Abstract: OBJECTIVE: We have recently shown the in vivo anti-obesity effects of sodium tungstate. In this study, we investigate the in vitro effects of sodium tungstate on adipocyte differentiation and function. METHODS: 3T3-F442A cells were allowed to differentiate in the presence of sodium tungstate, and were analyzed for triglyceride (TG) accumulation, adipocyte differentiation and mitochondrial oxygen consumption. RESULTS: Sodium tungstate treatment of adipose cells decreased TG accumulation and adipocyte differentiation. Expression of key genes for adipocyte function (aP2, ACC, fatty acid synthase (FAS) and lipoprotein lipase (LPL)) and differentiation (CCAAT enhancer-binding protein (C/EBP)alpha and peroxisome proliferator-activated receptor gamma (PPARgamma)) was reduced by sodium tungstate, whereas C/EBPbeta isoform LIP expression level was increased. TG accumulation and changes in C/EBPbeta expression were partially recovered by inactivating the erk1/2 pathway. Finally, tungstate treatment increased the oxygen consumption of adipose cells without changes in the expression of oxidative genes. CONCLUSIONS: Sodium tungstate inhibits adipocyte differentiation by promoting the translation of LIP, a master dominant-negative regulator of this process, and regulates the mitochondrial oxygen consumption of adipose cells. These effects contribute to the anti-obesity activity of sodium tungstate and confirm its potential as a powerful alternative for the treatment of obesity.

Abstract: OBJECTIVE: The aim of this study is to determine the effect of coenzyme Q (Q) on ob/ob mice treated or not with thiazolidinedione (TZD). DESIGN AND MEASUREMENTS: Ob/ob mice were treated with Q, Rosiglitazone or a combination of both molecules for 13 days; physical and metabolic parameters as well as oral glucose tolerance test were assessed. mRNA expression of genes of energy dissipation and storage were measured by real-time PCR. RESULTS: Q treatment improved some metabolic parameters in ob/ob mice. Surprisingly, cotreatment with Rosiglitazone and Q improved metabolic parameters and prevented TZD increase in body weight and adiposity, mainly by increasing lipid oxidation in adipose tissue, reducing lipid synthesis and balancing adipokine gene expression. CONCLUSIONS: Our finding suggests that Rosiglitazone and coenzyme Q bitherapy could prevent the body weight gain associated with adipogenesis and could improve the clinical use of these compounds.

Abstract: Sodium tungstate is a novel agent in the treatment of obesity. In diet-induced obese rats, it is able to reduce body weight gain by increasing energy expenditure. This study evaluated the role of leptin, a key regulator of energy homeostasis, in the tungstate antiobesity effect. Leptin receptor-deficient Zucker fa/fa rats and leptin-deficient ob/ob mice were treated with tungstate. In lean animals, tungstate administration reduced body weight gain and food intake and increased energy expenditure. However, in animals with deficiencies in the leptin system, treatment did not modify these parameters. In ob/ob mice in which leptin deficiency was restored through adipose tissue transplantation, treatment restored the tungstate-induced body weight gain and food intake reduction as well as energy expenditure increase. Furthermore, in animals in which tungstate administration increased energy expenditure, changes in the expression of key genes involved in brown adipose tissue thermogenesis were detected. Finally, the gene expression of the hypothalamic neuropeptides, Npy, Agrp, and Cart, involved in the leptin regulation of energy homeostasis, was also modified by tungstate in a leptin-dependent manner. In summary, the results indicate that the effectiveness of tungstate in reducing body weight gain is completely dependent on a functional leptin system.

Abstract: The status of adipose tissue changes rapidly. From a simple filler tissue, it successively acquires the status of metabolic active tissue, endocrine tissue, plastic tissue, and finally that of a large reservoir of cells suitable for cell therapy and regenerative medicine. All throughout this story, our knowledge has been largely dependent on genetic tools and gene transfer. Now, the time has come where gene transfer in adipose derived cells can be envisioned, not only for understanding the role or importance of one gene, but also to engineer adipose derived cells for the purpose of therapy by delivering secreted products. In this paper, after a brief overview of adipose tissues, a large part will be devoted to the use of virus-based gene transfer in transducing adipose tissue and cells which reside therein. We also critically review the use of adipose "specific" promoters and the applications already described in the literature.

Abstract: Our previous results demonstrated that tungstate decreased weight gain and adiposity in obese rats through increased thermogenesis and lipid oxidation, suggesting that brown adipose tissue was one of the targets of its antiobesity effect. To identify potential targets of tungstate, we used DIGE to compare brown adipose tissue protein extracts from the following experimental groups: untreated lean, tungstate-treated lean, untreated obese, and tungstate-treated obese rats. To distinguish direct targets of tungstate action from those that are secondary to body weight loss, we also included in the analysis an additional group consisting of obese rats that lose weight by caloric restriction. Hierarchical clustering of analysis of variance and t test contrasts clearly separated the different experimental groups. DIGE analysis identified 20 proteins as tungstate obesity direct targets involved in Krebs cycle, glycolysis, lipolysis and fatty acid oxidation, electron transport, and redox. Protein oxidation was decreased by tungstate treatment, confirming a role in redox processes; however, palmitate oxidation, as a measure of fatty acid beta-oxidation, was not altered by tungstate, thus questioning its putative function in fatty acid oxidation. Protein network analyses using Ingenuity Pathways Analysis highlighted peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC-1alpha) as a potential target. We confirmed by real time PCR that indeed tungstate up-regulates PGC-1alpha, and its major target, uncoupling protein 1, was also increased as shown by Western blot. These results illustrate the utility of proteomics and bioinformatics approaches to identify targets of obesity therapies and suggest that in brown adipose tissue tungstate modulates redox processes and increases energy dissipation through uncoupling and PGC-1alpha up-regulation, thus contributing to its overall antiobesity effect.

Abstract: Brown adipose tissue uncoupling protein-1 (UCP1) plays a major role in the control of energy balance in rodents. It has long been thought, however, that there is no physiologically relevant UCP1 expression in adult humans. In this study we show, using an original approach consisting of sorting cells from various tissues and differentiating them in an adipogenic medium, that a stationary population of skeletal muscle cells expressing the CD34 surface protein can differentiate in vitro into genuine brown adipocytes with a high level of UCP1 expression and uncoupled respiration. These cells can be expanded in culture, and their UCP1 mRNA expression is strongly increased by cell-permeating cAMP derivatives and a peroxisome-proliferator-activated receptor-gamma (PPARgamma) agonist. Furthermore, UCP1 mRNA was detected in the skeletal muscle of adult humans, and its expression was increased in vivo by PPARgamma agonist treatment. All the studies concerning UCP1 expression in adult humans have until now been focused on the white adipose tissue. Here we show for the first time the existence in human skeletal muscle and the prospective isolation of progenitor cells with a high potential for UCP1 expression. The discovery of this reservoir generates a new hope of treating obesity by acting on energy dissipation.

Abstract: The ability for the brain to sense peripheral fuel availability is mainly accomplished within the hypothalamus, which detects ongoing systemic nutrients and adjusts food intake and peripheral metabolism as needed. Here, we hypothesized that mitochondrial reactive oxygen species (ROS) could trigger sensing of nutrients within the hypothalamus. For this purpose, we induced acute hypertriglyceridemia in rats and examined the function of mitochondria in the hypothalamus. Hypertriglyceridemia led to a rapid increase in the mitochondrial respiration in the ventral hypothalamus together with a transient production of ROS. Cerebral inhibition of fatty acids-CoA mitochondrial uptake prevented the hypertriglyceridemia-stimulated ROS production, indicating that ROS derived from mitochondrial metabolism. The hypertriglyceridemia-stimulated ROS production was associated with change in the intracellular redox state without any noxious cytotoxic effects, suggesting that ROS function acutely as signaling molecules. Moreover, cerebral inhibition of hypertriglyceridemia-stimulated ROS production fully abolished the satiety related to the hypertriglyceridemia, suggesting that hypothalamic ROS production was required to restrain food intake during hypertriglyceridemia. Finally, we found that fasting disrupted the hypertriglyceridemia-stimulated ROS production, indicating that the redox mechanism of brain nutrient sensing could be modulated under physiological conditions. Altogether, these findings support the role of mitochondrial ROS as molecular actors implied in brain nutrient sensing.

Abstract: Due to the importance of fat tissues in both energy balance and in the associated disorders arising when such balance is not maintained, adipocyte differentiation has been extensively investigated in order to control and inhibit the enlargement of white adipose tissue. The ability of a cell to undergo adipocyte differentiation is one particular feature of all mesenchymal cells. Up until now, the peroxysome proliferator-activated receptor (PPAR) subtypes appear to be the keys and essential players capable of inducing and controlling adipocyte differentiation. In addition, it is now accepted that adipose cells present a broad plasticity that allows them to differentiate towards various mesodermal phenotypes. The role of PPARs in such plasticity is reviewed here, although no definite conclusion can yet be drawn. Many questions thus remain open concerning the definition of preadipocytes and the relative importance of PPARs in comparison to other master factors involved in the other mesodermal phenotypes.

Abstract: OBJECTIVE: Rosiglitazone displays powerful antidiabetes benefits but is associated with increased body weight and adipogenesis. Keeping in mind the concept of selective peroxisome proliferator-activated receptor (PPAR)gamma modulator, the aim of this study was to characterize the properties of a new PPARgamma ligand, S 26948, with special attention in body-weight gain. RESEARCH DESIGN AND METHODS: We used transient transfection and binding assays to characterized the binding characteristics of S 26948 and GST pull-down experiments to investigate its pattern of coactivator recruitment compared with rosiglitazone. We also assessed its adipogenic capacity in vitro using the 3T3-F442A cell line and its in vivo effects in ob/ob mice (for antidiabetes and antiobesity properties), as well as the homozygous human apolipoprotein E2 knocking mice (E2-KI) (for antiatherogenic capacity). RESULTS: S 26948 displayed pharmacological features of a high selective ligand for PPARgamma with low potency in promoting adipocyte differentiation. It also displayed a different coactivator recruitment profile compared with rosiglitazone, being unable to recruit DRIP205 or PPARgamma coactivator-1 alpha. In vivo experiments showed that S 26948 was as efficient in ameliorating glucose and lipid homeostasis as rosiglitazone, but it did not increase body and white adipose tissue weights and improved lipid oxidation in liver. In addition, S 26948 represented one of the few molecules of the PPARgamma ligand class able to decrease atherosclerotic lesions. CONCLUSIONS: These findings establish S 26948 as a selective PPARgamma ligand with distinctive coactivator recruitment and gene expression profile, reduced adipogenic effect, and improved biological responses in vivo.

Abstract: Literature on reactive oxygen species (ROS) effects on cell biology and physiopathology is huge and appears to be controversial. This could be explained by the fact that very few studies take into account the real subcellular source of ROS production, their chemical nature and the intensity of their production. In spite of the importance of the other sites of ROS production in the cell, we decided to focus on mitochondrial ROS. Besides their key role in bioenergetics and ATP synthesis, mitochondria are one of the main sites of ROS generation within the cell. 80 % of intracellular superoxide anion is provided by the mitochondrial respiratory chain. Mitochondrial ROS production is closely associated with activity of the respiratory chain and is modulated by environmental factors which can induce constraints on respiratory chain components. Nutrient availability as well as oxygen pressure can both modulate mitochondrial ROS production. When moderately produced, ROS specifically regulate intracellular signalling pathways by reversible oxidation of proteins such as transcription factors or proteins kinases. In this way, they can trigger cell adaptation to environmental changes as modifications of energetic metabolism or hypoxia. Indeed, we demonstrated that mitochondrial ROS act as key elements in the control of white adipose tissue development by specific up-regulation of the anti-adipogenic transcription factor CHOP-10/GADD153. However, when they are produced at high level and in a chronic manner, mitochondrial ROS can also have deleterious effects by massive and irreversible oxidation of their principals targets i.e. lipids, DNA and proteins. In these conditions, mitochondrial ROS are involved in aging process and in pathological situations as metabolic disease.

Abstract: A series of 1,3-dicarbonyl compounds having 2(3H)-benzazolonic heterocycles has been synthesized and tested for PPARgamma agonist activity. SAR were developed and revealed that 6-acyl-2(3H)-benzothiazolone derivatives with 1,3-dicarbonyl group were the most potent. IP administration of compound 22 exhibited comparable levels of glucose and triglyceride correction to PO administration of rosiglitazone in the ob/ob mouse studies.

Abstract: Using pharmacological tools, a role for opioid receptors in the regulation of food intake has been documented. However, the involvement of specific receptor subtypes remains questionable, and little information is available regarding a role for opioid receptors in energy metabolism. Using adult male mice lacking the mu-opioid receptor (MOR) gene (MOR-/-), we show that the MOR is not essential for the maintenance of normal levels of ad libitum food intake but does modulate the efficiency of energy storage during high-fat diets through the regulation of energy partitioning. When fed a regular diet, MOR-/- mice displayed only subtle alterations in energy homeostasis, suggesting a relative overuse of fat as a fuel source in the fed state. When fed a high-fat diet, MOR-/- mice were resistant to obesity and impaired glucose tolerance, despite having similar energy intake to wild-type mice. This resistance to obesity was associated with a strong induction of the expression of key mitochondrial enzymes involved in fatty acid oxidation within skeletal muscle. This metabolic role of the MOR, which is consistent with the properties of a "thrifty gene," suggests that the MOR pathway is a potential target for pharmacological intervention in the treatment of obesity associated with the intake of fatty diets.

Abstract: AIMS/HYPOTHESIS: Fibrates and thiazolidinediones are commonly used for the treatment of dyslipidemia and type 2 diabetes, respectively. The aim of this study was to investigate the effects on body weight as well as on glucose and lipid homeostasis of ligands for PPARalpha and PPARgamma, Fenofibrate and Rosiglitazone, alone or in association. METHODS: Ob/ob mice were divided into four groups: control, and mice daily injected (intraperitoneally), either with 10 mg/kg Rosiglitazone, 100 mg/kg Fenofibrate or both molecules. Body weight and food intake were monitored daily. After 13 days of treatment, mice were killed, and blood samples were collected for posterior metabolite quantification. The liver and adipose tissues were dissected and weighed. RESULTS: Body weight was significantly reduced or increased by Fenofibrate and Rosiglitazone, respectively. The effect of Rosiglitazone was prevented by coadministration of Fenofibrate. This was accompanied by a normalization of the daily food efficiency. Compared to those treated with Rosiglitazone, animals treated with Fenofibrate alone or in combination presented a decreased white adipose tissue mass. Fenofibrate or Rosiglitazone alone significantly reduced the levels of plasma lipid parameters. Surprisingly, Fenofibrate also decreased blood glucose levels in ob/ob mice, despite having no effect on insulin levels. By contrast, both glucose and insulin levels were decreased by Rosiglitazone treatment. Coadministration of both drugs improved all parameters as with Rosiglitazone. Fenofibrate restored almost normal hepatocyte morphology and significantly reduced the triglyceride content of the liver. This was accompanied by an increase in fatty acid oxidation in the liver in all groups receiving Fenofibrate. CONCLUSION/INTERPRETATION: These biological effects suggest that combined therapy with a PPARalpha and a PPARgamma ligand is more effective in ameliorating, specifically, lipid homeostasis than in activating any of this receptor separately. Furthermore, Fenofibrate prevents one of the most undesirable effects of Rosiglitazone, namely increased adiposity and body weight gain.

Abstract: C/EBPbeta (CCAAT/enhancer-binding protein beta) is a transcriptional regulator of the UCP1 (uncoupling protein-1) gene, the specific marker gene of brown adipocytes that is responsible for their thermogenic capacity. To investigate the role of C/EBPbeta in brown fat, we studied the C/EBPbeta-null mice. When placed in the cold, C/EBPbeta(-/-) mice did not maintain body temperature. This cold-sensitive phenotype occurred, although UCP1 and PGC-1alpha (peroxisome-proliferator-activated receptor gamma co-activator-1alpha) gene expression was unaltered in brown fat of C/EBPbeta(-/-) mice. The UCP1 gene promoter was repressed by the truncated inhibitory C/EBPbeta isoform LIP (liver-enriched transcriptional inhibitory protein, the truncated inhibitory C/EBPbeta isoform). Since C/EBPbeta-null mice lack both C/EBPbeta isoforms, active LAP (liver-enriched transcriptional activatory protein, the active C/EBPbeta isoform) and LIP, the absence of LIP may have a stronger effect than the absence of LAP upon UCP1 gene expression. Gene expression for UCP2 and UCP3 was not impaired in all tissues analysed. In primary brown adipocytes from C/EBPbeta(-/-) mice, induction of gene expression by noradrenaline was preserved. In contrast, the expression of genes related to lipid storage was impaired, as was the amount of triacylglycerol mobilized after acute cold exposure in brown fat from C/EBPbeta(-/-) mice. LPL (lipoprotein lipase) activity was also impaired in brown fat, but not in other tissues of C/EBPbeta(-/-) mice. LPL protein levels were also diminished, but this effect was independent of changes in LPL mRNA, suggesting that C/EBPbeta is involved in the post-transcriptional regulation of LPL gene expression in brown fat. In summary, defective thermoregulation owing to the lack of C/EBPbeta is associated with the reduced capacity to supply fatty acids as fuels to sustain brown fat thermogenesis.

Abstract: The developmental regulation of peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) gene expression was studied in mice and compared with that of marker genes of liver energy metabolism. The PGC-1alpha gene was highly expressed in fetal liver compared with that in adults and remained high in neonatal liver. The regulation of PGC-1alpha gene expression during the fetal and early neonatal periods was dissociated from that of gluconeogenic genes, i.e. the phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) genes. Only under the effects of starvation was PGC-1alpha gene expression induced in parallel to phosphoenolpyruvate carboxykinase and G6Pase mRNAs during the perinatal period. Furthermore, the PGC-1alpha gene was not regulated as part of the developmental program of gene expression associated with the maturation of hepatic gluconeogenesis, as revealed by the impaired PEPCK and G6Pase gene expression but unaltered PGC-1alpha mRNA levels in CCAAT/enhancer-binding protein-alpha-null fetus and neonates. Regulation of the PGC-1alpha gene and that of mitochondrial 3-hydroxy-3-methyl-glutaryl-coenzyme A synthase, acyl-coenzyme A oxidase, and long-chain acyl-coenzyme dehydrogenase, marker genes of lipid catabolism, were dissociated in fetuses and neonates. The expression of lipid catabolism genes was down-regulated in fasted neonates, whereas PGC-1alpha was oppositely regulated. The independent regulation of PGC-1alpha and lipid catabolism genes was also found in peroxisome proliferator-activated receptor-alpha-null neonates, in which PGC-1alpha mRNA levels were unaffected whereas gene expression for 3-hydroxy-3-methyl-glutaryl-coenzyme A synthase and acyl-coenzyme A oxidase was impaired. Thus, regulation of the PGC-1alpha gene is partially dissociated from the patterns of regulation of hepatic genes encoding enzymes involved in gluconeogenesis and lipid catabolism during fetal ontogeny and in response to the initiation of lactation.

Abstract: Recent reports emphasize the importance of mitochondria in white adipose tissue biology. In addition to their crucial role in energy homeostasis, mitochondria are the main site of reactive oxygen species generation. When moderately produced, they function as physiological signaling molecules. Thus, mitochondrial reactive oxygen species trigger hypoxia-dependent gene expression. Therefore the present study tested the implication of mitochondrial reactive oxygen species in adipocyte differentiation and their putative role in the hypoxia-dependent effect on this differentiation. Pharmacological manipulations of mitochondrial reactive oxygen species generation demonstrate a very strong and negative correlation between changes in mitochondrial reactive oxygen species and adipocyte differentiation of 3T3-F442A preadipocytes. Moreover, mitochondrial reactive oxygen species positively and specifically control expression of the adipogenic repressor CHOP-10/GADD153. Hypoxia (1% O2) strongly increased reactive oxygen species generation, hypoxia-inducible factor-1 and CHOP-10/GADD153 expression, and inhibited adipocyte differentiation. All of these hypoxia-dependent effects were partly prevented by antioxidants. By using hypoxia-inducible factor-1alpha (HIF-1alpha)-deficient mouse embryonic fibroblasts, HIF-1alpha was shown not to be required for hypoxia-mediated CHOP-10/GADD153 induction. Moreover, the comparison of hypoxia and CoCl2 effects on adipocyte differentiation of wild type or HIF-1alpha deficient mouse embryonic fibroblasts suggests the existence of at least two pathways dependent or not on the presence of HIF-1alpha. Together, these data demonstrate that mitochondrial reactive oxygen species control CHOP-10/GADD153 expression, are antiadipogenic signaling molecules, and trigger hypoxia-dependent inhibition of adipocyte differentiation.

Abstract: Brown fat differentiation in mice is fully achieved in fetuses at term and entails the acquisition of not only adipogenic but also thermogenic and oxidative mitochondrial capacities. The present study of the mice homozygous for a deletion in the gene for CCAAT/enhancer-binding protein alpha (C/EBPalpha-null mice) demonstrates that C/EBPalpha is essential for all of these processes. Developing brown fat from C/EBPalpha-null mice showed a lack of uncoupling protein-1 expression, impaired adipogenesis, and reduced size and number of mitochondria per cell when compared with wild-type mice. Furthermore, immature mitochondrial morphology was found in brown fat, but not in liver or heart, from C/EBPalpha-null mice. Concordantly, expression of both nuclear and mitochondrial genome-encoded genes for mitochondrial proteins was reduced in C/EBPalpha-null brown fat, although expression of mitochondrial rRNA and mitochondrial DNA content were unaltered. Expression of nuclear respiratory factor-2, thyroid hormone nuclear receptors, and peroxisome proliferator-activated receptor gamma coactivator-1, was delayed in C/EBPalpha-null brown fat. Iodothyronine 5'-deiodinase activity and thyroid hormone content were also reduced in brown fat from C/EBPalpha-null mice, indicating for the first time a crucial role for C/EBPalpha in controlling thyroid status in developing brown fat, which may contribute to impaired mitochondrial biogenesis and cell differentiation. When survival of C/EBPalpha-null mice was achieved by transgenically expressing C/EBPalpha only in the liver, a substantial recovery in brown fat differentiation was found by day 7 of postnatal age, which is associated with a compensatory overexpression of C/EBPdelta and C/EBPbeta.

Abstract: The differentiation of brown adipocytes during late fetal development or in cell culture is associated with enhanced mitochondrial biogenesis and increased gene expression for components of the respiratory chain/oxidative phosphorylation system. We have shown that this is due to a rise in mitochondrial DNA abundance and the corresponding increase in mitochondrial genome transcripts and gene products, as well as to the coordinate induction of nuclear-encoded genes for mitochondrial proteins. We studied how the expression of key components of the transcriptional regulation of mitochondrial biogenesis is regulated during this process. Changes in the expression of nuclear respiratory factor-2/GA-binding protein a and peroxisome proliferator-activated-receptor gamma coactivator-1 (increase) were opposite to those of nuclear respiratory factor-1 and Sp1 (decrease) during the developmental and differentiation-dependent induction of mitochondrial biogenesis in brown fat. These results indicate that the relative roles of transcription factors and coactivators in mediating mitochondrial biogenesis 'in vivo' are highly specific according to the cell type and stimulus that mediate the mitochondriogenic process.

Abstract: The expression of uncoupling protein (UCP)-3 mRNA in skeletal muscle is dramatically reduced during lactation in mice. The reduction in UCP-3 mRNA levels lowers the amount of the UCP-3 protein in skeletal muscle mitochondria during lactation. Spontaneous or abrupt weaning reverses the downregulation of the UCP-3 mRNA but not the reduction in UCP-3 protein levels. In lactating and virgin mice, however, fasting increases UCP-3 mRNA levels. Changes in UCP-3 mRNA occur in parallel with modifications in the levels of free fatty acids, which are reduced in lactation and are upregulated due to weaning or fasting. Modifications in the energy nutritional stress of lactating dams achieved by manipulating litter sizes do not influence UCP-3 mRNA levels in skeletal muscle. Conversely, when mice are fed a high-fat diet after parturition, the downregulation of UCP-3 mRNA and UCP-3 protein levels due to lactation is partially reversed, as is the reduction in serum free fatty acid levels. Treatment of lactating mice with a single injection of bezafibrate, an activator of the peroxisome proliferator-activated receptor (PPAR), raises UCP-3 mRNA in skeletal muscle to levels similar to those in virgin mice. 4-chloro-6-[(2,3-xylidine)-pirimidinylthio] acetic acid (WY-14,643), a specific ligand of the PPAR-alpha subtype, causes the most dramatic increase in UCP-3 mRNA, whereas troglitazone, a specific activator of PPAR-gamma, also significantly increases UCP-3 mRNA abundance in skeletal muscle of lactating mice. However, in virgin mice, bezafibrate and WY-14,643 do not significantly affect UCP-3 mRNA expression, whereas troglitazone is at least as effective as it is in lactating dams. It is proposed that the UCP-3 gene is regulated in skeletal muscle during lactation in response to changes in circulating free fatty acids by mechanisms involving activation of PPARs. The impaired expression of the UCP-3 gene is consistent with the involvement of UCP-3 gene regulation in the reduction of the use of fatty acids as fuel by the skeletal muscle and in impaired adaptative thermogenesis, both of which are major metabolic adaptations that occur during lactation.

Abstract: To investigate relationships between the uncoupling protein (UCP) family and oxidative metabolism in fat pads, we measured the cytochrome oxidase activity, used as an index of oxidative capacity, and the mRNA content encoding UCP1, UCP2 and UCP3. Most oxidative potential was found in the stromal-vascular fraction (SVF) of brown fat and in mature adipocytes of white fat (inguinal and periovarian). Considering the whole fat pads, the oxidative potential observed in mature white adipocytes fraction was not negligible compared with that of brown adipocytes fraction. UCP1 and UCP3 were expressed exclusively in mature brown adipocytes. Whatever the deposit, UCP2 mRNA was mainly localized in the SVF. These results indicate that, in fat, high oxidative potential is not necessarily linked to high UCPs transcripts content and point out the oxidative capacity of SVF from brown fat.

Abstract: The recently identified uncoupling protein-3 (UCP-3) gene, predicted to encode a new member of the family of uncoupling proteins, is preferentially expressed in skeletal muscle and has been related to phenotypes of obesity and type 2 diabetes. We have established that during mouse ontogeny, the expression of the UCP-3 gene is switched on in skeletal muscle just after birth. The induction of UCP-3 gene expression is dependent on the initiation of suckling and particularly on lipid intake. Treatment of newborn mice with activators of peroxisome proliferator-activated receptors (PPARs), such as clofibrate, bezafibrate, or (4-chloro-6-(2,3-xylidine)-pirimidinylthio)acetic acid (WY 14,643), mimics the action of food intake on UCP-3 gene expression. The specific ligand of PPAR-alpha WY 14,643 induces UCP-3 gene expression in a time- and dose-dependent manner, whereas the thiazolidinedione BRL 49653, specific for PPAR-gamma, has no effect. These treatments act without altering circulating free fatty acids. During development, skeletal muscle expresses constitutive levels of PPAR-delta mRNA, whereas expression of the PPAR-gamma gene is undetectable. PPAR-alpha gene expression is developmentally regulated in muscle as it is first expressed at birth, just before UCP-3 gene induction occurs. The induction of UCP-3 gene expression by WY 14,643 is impaired in skeletal muscle of premature neonates, which do not express PPAR-alpha. It is proposed that the UCP-3 gene is predominantly regulated in neonatal muscle by PPAR-alpha activation.

Abstract: The peroxisome proliferator-activated receptors (PPARs) are lipid-activated transcription factors involved in the regulation of lipid metabolism and adipocyte differentiation. Little is known, however, about the control of the expression of the genes encoding each of all three receptor subtypes: alpha, delta, and gamma. We have addressed this question in the brown adipocyte, the only cell type that co-expresses high levels of the three PPAR subtypes. Differentiation of brown adipocytes is associated with enhanced expression of PPAR genes. However, whereas PPARgamma and PPARdelta genes are already expressed in preadipocytes, the mRNA for PPARalpha appears suddenly in association with the acquisition of the terminally differentiated phenotype. Both retinoic acid isomers and PPAR agonists, specific for either PPARalpha or PPARgamma, regulate expression of each PPAR subtype gene in the opposite way: they up-regulate PPARalpha and down-regulate PPARgamma. The effects on PPARalpha mRNA are independent of protein synthesis, whereas inhibition of PPARgamma mRNA expression depends on protein synthesis, except when its specific ligand prostaglandin J2 is used. Our results indicate a strictly opposite autoregulation of PPAR subtypes, which supports specific physiological roles for them in controlling brown fat differentiation and thermogenic activity.

Abstract: Uncoupling protein-3 gene expression in skeletal muscle is up-regulated during postnatal development of mice. A high-carbohydrate diet at weaning induces a decrease in uncoupling protein-3 mRNA levels that does not occur when mice were weaned onto a high-fat diet. Uncoupling protein-3 mRNA levels do not increase in response to fasting in young pups. Only after day 15 of life, when fasting increases serum non-esterified fatty acids, uncoupling protein-3 mRNA is up-regulated by starvation. Over-nutrition or under-nutrition during lactation increases or decreases, respectively, uncoupling protein-3 mRNA expression in skeletal muscle. Regulation of uncoupling protein-3 gene expression in skeletal muscle during development is mediated by ontogenic and nutritional factors determining changes in circulating non-esterified fatty acids.

Abstract: The expression of uncoupling protein-2 (UCP2) mRNA is up-regulated during the differentiation of brown adipocytes in primary culture. When differentiation of brown adipocytes is impaired, UCP2 mRNA expression is down-regulated. 9-cis Retinoic acid causes a dose-dependent induction of UCP2 mRNA levels in brown adipocytes, whereas all-trans retinoic acid has no effect. Specific agonists of retinoid X receptors (RXR) induce UCP2 mRNA expression, whereas specific activators of retinoic acid receptors do not. 9-cis Retinoic acid, acting through RXR receptors, is identified as a major regulator of the expression of the UCP2 gene in the brown fat cell.