RESUMO
The uncoupling protein homologs UCP2 and UCP3 have been proposed as candidate genes for the regulation of lipid metabolism. Within the context of this hypothesis, we have compared, from fed and fasted rats, changes in gene expression of skeletal muscle UCP2 and UCP3 with those of carnitine palmitoyltransferase I and medium-chain acyl-CoA dehydrogenase, two key enzymes regulating lipid flux across the mitochondrial beta-oxidation pathway. In addition, changes in gene expression of peroxisome proliferator-activated receptor gamma, a nuclear transcription factor implicated in lipid metabolism, were also investigated. The results indicate that in response to fasting, the mRNA levels of UCP2, UCP3, carnitine palmitoyltransferase I and medium-chain acyl-CoA dehydrogenase are markedly increased, by three- to sevenfold, in the gastrocnemius and tibialis anterior (fast-twitch muscles, predominantly glycolytic or oxidative-glycolytic), but only mildly increased, by less than twofold, in the soleus (slow-twitch muscle, predominantly oxidative). Furthermore, such muscle-type dependency in fasting-induced transcriptional changes in UCP2, UCP3, carnitine palmitoyltransferase and medium-chain acyl-CoA dehydrogenase persists when the increase in circulating levels of free fatty acids during fasting is abolished by the anti-lipolytic agent nicotinic acid - with blunted responses only in the slow-twitch muscle contrasting with unabated increases in fast-twitch muscles. Independently of muscle type, however, the mRNA levels of peroxisome proliferator-activated receptor gamma are not altered during fasting. Taken together, these studies indicate a close association between fasting-induced changes in UCP2 and UCP3 gene expression with those of key regulators of lipid oxidation, and are hence consistent with the hypothesis that these UCP homologs may be involved in the regulation of lipid metabolism. Furthermore, they suggest that in response to fasting, neither the surge of free fatty acids in the circulation nor induction of the peroxisome proliferator-activated receptor gamma gene may be required for the marked upregulation of genes encoding the UCP homologs and key enzymes regulating lipid oxidation in fast-twitch muscles.
Assuntos
Proteínas de Transporte/metabolismo , Jejum/fisiologia , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/fisiologia , Proteínas/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Enzimas/metabolismo , Canais Iônicos , Metabolismo dos Lipídeos , Masculino , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Niacina/farmacologia , Oxirredução , Ratos , Ratos Sprague-Dawley , Proteína Desacopladora 2 , Proteína Desacopladora 3 , Regulação para CimaRESUMO
A role for uncoupling protein (UCP) 3 in fatty acid metabolism is reviewed within the context of our proposal, first put forward in 1998, that this homologue of UCP1 may be involved in the regulation of lipids as fuel substrate rather than in the mediation of thermogenesis. Since then, the demonstrations of muscle-type differences in UCP3 gene regulation in response to dietary manipulations (starvation, high-fat feeding) or to pharmacological interferences with the flux of lipid substrates between adipose-tissue stores and skeletal-muscle mitochondrial oxidation are all in accord with this proposed role for UCP3 in regulating lipids as fuel substrate. However, given the current limitations of gene-knockout technology for evaluating/interpreting the functional importance of genes encoding mitochondrial membrane proteins, the transition from 'associative' to 'cause-and-effect' evidence for a physiological role of UCP3 in regulating fatty acid metabolism will have to await the development of assays that are sensitive to changes in UCP3 activity. Furthermore, in evaluating the physiological regulators of UCP3, the available evidence points to the existence of adipose-derived factor(s) which, independently of circulating levels of free fatty acids, initiates events leading to the transcription of genes encoding UCP3 and key enzymes of lipid oxidation in the fast glycolytic or fast oxidative-glycolytic muscles, i.e. in the bulk of the skeletal-muscle mass. It is proposed that in tissues where UCP3 co-exists with UCP2 (skeletal muscle, brown adipose tissue, heart) they may act in concert in the overall regulation of lipid oxidation, concomitant to the prevention of lipid-induced oxidative damage.
Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Transporte/fisiologia , Ácidos Graxos/metabolismo , Tecido Adiposo/metabolismo , Animais , Humanos , Resistência à Insulina , Canais Iônicos , Metabolismo dos Lipídeos , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Modelos Biológicos , Músculo Esquelético/metabolismo , Temperatura , Proteína Desacopladora 3RESUMO
During fasting, when overall metabolism changes, the contribution of glucose and fatty acids (FA) to cardiac energy production alters as well. Here, we examined if the heart is able to adapt to such fasting-induced changes by modulation of its gene expression. Rats were fed ad libitum or fasted for 46 h, resulting in reduced circulating glucose levels and a 3-fold rise in FA. Besides changes in the cardiac activity or content of proteins involved in glucose or FA metabolism, mRNA levels also altered. The cardiac expression of genes coding for glucose-handling proteins (glucose transporter GLUT4, hexokinase I and II) was up to 70% lower in fasted than in fed rats. In contrast, the mRNA levels of various genes involved in FA transport and metabolism (FA translocase/CD36, muscle-type carnitine palmitoyl transferase 1, long-chain acyl-CoA dehydrogenase) and of the uncoupling protein UCP-3 increased over 50% in hearts of fasted rats. Surprisingly, mRNA levels of the fatty acid- activated transcription factors PPARalpha and PPARbeta/delta were reduced in hearts of fasted rats, whereas in livers, fasting led to a marked rise in PPARalpha mRNA. Reducing FA levels by nicotinic acid administration during the final 8 h of fasting did not affect the expression of the majority of metabolic genes, but totally abolished the induction of UCP-3. In conclusion, the adult rat heart responds to changes in nutritional status, as provoked by 46 h fasting, through adjustment of glucose as well as FA metabolism at the level of gene expression.
Assuntos
Jejum/fisiologia , Expressão Gênica , Proteínas Musculares , Miocárdio/metabolismo , Acil-CoA Desidrogenase , Acil-CoA Desidrogenase de Cadeia Longa/genética , Animais , Glicemia/metabolismo , Northern Blotting , Antígenos CD36 , Carnitina O-Palmitoiltransferase/genética , Proteínas de Transporte/genética , Metabolismo Energético/genética , Ensaio de Imunoadsorção Enzimática , Ácidos Graxos/sangue , Transportador de Glucose Tipo 4 , Hexoquinase/genética , Canais Iônicos , Masculino , Glicoproteínas de Membrana/genética , Proteínas Mitocondriais , Proteínas de Transporte de Monossacarídeos/genética , Niacina/farmacologia , Transportadores de Ânions Orgânicos/genética , RNA Mensageiro/análise , Ratos , Ratos Sprague-Dawley , Receptores Citoplasmáticos e Nucleares/genética , Fatores de Transcrição/genética , Proteína Desacopladora 3RESUMO
Toxoplasma gondii (Me49 strain) infection into Swiss Webster mice is followed by hypermetabolism and weight loss in the acute phase lasting 14 days. In the subsequent chronic phase of infection, mice showed either a resolution of hypermetabolism and partial weight recovery (Gainers) or persistent hypermetabolism, with stable weight loss (Non-Gainers). The hypermetabolic response was not associated with an augmentation in the thermogenic uncoupling protein 1 (UCP1) mRNA expression in interscapular brown adipose tissue (BAT), but rather UCP1 expression was reduced. Hypermetabolism is associated with high lipid oxidation as attested by a low respiratory quotient (RQ). Neither BAT nor sympathetic nervous system appear to be involved in the increased lipid utilization, since propranolol did not increase the lower RQ in infected mice. The mitochondrial lipid oxidation blocker mercaptoacetate did not reestablish the respiratory quotient RQ in acute infection (on day 4) and in chronically infected Non-Gainer mice. This suggests an important extra-mitochondrial mechanism of lipid oxidation. Increased lipid peroxidation was detected especially in serum, lung, spleen and liver, which are rich in macrophage-type cells. Following infection peritoneal macrophages exhibited an enhanced capacity to produce reactive oxygen species (ROS). Using IFN-gamma knockout mice we observed that not only the hypermetabolic response was ablated in these mice but there was not a marked increase in ROS production or preferential oxidation/peroxidation of lipids in the acute phase of infection prior to the cachectic phase. The present study described a novel hypermetabolic mechanism involving enhanced lipid peroxidation dependent on IFN-gamma, especially associated with tissues rich in macrophages.
Assuntos
Proteínas de Transporte/metabolismo , Citocinas/biossíntese , Interferon gama/metabolismo , Peroxidação de Lipídeos , Proteínas de Membrana/metabolismo , Toxoplasmose Animal/metabolismo , Tecido Adiposo Marrom/metabolismo , Animais , Peso Corporal , Encéfalo/metabolismo , Citocinas/sangue , Ingestão de Alimentos/fisiologia , Metabolismo Energético , Interferon gama/genética , Canais Iônicos , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Camundongos Knockout , Proteínas Mitocondriais , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Respiração , Baço/metabolismo , Toxoplasma , Proteína Desacopladora 1RESUMO
We have investigated the effect of 24-h fasting on basal proton leak and uncoupling protein (UCP) 3 expression at the protein level in subsarcolemmal and intermyofibrillar skeletal muscle mitochondria. In fed rats, the two mitochondrial populations displayed different proton leak, but the same protein content of UCP3. In addition, 24-h fasting, both at 24 and 29 degrees C, induced an increase in proton leak only in subsarcolemmal mitochondria, while UCP3 content increased in both the populations. From the present data, it appears that UCP3 does not control the basal proton leak of skeletal muscle mitochondria.
Assuntos
Proteínas de Transporte/metabolismo , Mitocôndrias Musculares/metabolismo , Músculo Esquelético/metabolismo , Miofibrilas/metabolismo , Sarcolema/metabolismo , Animais , Jejum , Canais Iônicos , Cinética , Masculino , Proteínas Mitocondriais , Prótons , Ratos , Ratos Wistar , Temperatura , Proteína Desacopladora 3RESUMO
During the past few years, there have been two major developments, if not revolutions, in the field of energy balance and weight regulation. The first at the molecular level, which was catalysed by developments in DNA screening technology together with the mapping of the human genome, has been the tremendous advances made in the identification of molecules that play a role in the control of food intake and metabolic rate. The second, at the systemic level, which centered upon the use of modern technologies or more robust analytical techniques for assessing human energy expenditure in response to starvation and overfeeding, has been the publication of several papers providing strong evidence that adaptive thermogenesis plays a much more important role in the regulation of body weight and body composition than previously thought. Within these same few years, several new members of the mitochondrial carrier protein family have been identified in a variety of tissues and organs. All apparently possess uncoupling properties in genetically-modified systems, with two of them (uncoupling protein (UCP) 2 and UCP3) being expressed in adipose tissues and skeletal muscles, which are generally recognised as important sites for variations in thermogenesis and/or in substrate oxidation. Considered as breakthrough discoveries, the cloning of these genes has generated considerable optimism for rapid advances in our molecular understanding of adaptive thermogenesis, and for the identification of new targets for pharmacological management of obesity and cachexia. The present paper traces first, from a historical perspective, the landmark events in the field of thermogenesis that led to the identification of these genes encoding candidate UCP, and then addresses the controversies and on-going debate about their physiological importance in adaptive thermogenesis, in lipid oxidation or in oxidative stress. The general conclusion is that UCP2 and UCP3 may have distinct primary functions, with UCP3 implicated in regulating the flux of lipid substrates across the mitochondria and UCP2 in the control of mitochondrial generation of reactive oxygen species. The distinct functions of these two UCP1 homologues have been incorporated in a conceptual model to illustrate how UCP2 and UCP3 may act in concert in the overall regulation of lipid oxidation concomitant to the prevention of lipid-induced oxidative damage.
Assuntos
Hipotálamo/fisiologia , Metabolismo dos Lipídeos , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Obesidade/metabolismo , Termogênese/fisiologia , Tecido Adiposo/metabolismo , Animais , Composição Corporal/fisiologia , Peso Corporal , Proteínas de Transporte/fisiologia , Proteínas Alimentares/metabolismo , Metabolismo Energético , Ácidos Graxos não Esterificados/metabolismo , Humanos , Canais Iônicos , Proteínas de Membrana/fisiologia , Proteínas de Desacoplamento Mitocondrial , Modelos Biológicos , Músculo Esquelético/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Proteínas/fisiologia , Inanição , Sistema Nervoso Simpático/metabolismo , Proteína Desacopladora 1 , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
We wished to gain insights into the role of skeletal muscle uncoupling protein-3 (UCP-3) in the elevated efficiency of fat recovery during refeeding after starvation. Previous observations have revealed that muscle UCP-3 expression is downregulated in rats during refeeding at 22 degrees C. Therefore, we investigated whether this also occurs during refeeding at thermoneutrality (29 C) or in the cold (6 C), since at these environmental temperatures the refed animals also show diminished thermogenesis and a higher rate of fat deposition than controls. The UCP-3 mRNA level in the skeletal muscles studied (soleus, gastrocnemius and tibialis anterior) was significantly lower in the refed group than in controls at thermoneutrality, but there were no such differences between these two groups in the cold. This effect of cold, namely abolishing refeeding-induced downregulation of skeletal muscle UCP, is specific to UCP-3 since the gene expression of skeletal muscle UCP-2 remained significantly lower in the refed than in the controls both at thermoneutrality and in the cold. These findings during refeeding in the cold therefore dissociate UCP-3 gene regulation from the adaptive reduction in thermogenesis that accelerates fat deposition during weight recovery. They also reveal differential responses of UCP-3 and UCP-2, whose significance is discussed in the light of our previously proposed hypothesis, which centers upon a role for these UCP homologues in the regulation of lipids as a fuel substrate.
Assuntos
Ração Animal , Proteínas de Transporte/genética , Temperatura Baixa , Expressão Gênica , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Inanição/genética , Animais , Regulação para Baixo , Canais Iônicos , Masculino , Proteínas/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Temperatura , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
Studies of starvation and refeeding have implicated the genes coding for uncoupling protein-3 and -2 (UCP3, UCP2) as candidate genes in the regulation of lipids as metabolic fuels in skeletal muscle. To gain insight into the role of free fatty acid (FFA) flux in regulating the expression of these muscle UCP homologues, we recently reported that, in response to the anti-lipolytic agent nicotinic acid, utilized to reduce FFA flux at the input supply (i.e. circulating) level in fed and fasted rats, expression of the UCP3 and UCP2 genes was reduced in the soleus (predominantly slow-oxidative fibres), but not in the gastrocnemius (predominantly fast-glycolytic fibres) or tibials anterior (predominantly fast-oxidative-glycolytic fibres) muscles. In the present study, we examined UCP2 and UCP3 gene expression in these muscles from fed or fasted rats treated with etomoxir, an inhibitor of FFA flux at the output (i.e. mitochondrial oxidation) level. Fasting per se resulted in a threefold increase in serum FFA (P < 0.001) and in marked increases in the messenger ribonucleic acid (mRNA) expression of both UCP2 and UCP3 in all three muscles (P < 0.001). Treatment with etomoxir had no significant effect on serum FFA in the fed rats, but further elevated serum FFA in the fasted rats (P < 0.001). The mRNA levels of both UCP3 and UCP2 in response to etomoxir were significantly reduced in the tibialis anterior muscle in both fed and fasted states (P < 0.01), unaltered in the gastrocnemius muscle in both fed and fasted states and unaltered in the soleus muscle in the fed state, but increased in the fasted state, in parallel with the etomoxir-induced changes in serum FFA levels. Taken together, these results suggest the existence of positive feedback loops between FFA flux and muscle UCPs only in oxidative muscles--with that loop operating at the input FFA supply level for muscles with predominantly slow-oxidative fibres, and at the output FFA oxidation level for muscles with predominantly fast-oxidative-glycolytic fibres.
Assuntos
Proteínas de Transporte/genética , Ácidos Graxos não Esterificados/fisiologia , Regulação da Expressão Gênica/fisiologia , Proteínas de Membrana Transportadoras , Mitocôndrias Musculares/metabolismo , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Proteínas/genética , Animais , Glicemia/análise , Ácidos Graxos não Esterificados/antagonistas & inibidores , Ácidos Graxos não Esterificados/sangue , Canais Iônicos , Masculino , Oxirredução , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
UCP2 and UCP3 are two recently cloned genes with high sequence homology to the gene for uncoupling protein (UCP)-1, which regulates thermogenesis in brown adipose tissue. In the context of the current debate about whether UCP2 and UCP3 in the skeletal muscle may also function as mediators of thermogenesis or as regulators of lipids as fuel substrate, we have examined their mRNA expressions in rat gastrocnemius muscle in response to dietary manipulations known to differentially affect thermogenesis during the phase of weight recovery after starvation. Compared with ad libitum-fed control rats, the refeeding of isocaloric amounts of a low-fat (high-carbohydrate) diet resulted in lower energy expenditure and lower mRNA levels of muscle UCP2 and UCP3. This downregulation of UCP homologs was abolished by the refeeding of a high-fat diet, even though energy expenditure was significantly lower during refeeding on the high-fat than on the low-fat diet. Furthermore, major alterations in the fatty acid composition of the refeeding diet in favor of n-6 polyunsaturated or medium-chain fatty acids resulted in significant increases in energy expenditure, but with no significant changes in the expression of skeletal muscle UCP homologs. Regression analysis of gastrocnemius UCP mRNA levels against parameters that included body composition, energy expenditure, and plasma levels of free fatty acids (FFAs), insulin, and glucose as well as the increase in plasma glucose after a glucose load, revealed that only the latter (an index of insulin resistance) could explain the variability in muscle UCP2 and UCP3 mRNA expressions (r = 0.41, P < 0.02; r = 0.45, P < 0.01, respectively). Taken together, these data are at variance with a role for skeletal muscle UCP2 and UCP3 in dietary regulation (or modulation) of thermogenesis. However, they are consistent with the notion that these UCP homologs may function as regulators of lipids as fuel substrate and raise the possibility that high-fat induced upregulation of muscle UCP2 and UCP3 may be more closely linked to insulin resistance than to changes in circulating FFAs.
Assuntos
Proteínas de Transporte/genética , Gorduras na Dieta/farmacologia , Expressão Gênica/fisiologia , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Proteínas/genética , Inanição/genética , Animais , Gorduras na Dieta/administração & dosagem , Relação Dose-Resposta a Droga , Ácidos Graxos/farmacologia , Resistência à Insulina/genética , Canais Iônicos , Masculino , Ratos , Ratos Sprague-Dawley , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
The relationship between UCP2 and UCP3 expression and mitochondrial proton conductance of rat skeletal muscle was examined. Rats were starved for 24 h and the levels of UCP2 and UCP3 mRNA and UCP3 protein were determined by Northern and Western blots. Proton conductance was measured by titrating mitochondrial respiration rate and membrane potential with malonate. Starvation increased UCP2 and UCP3 mRNA levels more than 5-fold and 4-fold, respectively, and UCP3 protein levels by 2-fold. However, proton conductance remained unchanged. These results suggest either that Northern and Western blots do not reflect the levels of active protein or that these UCPs do not catalyse the basal proton conductance in skeletal muscle mitochondria.
Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Membrana Transportadoras , Mitocôndrias/metabolismo , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Proteínas/metabolismo , Prótons , Desacopladores/metabolismo , Animais , Northern Blotting , Western Blotting , Proteínas de Transporte/genética , Eletrofisiologia , Feminino , Privação de Alimentos , Canais Iônicos , Cinética , Proteínas/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
Uncoupling proteins 3 and 2 (UCP3 and UCP2) are two newly cloned genes that have been implicated in the regulation of lipids as fuel substrate in skeletal muscle on the basis that their mRNA expressions are upregulated during starvation (when fat stores are being rapidly mobilized) and downregulated during the early phase of refeeding (when fat stores are being rapidly replenished). To test the hypothesis that circulating free fatty acids (FFAs) may have a physiological role as an interorgan signal linking these dynamic changes in the fat stores to skeletal muscle expression of UCP3 and UCP2, the mRNA levels of these UCP homologs were examined in fed and fasted rats treated with the antilipolytic agent nicotinic acid. In 46-h fasted rats, we observed a threefold increase in serum FFA levels and increases in UCP3 and UCP2 mRNA levels that were more marked in the gastrocnemius and tibialis anterior muscles (predominantly fast-twitch fibers) than in the soleus muscle (predominantly slow-twitch fibers). Treatment with nicotinic acid blunted the fasting-induced increase in serum FFA levels and prevented the increase in mRNA levels of UCP3 and UCP2 in the soleus muscle, but had little or no effect on the elevated mRNA levels of these UCP homologs in the gastrocnemius and tibialis anterior muscles. Furthermore, treatment of ad libitum-fed animals with nicotinic acid resulted in a twofold reduction in serum FFA levels (i.e., by a magnitude similar to that observed during early refeeding) and significant reductions in UCP3 and UCP2 mRNA levels in the soleus muscle, but not in the gastrocnemius or tibialis anterior muscles. These results revealed a muscle-type dependency in the way UCP2 and UCP3 gene expression in skeletal muscle is regulated, and suggest that the hypothesis that circulating FFAs function as an interorgan signal between fat stores and skeletal muscle UCP3 and UCP2 gene expression is adequate only for slow-twitch (oxidative) muscles. Consequently, a signal(s) other than circulating FFAs must be implicated in the link between dynamic changes in body fat stores and UCP expression in predominantly fast-twitch (glycolytic/oxidative-glycolytic) muscles, which constitute the major fiber type of the total skeletal muscle mass and which have high susceptibility to developing insulin resistance and impairment in substrate utilization in metabolic diseases.
Assuntos
Tecido Adiposo/metabolismo , Proteínas de Transporte/genética , Ácidos Graxos não Esterificados/sangue , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Proteínas/genética , Transdução de Sinais , Tecido Adiposo/efeitos dos fármacos , Animais , Glicemia/metabolismo , Proteínas de Transporte/fisiologia , Jejum , Alimentos , Expressão Gênica , Canais Iônicos , Masculino , Músculo Esquelético/efeitos dos fármacos , Niacina/farmacologia , Proteínas/fisiologia , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Desacopladores , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
The mRNA expressions of UCP2 and UCP3, two newly described genes with high sequence homology to the uncoupling protein UCP1 in brown adipose tissue (BAT), were examined in two skeletal muscles (gastrocnemius and soleus) as well as in interscapular BAT (IBAT) of the rat in response to food deprivation and controlled refeeding. In IBAT (a tissue highly dependent on lipids for thermogenesis), the pattern of mRNA expression of UCP2 and UCP3 closely follows that of UCP1: it was markedly down-regulated during food deprivation (when this tissue's thermogenesis and lipid fuel requirements are decreased) and restored to control levels by day 5 of refeeding. By contrast, in the gastrocnemius muscle (a mixed fiber type muscle with a high capacity to shift between glucose and lipids as fuel substrate), mRNA expression of both UCP2 and UCP3 mRNA was found to be markedly up-regulated during food deprivation (when this tissue's thermogenesis is also decreased but its lipid fuel utilization is increased). The expressions were subsequently found to be markedly down-regulated upon transition to refeeding, with mRNA levels remaining below control levels on days 3, 5, and 10 of refeeding (period of enhanced efficiency of body fat deposition). In the soleus muscle (an oxidative type muscle with higher dependency on lipids than the gastrocnemius, and hence with a lower capacity to shift between lipids and glucose as fuel substrate), UCP homologues were also found to be up-regulated during food deprivation, but changes in their mRNA expression contrast with those in the gastrocnemius muscle both in their much lower magnitude of response to food deprivation and in their more rapid restoration to control levels during refeeding. Up-regulation of UCP2 and UCP3 gene expressions in skeletal muscle during food deprivation was found to persist at thermoneutrality (i.e., under conditions of reduced thermoregulatory thermogenesis). Together, these tissue-dependent differential mRNA expressions of the UCP homologues in IBAT, gastrocnemius, and soleus muscles during food deprivation and refeeding are much more consistent with a role for UCP2 and UCP3 in the regulation of lipids as fuel substrate rather than as mediators of regulatory thermogenesis.
Assuntos
Tecido Adiposo Marrom/fisiologia , Metabolismo Energético/fisiologia , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Músculo Esquelético/fisiologia , Proteínas/metabolismo , Desacopladores/metabolismo , Animais , Composição Corporal , Regulação da Temperatura Corporal/fisiologia , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Privação de Alimentos/fisiologia , Regulação da Expressão Gênica , Canais Iônicos , Metabolismo dos Lipídeos , Masculino , Proteínas/genética , RNA Mensageiro/biossíntese , Ratos , Ratos Sprague-Dawley , Homologia de Sequência de Aminoácidos , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
Endurance exercise training has been shown to decrease diet-induced thermogenesis (DIT) in rats and humans. In rodents, most thermogenesis is thought to occur in brown adipose tissue via activation of the uncoupling protein-1 (UCP1) and in skeletal muscle. Since the level of UCP1 mRNA in rat BAT was reported to be unmodified by exercise training, the newly described uncoupling proteins UCP2 and UCP3 could be responsible for the decreased DIT in trained rats. UCP3 mRNA levels in endurance-trained rats were found to be reduced by 76% and 59% in tibialis anterior and soleus muscles, respectively. UCP2 mRNA levels were also decreased in tibialis anterior and in heart by 54% and 41%, respectively. Neither white adipose tissue UCP2 nor brown adipose tissue UCP1, UCP2, and UCP3 mRNA levels were modified. The results of this study show that a need for a higher metabolic efficiency is associated with decreased mRNA expression of the uncoupling proteins in skeletal and heart muscles, which would decrease energy dissipation in these tissues. The down-regulation of UCP3 and UCP2 expressions might also contribute to the rapid weight gain known to occur when exercise training ceased.
Assuntos
Proteínas de Transporte/biossíntese , Proteínas de Membrana/biossíntese , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Atividade Motora/fisiologia , Músculo Esquelético/metabolismo , Biossíntese de Proteínas , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Transporte/genética , Citrato (si)-Sintase/metabolismo , DNA Complementar , Humanos , Canais Iônicos , Masculino , Proteínas de Membrana/genética , Dados de Sequência Molecular , Proteínas/genética , RNA Mensageiro , Ratos , Homologia de Sequência de Aminoácidos , Proteína Desacopladora 1 , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
A new member of the uncoupling protein (UCP) family called UCP3 has recently been cloned and shown to be highly expressed in skeletal muscle of rodents and humans. In the present study, UCP3 was overexpressed in C2C12 myoblasts where it acts as an uncoupling protein. Changes in UCP3 mRNA expression were examined in rodent muscles under conditions known to modulate thermogenesis in brown adipose tissue. In skeletal muscle, UCP3 expression did not change in response to 48 h of cold exposure (6 degrees C), whereas it was decreased by 81% or increased 5.6-fold by 1 week of 50% food restriction or fasting, respectively. It was also decreased by 36% in soleus muscle of obese (fa/fa) as compared with lean Zucker rats. The unexpected rise of UCP3 mRNA level induced by fasting did not change in vitro muscle basal heat production rate but decreased by 31% the capacity to produce heat in response to the uncoupler carbonylcyanide p-trifluoromethoxyphenylhydrazone. This decrease may reflect underlying uncoupling by UCP3. Up-regulation of UCP3 mRNA after a 24-h fast was still observed in mice exposed at thermoneutrality. These results show that the increase in UCP3 expression induced by fasting is associated with the maintenance of thermogenesis measured in muscle in vitro and is not modulated by environmental temperature. The notion that UCP3 expression is modulated by food intake is of importance to better understand the pathophysiology of obesity in humans.
Assuntos
Proteínas de Transporte/genética , Alimentos , Músculo Esquelético/metabolismo , Temperatura , Animais , Calorimetria , Linhagem Celular , Feminino , Regulação da Expressão Gênica , Canais Iônicos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias Musculares/metabolismo , Proteínas Mitocondriais , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Ratos Zucker , Proteína Desacopladora 3RESUMO
Targeted disruption of mouse beta3-adrenoceptor was generated by homologous recombination, and validated by an acute in vivo study showing a complete lack of effect of the beta3-adrenoceptor agonist CL 316,243 on the metabolic rate of homozygous null (-/-) mice. In brown adipose tissue, beta3-adrenoceptor disruption induced a 66% decrease (P < 0.005) in beta1-adrenoceptor mRNA level, whereas leptin mRNA remained unchanged. Chronic energy balance studies in chow-fed mice showed that in -/- mice, body fat accumulation was favored (+41%, P < 0.01), with a slight increase in food intake (+6%, NS). These effects were accentuated by high fat feeding: -/- mice showed increased total body fat (+56%, P < 0.025) and food intake (+12%, P < 0.01), and a decrease in the fat-free dry mass (-10%, P < 0.05), which reflects a reduction in body protein content. Circulating leptin levels were not different in -/- and control mice regardless of diet. The significant shift to the right in the positive correlation between circulating leptin and percentage of body fat in high fat-fed -/- mice suggests that the threshold of body fat content inducing leptin secretion is higher in -/- than in control mice. Taken together, these studies demonstrate that beta3-adrenoceptor disruption creates conditions which predispose to the development of obesity.
Assuntos
Composição Corporal , Proteínas/fisiologia , Receptores Adrenérgicos beta/fisiologia , Tecido Adiposo/fisiologia , Animais , Northern Blotting , Regulação da Temperatura Corporal , Células Cultivadas , Gorduras na Dieta/administração & dosagem , Metabolismo Energético , Leptina , Masculino , Camundongos , Proteínas/análise , Receptores Adrenérgicos beta/genética , Receptores Adrenérgicos beta 1/fisiologia , Receptores Adrenérgicos beta 3 , Receptores para LeptinaRESUMO
The control of uncoupling protein-2 (UCP2) mRNA expression in rat brown adipose tissue (BAT), heart and skeletal muscles was examined. Cold exposure (48 h) increased UCP2 mRNA in BAT, heart and soleus muscle by 2.4-, 4.3- and 2.6-fold, respectively. Fasting (48 h) had no effect on UCP2 mRNA expression neither in BAT nor in heart, but markedly increased it in skeletal muscles. While the upregulation of UCP2 mRNA in response to cold exposure is in line with a putative uncoupling role for this protein in thermoregulatory thermogenesis, the unexpected upregulation of UCP2 in skeletal muscles in response to fasting seems inconsistent with its role as an uncoupling protein involved in dietary regulation of thermogenesis.
Assuntos
Temperatura Baixa , Jejum , Regulação da Expressão Gênica , Proteínas de Membrana Transportadoras , Proteínas Mitocondriais , Proteínas/genética , Tecido Adiposo Marrom/metabolismo , Animais , Regulação da Temperatura Corporal , Canais Iônicos , Masculino , Dados de Sequência Molecular , Músculo Esquelético/metabolismo , Miocárdio/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Proteína Desacopladora 2RESUMO
Brown adipose tissue (BAT) and skeletal muscle are important sites of nonshivering thermogenesis. The uncoupling protein-1 (UCP1) is the main effector of nonshivering thermogenesis in BAT and the recently described ubiquitous UCP2 [1] has been implicated in energy balance. In an attempt to better understand the biochemical events underlying nonshivering thermogenesis in muscle, we screened a human skeletal muscle cDNA library and isolated three clones: UCP2, UCP3L and UCP3S. The novel UCP3 was 57% and 73% identical to human UCP1 and UCP2, respectively, highly skeletal muscle-specific and its expression was unaffected by cold acclimation. This new member of the UCP family is a candidate protein for the modulation of the respiratory control in skeletal muscle.
Assuntos
Proteínas de Transporte/genética , Regulação da Expressão Gênica , Proteínas de Membrana Transportadoras , Mitocôndrias Musculares/metabolismo , Proteínas Mitocondriais , Músculo Esquelético/metabolismo , Tecido Adiposo/metabolismo , Tecido Adiposo Marrom/metabolismo , Sequência de Aminoácidos , Northern Blotting , Regulação da Temperatura Corporal/fisiologia , Proteínas de Transporte/química , Proteínas de Transporte/metabolismo , Clonagem Molecular , Sondas de DNA , Metabolismo Energético , Humanos , Canais Iônicos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Dados de Sequência Molecular , Proteínas/química , Proteínas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Homologia de Sequência de Aminoácidos , Proteína Desacopladora 1 , Proteína Desacopladora 2 , Proteína Desacopladora 3RESUMO
Exon amplification has been applied to a 2.5 Mb region of chromosome 21 that has been associated with some features of Down syndrome (DS). Identification of the majority of genes from this region will facilitate the correlation of the over-expression of particular genes with specific phenotypes of DS. Over 100 gene fragments have been isolated from this 2.5 Mb segment. The exons have been characterized by sequence analysis, comparison with public databases and expansion to cDNA clones. Localization of the exons to chromosome 21 has been determined by hybridization to genomic Southern blots and to YAC and cosmid clones representing the region. This has resulted in a higher resolution physical map with a marker approximately every 25 kb. This integrated physical and transcript map will be valuable for fine mapping of DNA from individuals with partial aneuploidy of chromosome 21 as well as for assessing and ultimately generating a complete gene map of this segment of the genome.