The mitochondrial uncoupling protein (UCP) in the mitochondrial inner membrane of mammalian brown adipose tissue generates heat by uncoupling oxidative phosphorylation. This process protects against cold and regulates energy balance. Manipulation of thermogenesis could be an effective strategy against obesity. Here we determine the role of UCP in the regulation of body mass by targeted inactivation of the gene encoding it. We find that UCP-deficient mice consume less oxygen after treatment with a beta3-adrenergic-receptor agonist and that they are sensitive to cold, indicating that their thermoregulation is defective. However, this deficiency caused neither hyperphagia nor obesity in mice fed on either a standard or a high-fat diet. We propose that the loss of UCP may be compensated by UCP2, a newly discovered homologue of UCP; this gene is ubiquitously expressed and is induced in the brown fat of UCP-deficient mice.

The mRNA levels for the mitochondrial uncoupling protein (UCP1) in fat tissues of A/J and C57BL/6J inbred strains of mice varied in a regional-specific manner after stimulation of adrenergic signaling by cold exposure or treatment with a ␤ 3-adrenergic agonist. While the differences between strains were minimal in interscapular brown fat, large differences occurred in white fat tissues, particularly in retroperitoneal fat. Among the AXB recombinant inbred strains, the Ucp1 mRNA levels varied up to 130-fold. This large induction at the mRNA level was accompanied by a corresponding increase in brown adipocytes as revealed by immunohistology with anti-UCP1 antibodies. A high capacity to induce brown fat in areas of traditional white fat had no impact on the ability to gain weight in response to high fat and sucrose diets, but did correlate with the loss of weight in response to treatment with a ␤ 3-adrenergic agonist (CL 316,243). This genetic variation in mice provides an experimental approach to identify genes controlling the induction of brown adipocytes in white fat tissues.

A forkhead-type transcription factor, DAF-16, is located in the most downstream part of the insulin signalling pathway via PI3K (phosphoinositide 3-kinase). It is essential for the extension of life-span and is also involved in dauer formation induced by food deprivation in Caenorhabditis elegans. In the present study, we addressed whether or not FOXO members AFX, FKHR (forkhead homologue in rhabdomyosarcoma) and FKHRL1 (FKHR-like protein 1), mammalian counterparts of DAF-16, are involved in starvation stress. We found a remarkable selective induction of FKHR and FKHRL1 transcripts in skeletal muscle of mice during starvation. The induction of FKHR gene expression was observed at 6 h after food deprivation, peaked at 12 h, and returned to the basal level by 24 h of refeeding. The induction was also found in skeletal muscle of mice with glucocorticoid treatment. Moreover, we found that the levels of PDK4 (pyruvate dehydrogenase kinase 4) gene expression were up-regulated through the direct binding of FKHR to the promoter region of the gene in C2C12 cells. These results suggest that FKHR has an important role in the regulation of energy metabolism, at least in part, through the up-regulation of PDK4 gene expression in skeletal muscle during starvation.

Sympathetic stimulation activates glucose utilization in parallel with fatty acid oxidation and thermogenesis in brown adipose tissue (BAT) through the beta-adrenergic receptors. To clarify the roles of the principal thermogenic molecule mitochondrial uncoupling protein 1 (UCP1) in the sympathetically stimulated glucose utilization, we investigated the uptake of 2-deoxyglucose (2-DG) into BAT and some other tissues of UCP1-knockout (KO) mice in vivo. In wild-type (WT) mice, administration of norepinephrine (NE) accelerated the disappearance of plasma 2-DG and increased 2-DG uptake into BAT and heart without any rise of plasma insulin level. In UCP1-KO mice, the stimulatory effect of NE on 2-DG uptake into BAT, but not into heart, disappeared completely. Insulin administration increased 2-DG uptake into BAT and also heart similarly in WT and UCP1-KO mice. NE also increased the activity of AMP-activated protein kinase (AMP kinase) in BAT of WT but not UCP1-KO mice. Our results, together with reports that the activation of AMP kinase increases glucose transport in myocytes, suggest that the sympathetically stimulated glucose utilization in BAT is due to the serial activation of UCP1 and AMP kinase.

SummaryLoss of nonshivering thermogenesis in mice by inactivation of the mitochondrial uncoupling protein gene ( Ucp1 -/-mice) causes increased sensitivity to cold and unexpected resistance to diet-induced obesity at a young age. To clarify the role of UCP1 in body weight regulation throughout life and influence of UCP1 deficiency on longevity, we longitudinally analyzed the phenotypes of Ucp1 -/-mice maintained in a room at 23 °°°° C. There was no difference in body weight and lifespan between genotypes under the standard chow diet condition, whereas the mutant mice developed obesity with age under the high-fat (HF) diet condition. Compared with Ucp1 +/+ mice, Ucp1 -/-mice showed increased expression of genes related to thermogenesis and fatty acid metabolism, such as β β β β 3-adrenergic receptor, in adipose tissues of the 3-month-old mutants; however, the augmented expression was reduced in Ucp1 +/+ mice in 11-month-old Ucp1 -/-mice fed the HF diet. Likewise, the increased levels of UCP3 and cAMPdependent protein kinase in the brown adipose tissue of Ucp1 -/-mice given the standard diet were decreased significantly in that of Ucp1 -/-mice fed the HF diet, which animals showed impaired norepinephrine-induced lipolysis in their adipose tissues. These results suggest profound attenuation of β β β β -adrenergic responsiveness and fatty acid utilization in Ucp1 -/-mice fed the HF diet, bringing them to late-onset obesity. Our findings provide evidence that UCP1 is neither essential for body weight regulation nor for longevity under conditions of standard diet and normal housing temperature, but deficiency increases susceptibility to obesity with age in combination with HF diet.

. Indispensable role of mitochondrial UCP1 for antiobesity effect of ␤ 3-adrenergic stimulation. Am J Physiol Endocrinol Metab 290: E1014 -E1021, 2006. First published December 20, 2005 doi:10.1152/ajpendo.00105.2005.-Mitochondrial uncoupling protein-1 (UCP1) has been thought to be a key molecule for thermogenesis during cold exposure and spontaneous hyperphagia and thereby in the autonomic regulation of energy expenditure and adiposity. However, UCP1 knockout (KO) mice were reported to be cold intolerant but unexpectedly did not get obese even after hyperphagia, implying that UCP1 may not be involved in the regulation of adiposity. Treatment of obese animals with ␤ 3-adrenergic agonists is known to increase lipid mobilization, induce UCP1, and, finally, reduce body fat content. To obtain direct evidence for the role of UCP1 in the anti-obesity effect of ␤ 3-adrenergic stimulation, in the present study, UCP1-KO and wild-type (WT) mice were fed on cafeteria diets for 8 wk and then given a ␤ 3-adrenergic agonist, CL-316,243 (CL), or saline for 2 wk. A single injection of CL increased whole body oxygen consumption and brown fat temperature in WT mice but not in KO mice, and it elicited almost the same plasma free fatty acid response in WT and KO mice. WT and KO mice increased similarly their body and white fat pad weights on cafeteria diets compared with those on laboratory chow. Daily treatment with CL resulted in a marked reduction of white fat pad weight and the size of adipocytes in WT mice, but not in KO mice. Compared with WT mice, KO mice expressed increased levels of UCP2 in brown fat but decreased levels in white fat and comparable levels of UCP3. It was concluded that the anti-obesity effect of ␤ 3-adrenergic stimulation is largely attributable to UCP1, but less to UCP2 and UCP3, and thereby to UCP1-dependent degradation of fatty acids released from white adipose tissue.uncoupling protein-1; adiposity; CL-316,243; energy expenditure; hyperphagia UNCOUPLING PROTEIN (UCP) IS A MOLECULE, as its name suggests, that uncouples mitochondrial oxidative phosphorylation by bypassing the electrochemical gradient across the inner membrane from the F1-ATPase and thereby dissipates energy as heat. Among several isoforms of the UCP family so far reported in mammals, UCP1 is the only one whose physiological importance has been firmly established; that is, UCP1 is present exclusively in brown adipose tissue (BAT), an organ specified for nonshivering thermogenesis during cold acclimation, arousal from hibernation, and recovery from anesthetic hypothermia (2). UCP1 has also been proposed to be involved in diet-induced thermogenesis, as well as cold-induced nonshivering thermogenesis, and play a significant role in the control of energy expenditure and whole body energy balance. This is supported by the observations, for example, that spontaneous overfeeding of highly palatable diets and/or high-fat diets gives rise to increased energy expenditure (oxygen consumption) in association with BAT hyperplasia and increased UCP1 c...

1. The present study was undertaken to investigate the effects of hypobaric hypoxia, equivalent to an altitude of 5500 m, on antioxidant enzymes in rats. 2. Malondialdehyde levels in serum, heart, lung, liver and kidney of hypobaric-hypoxic rats were all significantly higher than in control rats by day 21 of exposure (P < 0 05), indicating increased oxidative stress. 3. Superoxide dismutase (SOD) catalyses the conversion of the superoxide anion to H202 and 02 The concentration of immunoreactive Mn-SOD in the serum of hypobaric-hypoxic rats was raised significantly from day 5 onwards, whereas in liver and lung, it had decreased significantly by day 21 (P < 0 05). 4. Glutathione peroxidase (GSH-Px) catalyses H202 and certain lipid peroxides. By day 21, GSH-Px activity had increased significantly in the heart and lungs, but decreased significantly in the liver (P < 0'05). 5. Catalase catalyses H202. Catalase activity in the liver and kidney of hypobaric-hypoxic rats was significantly decreased on day 1 (P < 0 05) though levels then recovered. 6. Mn-SOD mRNA in the liver of hypobaric-hypoxic rats was induced during the experiment, the effect being exceptionally marked, especially during the first 3 days of exposure to hypobaric hypoxia. 7. These results suggest that the liver may be more vulnerable than the other organs tested to oxidative stress under hypobaric hypoxia.Oxidative stress is an important cause of in-cell damage. Consequently, numerous studies have been published on the effects on the antioxidant systems of conditions in which the level of tissue oxygenation is changed such as: (i) hypoxia

In C. elegans, insulin-like hormone signal pathway plays a significant role in longevity. In particular, daf-16 gene product is indispensable factor for this lifespan-extension. This signal pathway is critical for dauer formation, which is a similar state to hibernation in mammals. We examined the expression level of mammalian daf-16 homologues, Foxo 1,3, and 4 (FKHR, FKHRL1, and AFX) mRNAs in the rat skeletal muscles during aging and in 30% caloric restricted of ad libitum fed. The expression level of AFX mRNA was significantly higher at 6 and 12 months than at 3 and 26 months, and FKHRL1 expression was significantly higher at 6 months than at 3 and 26 months but FKHR expression showed no significant change with age. We observed a characteristic expression of AFX and FKHR mRNAs to be significantly higher in the second day in caloric restriction by every-other-day feeding than in ad libitum fed. This suggests that caloric restriction may increase the expression of FKHR-family genes and prevent the aging process in the skeletal muscles.