepatic triacylglycerol accumulation and accelerated insulin resistance. Altered glucose clearance in TIMP1-injected mice was due to changes in adipose tissue glucose uptake, whereas muscle glucose clearance remained unaffected. CONCLUSIONS/INTERPRETATION: TIMP1 is a negative regulator of adipogenesis. In vivo, TIMP1 leads to enlarged adipocytes in the state of overnutrition. This might contribute to the detrimental metabolic consequences seen in TIMP1-injected mice, such as systemic fatty acid overload, hepatic lipid accumulation and insulin resistance.
Asunto(s)
Adipogénesis/fisiología , Obesidad/metabolismo , Obesidad/patología , Inhibidor Tisular de Metaloproteinasa-1/fisiología , Adipocitos/efectos de los fármacos , Adipocitos/patología , Animales , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Grasas de la Dieta/farmacología , Modelos Animales de Enfermedad , Ácidos Grasos no Esterificados/metabolismo , Humanos , Resistencia a la Insulina/fisiología , Metabolismo de los Lípidos/efectos de los fármacos , Metabolismo de los Lípidos/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/fisiopatología , Inhibidor Tisular de Metaloproteinasa-1/farmacología , Triglicéridos/metabolismoRESUMEN
Acute inflammation in humans is associated with transient insulin resistance (IR) and dyslipidemia. Chronic low-grade inflammation is a pathogenic component of IR and adipose tissue dysfunction in obesity-induced type 2 diabetes. Because feline diabetes closely resembles human type 2 diabetes, we studied whether lipopolysaccharide (LPS)-induced subacute inflammation, in the absence of obesity, is the potential primary cause of IR and metabolic disorders. Cats received increasing iv doses (10-1000 ng/kg(-1) · h(-1)) of LPS (n = 5) or saline (n = 5) for 10 d. Body temperature, proinflammatory and metabolic markers, and insulin sensitivity were measured daily. Tissue mRNA and protein expression were quantified on d 10. LPS infusion increased circulating and tissue markers of inflammation. Based on the homeostasis model assessment, endotoxemia induced transient IR and ß-cell dysfunction. At the whole-body level, IR reverted after the 10-d treatment; however, tissue-specific indications of IR were observed, such as down-regulation of adipose glucose transporter 4, hepatic peroxisome proliferative activated receptor-γ1 and -2, and muscle insulin receptor substrate-1. In adipose tissue, increased hormone-sensitive lipase activity led to reduced adipocyte size, concomitant with increased plasma and hepatic triglyceride content and decreased total and high-density lipoprotein cholesterol levels. Prolonged LPS-induced inflammation caused acute IR, followed by long-lasting tissue-specific dysfunctions of lipid-, glucose-, and insulin metabolism-related targets; this ultimately resulted in dyslipidemia but not whole-body IR. Endotoxemia in cats may provide a promising model to study the cross talk between metabolic and inflammatory responses in the development of adipose tissue dysfunction and IR.
Asunto(s)
Tejido Adiposo/fisiopatología , Endotoxemia/metabolismo , Inflamación/inducido químicamente , Metabolismo de los Lípidos/fisiología , Lipoproteínas/metabolismo , Animales , Gatos , Endotoxemia/inducido químicamente , Regulación de la Expresión Génica , Resistencia a la Insulina , Células Secretoras de Insulina , Lipopolisacáridos , MasculinoRESUMEN
Translation of nutrient stimuli through intracellular signaling is important for adaptation and regulation of metabolic processes, while deregulation by either genetic or environmental factors predisposes towards the development of metabolic disorders. Besides providing energy, fatty acids act as prominent signaling molecules by altering cell membrane structures, affecting the lipid modification status of proteins, and by modulating ligand-activated nuclear receptor activity. Given their highly hydrophobic nature, fatty acids in the aqueous intracellular compartment are bound to small intracellular lipid binding proteins which function as intracellular carriers of these hydrophobic components. This review describes recent advances in identifying intracellular pathways for cytosolic fatty acid signaling through ligand activated receptors by means of small intracellular lipid binding proteins. The mechanism behind intracellular fatty acid transport and subsequent nuclear receptor activation is an emerging concept, and advances in understanding this process provide new potential therapeutic targets towards the treatment of metabolic disorders.
Asunto(s)
Proteínas de Unión a Ácidos Grasos/metabolismo , Ácidos Grasos/metabolismo , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Animales , Humanos , Metabolismo , Receptores Activados del Proliferador del Peroxisoma/fisiología , Transducción de SeñalRESUMEN
Due to the growing numbers of counterfeited products on the world market, there is a huge demand for new and forgery-proof marking systems. We developed a unique system using "molecular beacons" with well adapted thermodynamic parameters. This marking system consists of the three components: DNA tag (a label or directly printing), detection pen (contains the "molecular beacon " solution), and DNA-scanner (reads the specific signal triggered by the detection pen even at daylight). The vast coding capacity of DNA combined with the highly specific signal offers a degree of security that is unmatched by conventional identification technologies.
Asunto(s)
ADN/química , Fraude/prevención & control , Biología Molecular/métodos , Oligonucleótidos/química , Técnicas Biosensibles , Colorantes Fluorescentes/farmacología , Cinética , Desnaturalización de Ácido Nucleico , Hibridación de Ácido Nucleico , Sondas de Oligonucleótidos/química , Temperatura , Termodinámica , Factores de TiempoRESUMEN
Peroxisome proliferator-activated receptor alpha (PPARalpha) is a key regulator of lipid homeostasis in hepatocytes and target for fatty acids and hypolipidemic drugs. How these signaling molecules reach the nuclear receptor is not known; however, similarities in ligand specificity suggest the liver fatty acid binding protein (L-FABP) as a possible candidate. In localization studies using laser-scanning microscopy, we show that L-FABP and PPARalpha colocalize in the nucleus of mouse primary hepatocytes. Furthermore, we demonstrate by pull-down assay and immunocoprecipitation that L-FABP interacts directly with PPARalpha. In a cell biological approach with the aid of a mammalian two-hybrid system, we provide evidence that L-FABP interacts with PPARalpha and PPARgamma but not with PPARbeta and retinoid X receptor-alpha by protein-protein contacts. In addition, we demonstrate that the observed interaction of both proteins is independent of ligand binding. Final and quantitative proof for L-FABP mediation was obtained in transactivation assays upon incubation of transiently and stably transfected HepG2 cells with saturated, monounsaturated, and polyunsaturated fatty acids as well as with hypolipidemic drugs. With all ligands applied, we observed strict correlation of PPARalpha and PPARgamma transactivation with intracellular concentrations of L-FABP. This correlation constitutes a nucleus-directed signaling by fatty acids and hypolipidemic drugs where L-FABP acts as a cytosolic gateway for these PPARalpha and PPARgamma agonists. Thus, L-FABP and the respective PPARs could serve as targets for nutrients and drugs to affect expression of PPAR-sensitive genes.
Asunto(s)
Proteínas Portadoras/fisiología , Núcleo Celular/metabolismo , Ácidos Grasos/fisiología , Regulación de la Expresión Génica , Hipolipemiantes/farmacología , Proteínas de Neoplasias , Proteínas del Tejido Nervioso , Receptores Citoplasmáticos y Nucleares/genética , Factores de Transcripción/genética , Proteínas Supresoras de Tumor , Animales , Línea Celular , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos , Humanos , Hígado/metabolismo , Ratones , Unión Proteica , Transducción de Señal , Activación Transcripcional/efectos de los fármacos , Activación Transcripcional/fisiología , Técnicas del Sistema de Dos HíbridosRESUMEN
Liver-type fatty acid binding protein (L-FABP) has been proposed to be involved in the transport of fatty acids and peroxisome proliferators from the cytosol into the nucleus for interaction with the peroxisome proliferator-activated receptors (PPARs). On the basis of this premise, we investigated by isothermal titration calorimetry the binding of myristic, stearic, oleic, and docosahexaenoic acids to three orthologous L-FABPs and compared these results to those obtained for several xenobiotics [Wy14,643, bezafibrate, 5,8,11,14-eicosatetraynoic acid (ETYA), and BRL48,482] known for their peroxisome proliferating activity in rodents. Recombinant human, murine, and bovine L-FABPs were analyzed and the thermodynamic data were obtained. Our studies showed that fatty acids bound with a stoichiometry of 2:1, fatty acid to protein, with dissociation constants for the first binding site in the nanomolar range. With dissociation constants above 1 microM the drug peroxisome proliferators showed weaker binding, with the exception of arachidonate analogue ETYA, which bound with a similar affinity as the natural fatty acid. Some of the thermodynamic data obtained for fatty acid binding could be explained by differences in protein structure. Moreover, our results revealed that binding affinities were not determined by ligand solubility in the aqueous phase.
Asunto(s)
Proteínas Portadoras/metabolismo , Ácidos Grasos/metabolismo , Hígado/metabolismo , Proteína P2 de Mielina/metabolismo , Proteínas de Neoplasias , Proteínas del Tejido Nervioso , Proliferadores de Peroxisomas/metabolismo , Proteínas Supresoras de Tumor , Ácido 5,8,11,14-Eicosatetrainoico/metabolismo , Secuencia de Aminoácidos , Animales , Bezafibrato/química , Bezafibrato/metabolismo , Calorimetría , Proteínas Portadoras/química , Bovinos , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos , Ácidos Grasos/química , Humanos , Ligandos , Ratones , Datos de Secuencia Molecular , Proteína P2 de Mielina/química , Proliferadores de Peroxisomas/química , Unión Proteica , Pirimidinas/química , Pirimidinas/metabolismoRESUMEN
The aim of the present study was to investigate whether lipid metabolism in the nuclei is affected by changes in the metabolism of free fatty acids in the liver. The experiments were carried out on 3 groups of rats: 1 - control-male, 2 - female, and 3 - male, treated with bezafibrate (a peroxisome proliferator). The rats received 14C-palmitic acid intravenously. Thirty min later liver samples and blood from the abdominal aorta were taken. The liver nuclei were isolated in sucrose gradient. Lipids were extracted from the nuclei and the liver homogenate and subsequently separated into the following fractions: phospholipids, mono, di- and triacylglycerols, free fatty acids, cholesterol and cholesterol esters. The radioactivity of each fraction was counted. Furthermore, the content of free fatty acids and the fatty acid binding proteins was measured. It was found that radioactivity was present in each lipid fraction obtained from the liver homogenate and from the nuclei. In the female group, the total radioactivity of lipids in the liver homogenate was lower, whereas in the nuclei it was higher in comparison to the male group. The reduction in the radioactivity in the liver was mostly accounted for by decreased radioactivity in the fraction oftriacylglycerols and phospholipids. In the nuclei, the radioactivity of the fraction of phospholipids, free fatty acids and diacylglycerols was elevated. Bezafibrate did not affect the total radioactivity of lipids in the liver and reduced it in the nuclei. In the liver, the drug increased radioactivity mostly in the fraction of phospholipids and reduced it mainly in the fraction of triacylglycerols. In the nuclei, the radioactivity of each lipid fraction examined was reduced. The content of the fraction of free fatty acids in the liver and in the nuclei in the female and in the bezafibrate-treated groups did not differ from the respective value in the control group. The content of fatty acid binding proteins in the nuclei of the female and bezafibrate-treated groups increased in parallel to the elevation in their content in the cytosol. It is concluded that the female sex hormones and bezafibrate influence the transport of selected lipids into the nuclei. The effects seem to be a consequence of the action of these factors directly on the nucleus.
Asunto(s)
Bezafibrato/farmacología , Metabolismo de los Lípidos , Hígado/efectos de los fármacos , Hígado/metabolismo , Proteínas de Neoplasias , Proteínas del Tejido Nervioso , Ácido Palmítico/metabolismo , Transporte Activo de Núcleo Celular/efectos de los fármacos , Animales , Radioisótopos de Carbono , Proteínas Portadoras/metabolismo , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos , Ácidos Grasos no Esterificados/metabolismo , Femenino , Masculino , Ácido Palmítico/sangre , Proliferadores de Peroxisomas/farmacología , Ratas , Ratas Wistar , Caracteres SexualesRESUMEN
Branched-chain phytanic acid is metabolized in liver peroxisomes. Sterol carrier protein 2/sterol carrier protein x (SCP2/SCPx) knockout mice, which develop a phenotype with a deficiency in phytanic acid degradation, accumulate dramatically high concentrations of this fatty acid in serum (Seedorf at al. 1998. Genes Dev. 12: 1189-1201) and liver. Concomitantly, a 6.9-fold induction of liver fatty acid binding protein (L-FABP) expression is observed in comparison to wild-type animals fed standard chow, possibly mediated by the peroxisome proliferator-activated receptor alpha (PPARalpha). Cytosolic transport of phytanic acid to either peroxisomal membranes or to the nucleus for activation of PPARalpha may be mediated by L-FABP, which gives rise to the question whether phytanic acid is a transactivator of this protein. Here we show first that phytanic acid binds to recombinant L-FABP with high affinity. Then the increase of the in vivo phytanic acid concentration by phytol feeding to mice results in a 4-fold induction of L-FABP expression in liver, which is in the order of that attained with bezafibrate, a known peroxisome proliferator. Finally to test in vitro whether this induction is conferred by phytanic acid, we cotransfected HepG2 cells with an expression plasmid for murine PPARalpha and a CAT-reporter gene with 176 bp of the murine L-FABP promoter, containing the peroxisome proliferator responsive element (PPRE). After incubation with phytanic acid, we observed a 3.2-fold induction of CAT expression. These findings, both in vivo and in vitro, demonstrate that phytanic acid is a transcriptional activator of L-FABP expression and that this effect is mediated via PPARalpha.
Asunto(s)
Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Proteína P2 de Mielina/genética , Proteína P2 de Mielina/metabolismo , Proteínas de Neoplasias , Proteínas del Tejido Nervioso , Ácido Fitánico/metabolismo , Ácido Fitánico/farmacología , Transcripción Genética/efectos de los fármacos , Animales , Bezafibrato/farmacología , Línea Celular , Cloranfenicol O-Acetiltransferasa/genética , ADN Complementario , Escherichia coli/genética , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos , Expresión Génica/efectos de los fármacos , Hipolipemiantes/farmacología , Hígado/metabolismo , Ratones , Fitol/farmacología , Ratas , Receptores Citoplasmáticos y Nucleares/genética , Proteínas Recombinantes/metabolismo , Factores de Transcripción/genética , Activación Transcripcional , TransfecciónRESUMEN
The liver-type fatty acid binding protein (L-FABP), a member of a family of mostly cytosolic 14-15 kDa proteins known to bind fatty acids in vitro and in vivo, is discussed to play a role in fatty acid uptake. Cells of the hepatoma HepG2 cell line endogenously express this protein to approximately 0.2% of cytosolic proteins and served as a model to study the effect of L-FABP on fatty acid uptake, by manipulating L-FABP expression in two approaches. First, L-FABP content was more than doubled upon treating the cells with the potent peroxisome proliferators bezafibrate and Wy14,643 and incubation of these cells with [1-14C]oleic acid led to an increase in fatty acid uptake rate from 0.55 to 0.74 and 0.98 nmol/min per mg protein, respectively. In the second approach L-FABP expression was reduced by stable transfection with antisense L-FABP mRNA yielding seven clones with L-FABP contents ranging from 0.03% to 0.14% of cytosolic proteins. This reduction to one sixth of normal L-FABP content reduced the rate of [1-14C]oleic acid uptake from 0.55 to 0. 19 nmol/min per mg protein, i.e., by 66%. The analysis of peroxisome proliferator-treated cells and L-FABP mRNA antisense clones revealed a direct correlation between L-FABP content and fatty acid uptake.
Asunto(s)
Proteínas Portadoras/biosíntesis , Ácidos Grasos/metabolismo , Proteína P2 de Mielina/biosíntesis , Proteínas de Neoplasias , Proliferadores de Peroxisomas/farmacología , Proteínas Supresoras de Tumor , Bezafibrato/farmacología , Carcinoma Hepatocelular , Proteínas Portadoras/genética , Proteína de Unión a los Ácidos Grasos 7 , Proteínas de Unión a Ácidos Grasos , Humanos , Proteína P2 de Mielina/genética , Pirimidinas/farmacología , ARN sin Sentido , Transfección , Células Tumorales CultivadasRESUMEN
We showed recently that a targeted null mutation in the murine sterol carrier protein 2-/sterol carrier protein x-gene (Scp2) leads to defective peroxisomal catabolism of 3,7,11, 15-tetramethylhexadecanoic acid (phytanic acid), peroxisome proliferation, hypolipidemia, and enhanced hepatic expression of several genes that have been demonstrated to be transcriptionally regulated by the peroxisome proliferator-activated receptor alpha (PPARalpha). As a broad range of fatty acids activates PPARalpha in vitro, we examined whether the latter effects could be because of phytanic acid-induced activation of this transcription factor. Dietary phytol supplementation was used to modulate the concentration of phytanic acid in C57Bl/6 and Scp2 (-/-) mice. We found that the serum concentrations of phytanic acid correlated well with the expression of genes encoding peroxisomal beta-oxidation enzymes and liver fatty acid-binding protein, which have all been demonstrated to contain functionally active peroxisome proliferator response elements in their promoter regions. In accordance with these findings, a stimulating effect on acyl-CoA oxidase gene expression was also observed after incubation of the rat hepatoma cell line MH1C1 with phytanic acid. Moreover, reporter gene studies revealed that phytanic acid induces the expression of a peroxisome proliferator response element-driven chloramphenicol transferase reporter gene comparable with strong peroxisome proliferators. In addition, the ability of phytanic acid to act as an inductor of PPARalpha-dependent gene expression corresponded with high affinity binding of this dietary branched chain fatty acid to recombinant PPARalpha. We conclude that phytanic acid can be considered as a bona fide physiological ligand of murine PPARalpha.
Asunto(s)
Proteínas Portadoras/fisiología , Proteínas de Unión al ADN/metabolismo , Ácido Fitánico/farmacología , Proteínas de Plantas , Receptores Citoplasmáticos y Nucleares/metabolismo , Factores de Transcripción/metabolismo , Acil-CoA Oxidasa , Animales , Proteínas Portadoras/genética , Ligandos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidorreductasas/genética , Oxidorreductasas/metabolismo , Trastorno Peroxisomal/genética , Trastorno Peroxisomal/metabolismo , ARN Mensajero/metabolismo , Ratas , Células Tumorales CultivadasRESUMEN
Moclobemide was compared to placebo in two parallel groups of depressed patients, in a multicenter randomized, double-blind study of six weeks treatment duration. Forty seven patients participated in the study: 23 received moclobemide (flexible dose 300-600 mg/day) and 24 placebo. They were evaluated weekly for efficacy and tolerability. Moclobemide was more efficacious than placebo as judged by analysis on the total score on the Hamilton depression scale (p < 0.05) and by the overall assessment of efficacy (p < 0.01). Moclobemide was also more effective than placebo in the subgroup with neurotic depression (p < 0.05). In addition, the number of patients prematurely terminating treatment for inefficacy, was higher in the placebo than in the moclobemide group (12 versus 2, p < 0.01). The number and the severity of side-effects tended to be slightly greater in the moclobemide than in the placebo group, but this did not reach a level of significance. Cardiovascular tolerability was good in both treatment groups. No hypertensive crisis was reported. Hematology, clinical chemistry and urine analysis were not affected by the treatment in any clinically significant fashion.