Comparison of Atrogin-1/MAFbx mRNA Expression in the Gizzards of Egg- and Meat-Type Chickens

We examined atrogin-1/MAFbx mRNA expression in the smooth muscle of gizzards from egg- and meat-type chickens. Gizzard weight relative to body weight was significantly lower in the meat-type chickens than in the egg-type at 14 d of age. In contrast, the level of atrogin-1/MAFbx mRNA in the gizzard was significantly higher in the meat-type chickens than in the egg-type chickens. Thus atrogin-1/MAFbx mRNA expression in the smooth muscle of the gizzard was higher in meat-type chickens than in egg-type chickens, in contrast to its expression in the skeletal muscles.     

Cold-induced mitochondrial uncoupling and expression of chicken UCP and ANT mRNA in chicken skeletal muscle

Although bird species studied thus far have no distinct brown adipose tissue (BAT) or a related thermogenic tissue, there is now strong evidence that non-shivering mechanisms in birds may play an important role during cold exposure. Recently, increased expression of the duckling homolog of the avian uncoupling protein (avUCP) was demonstrated in cold-acclimated ducklings [Raimbault et al., Biochem. J. 353 (2001) 441-444]. Among the mitochondrial anion carriers, roles for the ATP/ADP antiporter (ANT) as well as UCP variants in thermogenesis are proposed. The present experiments were conducted (i) to examine the effects of cold acclimation on the fatty acid-induced uncoupling of oxidative phosphorylation in skeletal muscle mitochondria and (ii) to clone the cDNA of UCP and ANT homologs from chicken skeletal muscle and study differences compared to controls in expression levels of their mRNAs in the skeletal muscle of cold-acclimated chickens. The results obtained here show that suppression of palmitate-induced uncoupling by carboxyatractylate was greater in the subsarcolemmal skeletal muscle mitochondria from cold-acclimated chickens than that for control birds. An increase in mRNA levels of avANT and, to lesser degree, of avUCP in the skeletal muscle of cold-acclimated chickens was also found. Taken together, the present studies on cold-acclimated chickens suggest that the simultaneous increments in levels of avANT and avUCP mRNA expression may be involved in the regulation of thermogenesis in skeletal muscle.     

Cold exposure differently influences mitochondrial energy efficiency in rat liver and skeletal muscle

This study deals with mitochondrial energy efficiency in liver and skeletal muscle mitochondria in 15 days cold exposed rats. Cold exposure strongly increases the sensitivity to uncoupling by added palmitate of skeletal muscle but not liver mitochondria, while mitochondrial energy coupling in the absence of fatty acids is only slightly affected by cold in liver and skeletal muscle. In addition, uncoupling protein 3 content does not follow changes in skeletal muscle mitochondrial coupling. It is therefore concluded that skeletal muscle could play a direct thermogenic role based on fatty acid-induced mild uncoupling of mitochondrial oxidative phosphorylation.     

Membrane potential and H2O2 production in duodenal mitochondria from broiler chickens (Gallus gallus domesticus) with low and high feed efficiency

Increased hydrogen peroxide (H2O2) production was observed in duodenal mitochondria obtained from broiler chickens with low feed efficiency (FE). As a decrease in mitochondrial membrane potential (Deltapsi(m)) due to mild uncoupling of oxidative phosphorylation reduces reactive oxygen species production, this study was conducted to evaluate the effect of uncoupling on Deltapsi(m) and H2O2 production in duodenal mitochondria isolated from broilers with low (0.48+/-0.02) and high (0.68+/-0.01) FE. Membrane potential and H2O2 production were measured fluorometrically and in the presence of different levels of an uncoupler, carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP). The Deltapsi(m) was higher (P相似文献   

Tissue variation of mitochondrial oxidative phosphorylation efficiency in cold-acclimated ducklings

We investigated the oxidative phosphorylation efficiency of liver and gastrocnemius muscle mitochondria in thermoneutral and cold-acclimated ducklings. The yield of oxidative phosphorylation was lower in muscle than in liver mitochondria, a difference that was associated with a higher proton conductance in muscle mitochondria. Cold exposure did not affect oxidative phosphorylation efficiency or basal proton leak in mitochondria. We conclude that the basal proton conductance of mitochondria may regulate mitochondrial oxidative phosphorylation efficiency, but is not an important contributor to thermogenic processes in cold-acclimated ducklings.     

The ability of genetically lean or fat slow-growing chickens to synthesize and store lipids is not altered by the dietary energy source

The increasing use of unconventional feedstuffs in chicken’s diets results in the substitution of starch by lipids as the main dietary energy source. To evaluate the responses of genetically fat or lean chickens to these diets, males of two experimental lines divergently selected for abdominal fat content were fed isocaloric, isonitrogenous diets with either high lipid (80 g/kg), high fiber (64 g/kg) contents (HL), or low lipid (20 g/kg), low fiber (21 g/kg) contents (LL) from 22 to 63 days of age. The diet had no effect on growth performance and did not affect body composition evaluated at 63 days of age. Glycolytic and oxidative energy metabolisms in the liver and glycogen storage in liver and Sartorius muscle at 63 days of age were greater in chicken fed LL diet compared with chicken fed HL diet. In Pectoralis major (PM) muscle, energy metabolisms and glycogen content were not different between diets. There were no dietary-associated differences in lipid contents of the liver, muscles and abdominal fat. However, the percentages of saturated (SFA) and monounsaturated fatty acids (MUFA) in tissue lipids were generally higher, whereas percentages of polyunsaturated fatty acids (PUFA) were lower for diet LL than for diet HL. The fat line had a greater feed intake and average daily gain, but gain to feed ratio was lower in that line compared with the lean line. Fat chickens were heavier than lean chickens at 63 days of age. Their carcass fatness was higher and their muscle yield was lower than those of lean chickens. The oxidative enzyme activities in the liver were lower in the fat line than in the lean line, but line did not affect energy metabolism in muscles. The hepatic glycogen content was not different between lines, whereas glycogen content and glycolytic potential were higher in the PM muscle of fat chickens compared with lean chickens. Lipid contents in the liver, muscles and abdominal fat did not differ between lines, but fat chickens stored less MUFA and more PUFA in abdominal fat and muscles than lean chickens. Except for the fatty acid composition of liver and abdominal fat, no interaction between line and diet was observed. In conclusion, the amount of lipids stored in muscles and fatty tissues by lean or fat chickens did not depend on the dietary energy source.     

Expression of an uncoupling protein gene homolog in chickens

An avian uncoupling protein (UCP) gene homolog was recently sequenced from skeletal muscle and was proposed to have a role in thermogenesis in chickens, ducks and hummingbirds. Since mammalian UCP 2 and UCP 3 also appear to have functions associated with energy and substrate partitioning and body weight regulation, the purpose of this study was to further characterize chicken UCP under conditions of nutritional stress and/or leptin administration. Male 3-week-old chickens were starved for 24 or 48 h and then half of each group was refed for an additional 24 h. In a follow-up experiment, chickens were fed or starved for 48 h with or without leptin administration. Feed deprivation increased UCP mRNA expression in skeletal muscle by up to 260% (P<0.001), and in a time-dependent manner in pectoralis muscle. Refeeding for 24 h normalized muscle UCP mRNA levels. Leptin administration had no effect on muscle UCP. Chicken muscle UCP mRNA levels were highly correlated with plasma triglyceride and non-esterified fatty acid (NEFA) concentrations, and with circulating levels of insulin, insulin-like growth factor (IGF)-I and IGF-II. These results suggest that, as in mammals, avian UCP is up-regulated during feed deprivation and is highly correlated with increased fatty acid oxidation and flux into skeletal muscle.     

Deletion of Kinin B2 Receptor Alters Muscle Metabolism and Exercise Performance

Metabolic syndrome is a cluster of metabolic risk factors such as obesity, diabetes and cardiovascular diseases. Mitochondria is the main site of ATP production and its dysfunction leads to decreased oxidative phosphorylation, resulting in lipid accumulation and insulin resistance. Our group has demonstrated that kinins can modulate glucose and lipid metabolism as well as skeletal muscle mass. By using B2 receptor knockout mice (B2R^-/-) we investigated whether kinin action affects weight gain and physical performance of the animals. Our results show that B2R^-/- mice are resistant to high fat diet-induced obesity, have higher glucose tolerance as well as increased mitochondrial mass. These features are accompanied by higher energy expenditure and a lower feed efficiency associated with an increase in the proportion of type I fibers and intermediary fibers characterized by higher mitochondrial content and increased expression of genes related to oxidative metabolism. Additionally, the increased percentage of oxidative skeletal muscle fibers and mitochondrial apparatus in B2R^-/- mice is coupled with a higher aerobic exercise performance. Taken together, our data give support to the involvement of kinins in skeletal muscle fiber type distribution and muscle metabolism, which ultimately protects against fat-induced obesity and improves aerobic exercise performance.     

Skeletal muscle respiratory uncoupling prevents diet-induced obesity and insulin resistance in mice

To determine whether uncoupling respiration from oxidative phosphorylation in skeletal muscle is a suitable treatment for obesity and type 2 diabetes, we generated transgenic mice expressing the mitochondrial uncoupling protein (Ucp) in skeletal muscle. Skeletal muscle oxygen consumption was 98% higher in Ucp-L mice (with low expression) and 246% higher in Ucp-H mice (with high expression) than in wild-type mice. Ucp mice fed a chow diet had the same food intake as wild-type mice, but weighed less and had lower levels of glucose and triglycerides and better glucose tolerance than did control mice. Ucp-L mice were resistant to obesity induced by two different high-fat diets. Ucp-L mice fed a high-fat diet had less adiposity, lower levels of glucose, insulin and cholesterol, and an increased metabolic rate at rest and with exercise. They were also more responsive to insulin, and had enhanced glucose transport in skeletal muscle in the setting of increased muscle triglyceride content. These data suggest that manipulating respiratory uncoupling in muscle is a viable treatment for obesity and its metabolic sequelae.     

Altered development and protein metabolism in skeletal muscles of broiler chickens (Gallus gallus domesticus) by corticosterone

Two trials were conducted to investigate the effect of corticosterone (CORT) on protein metabolism and the amino acid composition in muscle tissues of broiler chickens (Gallus gallus domesticus). In Trial 1, two groups of 30 broiler chickens were subjected to control or CORT treatment (30 mg/kg diet) from 28 to 39 days of age. In Trial 2, three groups of chickens of 28 days of age were randomly subjected to one of the following treatments for 7 days: CORT (30 mg/kg diet), pair-fed (maintaining the same feed intake as CORT treatment) and control treatments. The body mass gain and feed efficiency was significantly decreased by CORT treatment, while the food intake was decreased. The breast and thigh masses (% body mass) were significantly suppressed by CORT treatment, while the abdominal fat and liver masses (%) were obviously increased. The plasma levels of glucose, urate and total amino acid were significantly elevated by CORT treatment. The capacity for protein synthesis, estimated by RNA:protein ratio, were significantly suppressed by CORT in M. pectoralis major and M. biceps femoris. The 3-methylhistidine concentrations were significantly increased in both M. pectoralis major and M. biceps femoris of CORT chickens, compared to control but not the pair-fed chickens. The amino acid composition of M. pectoralis major and M. biceps femoris was not significantly affected by CORT treatment. In conclusion, the arrested growth in skeletal muscles induced by CORT administration has tissue specificity. The CORT treatment retards the growth of skeletal muscle by suppressed protein synthesis and augmented protein catabolism.     

Upregulation of uncoupling protein-3 in skeletal muscle during exercise: a potential antioxidant function

Uncoupling protein-3 (UCP3) expression has been shown to increase dramatically in response to muscular contraction, but the physiological significance of UCP3 upregulation is still elusive. In this study, UCP3 mRNA and protein expression were investigated along with mitochondrial respiratory function, reactive oxygen species (ROS) generation, and antioxidant defense in rat skeletal muscle during and after an acute bout of prolonged exercise. UCP3 mRNA expression was elevated sharply at 45 min of exercise, reaching 7- to 8-fold above resting level at 150 min. The increase in UCP3 protein content showed a latent response but was elevated approximately 1.9-fold at 120 min of exercise. Both UCP3 mRNA and UCP3 protein gradually returned to resting levels 24 h postexercise. Mitochondrial ROS production was progressively increased during exercise. However, ROS showed a dramatic drop at 150 min although their levels remained severalfold higher during the recovery. Mitochondrial State 4 respiration rate was increased by 46 and 58% (p < 0.05) at 90 and 120 min, respectively, but returned to resting rate at 150 min, when State 3 respiration and respiratory control index (RCI) were suppressed. ADP-to-oxygen consumption (P/O) ratio and ATP synthase activity were lowered at 3 h postexercise, whereas proton motive force and mitochondrial malondialdehyde content were unchanged. Manganese superoxide dismutase gene expression was not affected by exercise except for an increase in mRNA abundance at 3 h postexercise. These data demonstrate that UCP3 expression in rat skeletal muscle can be rapidly upregulated during prolonged exercise, possibly owing to increased ROS generation. Increased UCP3 may partially alleviate the proton gradient across the inner membrane, thereby reducing further ROS production by the electron transport chain. However, prolonged exercise caused a decrease in energy coupling efficiency in muscle mitochondria revealed by an increased respiration rate due to proton leak (State 4/State 3 ratio) and decreased RCI. We thus propose that the compromise of the oxidative phosphorylation efficiency due to UCP3 upregulation may serve an antioxidant function to protect the muscle mitochondria from exercise-induced oxidative stress     

Expression of uncoupling protein-3 in subsarcolemmal and intermyofibrillar mitochondria of various mouse muscle types and its modulation by fasting.

Uncoupling protein-3 (UCP3) is a mitochondrial inner-membrane protein abundantly expressed in rodent and human skeletal muscle which may be involved in energy dissipation. Many studies have been performed on the metabolic regulation of UCP3 mRNA level, but little is known about UCP3 expression at the protein level. Two populations of mitochondria have been described in skeletal muscle, subsarcolemmal (SS) and intermyofibrillar (IMF), which differ in their intracellular localization and possibly also their metabolic role. To examine if UCP3 is differentially expressed in these two populations and in different mouse muscle types, we developed a new protocol for isolation of SS and IMF mitochondria and carefully validated a new UCP3 antibody. The data show that the density of UCP3 is higher in the mitochondria of glycolytic muscles (tibialis anterior and gastrocnemius) than in those of oxidative muscle (soleus). They also show that SS mitochondria contain more UCP3 per mg of protein than IMF mitochondria. Taken together, these results suggest that oxidative muscle and the mitochondria most closely associated with myofibrils are most efficient at producing ATP. We then determined the effect of a 24-h fast, which greatly increases UCP3 mRNA (16.4-fold) in muscle, on UCP3 protein expression in gastrocnemius mitochondria. We found that fasting moderately increases (1.5-fold) or does not change UCP3 protein in gastrocnemius SS or IMF mitochondria, respectively. These results show that modulation of UCP3 expression at the mRNA level does not necessarily result in similar changes at the protein level and indicate that UCP3 density in SS and IMF mitochondria can be differently affected by metabolic changes.     

一次性力竭运动对大鼠PHB1表达及线粒体功能的影响

目的：观察一次性力竭运动后大鼠脑、心、骨骼肌组织和线粒体中PHB1含量的变化及对大鼠线粒体功能的影响,探寻PHB1与线粒体功能和能量代谢的关系。方法：健康雄性SD大鼠40只,随机分为2组（n=20）：对照组和一次性力竭运动组,大鼠进行一次性急性跑台运动建立力竭运动模型。收集各组大鼠的心、脑和骨骼肌组织样品并提取线粒体,检测其呼吸功能和ROS的变化。用Western blot方法检测组织和线粒体中PHB1蛋白表达水平;用分光光度计检测各器官中ATP含量以及线粒体中复合体V活性（ATP合酶活性）。结果：①一次性力竭运动后脑、心肌、骨骼肌中ATP含量显著性降低;②一次性力竭运动后脑、心肌、骨骼肌线粒体中复合体V活性、RCR、ROS显著性降低,ST4均显著性升高,ST3无显著性差异。③一次性力竭运动后心、脑、骨骼肌线粒体中PHB1的表达显著性减少。④通过相关性分析得出：一次性力竭运动后心、脑、骨骼肌中ATP含量与心、脑、骨骼肌中复合体V活性呈正相关;心、脑、骨骼肌中ATP含量和心、脑骨骼肌中PHB1的表达呈正相关。结论：一次性力竭运动后,降低线粒体氧化磷酸化功能,使大鼠脑、骨骼肌线粒体内ROS生成增加,PHB1的表达、ATP含量和复合体V活性均下降。一次性力竭运动使得大鼠线粒体内PHB1表达降低,线粒体功能减弱,机体能量代谢降低。     

Recombinant human uncoupling protein-3 increases thermogenesis in yeast cells

The long form of human uncoupling protein-3 (hUCP3L) is highly homologous to thermogenin (UCPI), the uncoupling protein of brown fat mitochondria, but is expressed predominantly in skeletal muscle. Its putative role is to regulate the coupling efficiency of oxidative phosphorylation and thus thermogenesis in skeletal muscle, a major thermogenic tissue in higher mammals. To study the functional relevance of hUCP3L, the protein was expressed in yeast cells under the control of the galactose promoter. Expression of hUCP3L induced a series of phenotype changes in the yeast cells. The cellular growth and the mitochondrial membrane potential were both diminished. The portion of cellular respiration coupled to oxidative phosphorylation decreased from 57% to 11% (P<0.001) and the cellular heat production, as measured by direct microcalorimetry, was increased by 33.3 +/- 3.2% (P<0.001) after induction of UCP3L. These observations demonstrate for the first time the intrinsic thermogenic properties of hUCP3L in intact cells.     

Distribution of the ATPase inhibitor proteins of mitochondria in mammalian tissues including fibroblasts from a patient with Luft's disease.

The mitochondrial ATPase inhibitor proteins--the Pullman-Monroy inhibitor (PMI) and the Ca(2+)-binding protein (CaBI)--have a wide distribution, both being present in mitochondria of bovine heart and kidney, rat liver and brain, two mitochondrial populations of rabbit skeletal muscle, and mitochondria from human fibroblasts and the human breast cancer cell line T-47-D. The ratio of CaBI to PMI was highest in heart and skeletal muscle mitochondria. The subsarcolemmal fraction of skeletal muscle had 2.6-times as much CaBI as did the intermyofibrillar. The ratio of CaBI to PMI in the mitochondria of the other normal tissues and fibroblasts was close to 1. In contrast, mitochondria from T-47D cells had 1.5-times as much PMI as CaBI whilst mitochondria from fibroblasts from a patient with Luft's disease showed a virtual lack of PMI. The specific ATPase, ATP-synthetase and succinate dehydrogenase activities of the Luft's mitochondria were, however, in the normal range. The specific ATP synthetase activity of the T-47D cells was significantly higher than normal. We conclude that tissues like heart and skeletal muscle which experience wide fluctuations in intracellular Ca2+ have a greater need for CaBI. Why lack of PMI could lead to 'loose' coupling of oxidative phosphorylation in skeletal muscle of Luft's patients, but not in fibroblasts is discussed.