NEW MEMBERS OF UNCOUPLING PROTEIN FAMILY IMPLICATED IN ENERGY METABOLISM

1. The regulation of energy metabolism involves food intake and energy expenditure. Mitochondrial uncoupling proteins (UCP) are implicated in energy expenditure. 2. cDNA of a homologue of UCP highly expressed in rat skeletal muscle, UCP-3, is isolated and sequenced. Rat UCP-2 cDNA is also isolated and sequenced. 3. Rat UCP-3 cDNA probe hybridized two bands, a major band at 2.5 kb and a minor band at 2.8 kb in rat tissues. The mRNA was expressed at the highest level in the skeletal muscle, and moderately in the interscapular brown adipose tissue (BAT). Only weak signals were detected in the epididymal white adipose tissue (WAT) and the heart. Rat UCP-2 cDNA probe hybridized a 1.7 kb band detected widely in the whole body, especially abundant in the lung and the spleen. In contrast to the UCP-3 gene expression, the UCP-2 gene expression was expressed at substantial levels in the WAT and only at slight levels in the skeletal muscle and BAT. 4. The UCP-3 gene expression is augmented two-fold in the gastrocnemius muscle from rats fed a high-fat diet (P < 0.05). The UCP-3 mRNA levels remained unchanged in the interscapular BAT, and epididymal WAT. The levels of the UCP-2 gene expression are augmented significantly in the epididymal WAT (1.6-fold; P < 0.05), while no significant increase is observed in the gastrocnemius muscle and interscapular BAT.     

DNA microarray analysis of white adipose tissue from obese (fa/fa) Zucker rats treated with a beta3-adrenoceptor agonist, KTO-7924.

We aimed to examine the effects of KTO-7924 (beta3-adrenoceptor agonist) on lipid metabolism and mRNA expressions in retroperitoneal white adipose tissue (RP WAT) in obese (fa/fa) Zucker rats using DNA microarray. Oral KTO-7924 for 28 days significantly decreased RP WAT weight, plasma triglyceride, free fatty acid, and insulin, and improved insulin resistance in oral glucose tolerance tests. In RP WAT of KTO-7924-treated rats, DNA microarray analysis revealed specifically enhanced mRNA expressions of uncoupling protein 1 (UCP1) and cytochrome c oxidase subunit VIII-H (COX8H), which are reportedly highly expressed in brown adipose tissue (BAT). Since these mRNA expression levels in RP WAT were significantly lower in obese (fa/fa) Zucker rats than in lean Zucker rats, these genes may be important in lipid metabolism. Our results imply that in obese (fa/fa) Zucker rats, continuous stimulation of beta3-adrenoceptors by KTO-7924 causes BAT-like adipocytes to appear in RP WAT, and improves lipid metabolism.     

Induction of uncoupling protein-1 and -3 in brown adipose tissue by kaki-tannin in type 2 diabetic NSY/Hos mice

Kaki-tannin, a highly polymerized-tannin from the young fruits of persimmon (Diospyros kaki 'Hachiya'), has been shown to have bile acid-binding activity. To verify the effect of kaki-tannin on the metabolism of lipid and glucose in type 2 diabetes, type 2 diabetic NSY/Hos mice were fed an AIN76-modified high fat diet supplemented with 1% (w/w) kaki-tannin for 8weeks. Kaki-tannin induced a 2-fold increase in fecal bile acid excretion and was significantly effective in the prevention of a rise in plasma cholesterol, triglyceride, and insulin levels. Kaki-tannin treatment also prevented fatty liver. To identify the molecular mechanism underlying these effects, gene expression analysis was performed on liver, brown adipose tissue (BAT), and skeletal muscle. The genes related to cholesterol metabolism, including 3-hydroxy-3-methylglutaryl-coenzyme A reductase and sterol regulatory element-binding protein 2, were increased in the liver of the kaki-tannin group. Interestingly, the uncoupling protein-1 (UCP1) gene and the UCP3 gene were significantly increased in the BAT of the kaki-tannin group, which was also confirmed at the protein level. These findings indicated that induction of UCP1 and UCP3 in the BAT by kaki-tannin treatment might influence the energy metabolism, thus contributing beneficial effects to type 2 diabetic NSY/Hos mice.     

Peripheral injection of risperidone,an atypical antipsychotic,alters the bodyweight gain of rats

1. Risperidone is an atypical antipsychotic drug that possesses 5-hydroxytryptamine 5-HT2 receptor antagonism combined with milder dopamine D2 receptor antagonism. 2. Excessive bodyweight gain is one of the side-effects of antipsychotics. Risperidone treatment causes a greater increase in the body mass of patients than treatment with conventional antipsychotics, such as haloperidol. Therefore, the present study was undertaken in order to address the aetiology of the risperidone-induced bodyweight change in rats by examining the expression of leptin, an appetite-regulating hormone produced in white adipose tissue (WAT), and uncoupling protein (UCP)-1, a substance promoting energy expenditure in the brown adipose tissues (BAT). 3. Eight-week-old male rats were injected subcutaneously with risperidone (0.005, 0.05 or 0.5 mg/kg) twice daily for 21 days. Both bodyweight and food intake were monitored daily. On day 21, rats were decapitated and their serum leptin and prolactin concentrations were measured. Expression levels of leptin, Ucp1 and beta3-adrenoceptor (beta3-AR) genes in WAT and BAT were quantified using real-time polymerase chain reaction amplification. 4. Injection of 0.005 mg/kg risperidone into rats increased food intake and the rate of bodyweight gain, as well as the augmentation of leptin gene expression in WAT. Injection of 0.05 mg/kg risperidone increased food intake and leptin gene expression in WAT, but the rate of bodyweight gain was not affected. Injection of 0.5 mg/kg risperidone caused a reduction in bodyweight gain, as well as enhanced Ucp1 gene expression in BAT and serum prolactin concentrations. The serum leptin concentration and beta3-AR gene expression in WAT and BAT were not affected by injection of 0.5 mg/kg risperidone. 5. Although the changes in food intake observed in risperidone-injected rats were rationalized neither by serum leptin nor prolactin concentrations, the reduction in the rate of bodyweight gain following injection of 0.5 mg/kg can be explained, in part, by increased energy expenditure, as revealed by the remarkable increase in the UCP-1 mRNA expression level in BAT. The role of leptin in risperidone-induced alterations in bodyweight gain remain to be clarified.     

microRNA-32 promotes white fat browning through augmenting FGF21 expression in brown adipose tissue

OBJECTIVE To investigate the effect of microRNA-32 on cold-induced thermogenesis and brown adipocyte energy metabolism.METHODS To apply the cold-induced thermogenesis model in mice,8-10 week old male C57Bl6 mice were placed within a 6℃fridge for 7d.Control microRNA inhibitor or miR-32 inhibitor(10mg·kg-1)was administered via intraperitoneal injection 16 hbefore the mice were placed in the fridge.Daily core body temperatures were taken using a rectal temperature probe.Mice were euthanized after 7dand brown adipose tissue(BAT),inguinal and epididymal white adipose tissue(WAT),skeletal muscle and liver tissue analysed for changes in morphology and gene expression.RESULTS miR-32 inhibition in vivoinhibits the emergence of beige cells,which function like BAT cells,within WAT.In silico prediction and gene ontology analysis identified Tob1 as a likely target gene of miR-32.miR-32 inhibition led to increased expression of Tob1 whilst mutation of target sequence abolished this effect.Expression of brown adipose markers such as Ucp1,Pgc1α,Pparαand Prdm16 were significantly reduced in inguinal white adipose tissue(P<0.05).There was also a significant decrease in serumfgf21 levels due to the inhibition of Fgf21 expression in BAT(P<0.05).p38/MAPK signalling in brown adipose tissue was also significantly inhibited within brown adipose tissue leading to decreased fgf21 expression and secretion.CONCLUSION Our study shows that miR-32 plays a crucial role in stimulating beige cell emergence by activating p38/MAPK signalling during cold thermogenesis.miR-32 may prove effective as a treatment for obesity by activating cold-induced thermogenesis leading to increased energy metabolism.     

The H₁-H₂ domain of the α₁ isoform of Na+-K+-ATPase is involved in ouabain toxicity in rat ventricular myocytes

Metals and metalloid species are involved in homeostasis in energy systems such as glucose metabolism. Enlarged adipocytes are one of the most important causes of obesity-associated diseases. In this study, we studied the possibility that various metals, namely, CoCl₂, HgCl₂, NaAsO₂ and MnCl₂ pose risk to or have beneficial effects on white adipose tissue (WAT). Exposure to the four metals resulted in decreases in WAT weight and the size of enlarged adipocytes in mice fed a high-fat diet (HFD) without changes in liver weight, suggesting that the size and function of adipocytes are sensitive to metals. Repeated administration of CoCl₂ significantly increased serum leptin, adiponectin and high-density lipoprotein (HDL) cholesterol levels and normalized glucose level and adipose cell size in mice fed HFD. In contrast, HgCl₂ treatment significantly decreased serum leptin level with the down-regulation of leptin mRNA expression in WAT and a reduction in adipocyte size. Next, we tried to investigate possible factors that affect adipocyte size. Repeated exposure to HgCl₂ significantly decreased the expression levels of factors upon the regulation of energy such as the PPARα and PPARγ mRNA expression levels in adipocytes, whereas CoCl₂ had little effect on those genes expressions compared with that in the case of the mice fed HFD with a vehicle. In addition, repeated administration of CoCl₂ enhanced AMPK activation in a dose-dependent manner in the liver, skeletal muscle and WAT; HgCl₂ treatment also enhanced AMPK activation in the liver. Thus, both Co and Hg reduced WAT weight and the size of enlarged adipocytes, possibly mediated by AMKP activation in the mice fed HFD. However, inorganic cobalt may have a preventive role in obesity-related diseases through increased leptin, adiponectin and HDL-cholesterol levels, whereas inorganic mercury may accelerate the development of such diseases. These results may lead to the development of new approaches to establishing the role of metals in adipose tissue of obesity-related diseases.     

Expression profiles of three isoforms of inositol 1,4,5-trisphosphate receptor in brown adipose tissue of the rat

The thermogenic function of brown adipose tissue (BAT) is known to be mainly regulated by a signal transduction cascade via beta-adrenoceptor. However, recent studies indicated that the alpha-adrenoceptor and its downstream signal transduction cascade, causing elevation of the cellular Ca(2+) level, are also important for the regulation of this function of BAT. In the present study, expression profiles of 3 isoforms of the inositol 1,4,5-trisphosphate (IP(3)) receptor, known as one of the major components of the machinery regulating the intracellular Ca(2+) concentration in the BAT of rats, were investigated by Northern analysis. Of these three IP(3) receptor isoforms, the type 2 one was found to be the most abundant of the three in BAT. Furthermore, when rats were exposed to the cold, under which condition the thermogenic function of BAT is known to be stimulated, the expression levels of types 1 and 2 isoforms of IP(3) receptor were remarkably elevated. The results of Western analysis supported the predominant expression of the type 2 isoform in BAT. However, different from the results of Northern analysis, the expression levels of types 1 and 2 isoforms of IP(3) receptor protein in BAT were not influenced by exposure of the animals to the cold.     

The phenotype of a flavin-containing monooyxgenase knockout mouse implicates the drug-metabolizing enzyme FMO1 as a novel regulator of energy balance

Flavin-containing monooxygenases (FMOs) of mammals are thought to be involved exclusively in the metabolism of foreign chemicals. Here, we report the unexpected finding that mice lacking Fmos 1, 2 and 4 exhibit a lean phenotype and, despite similar food intake, weigh less and store less triglyceride in white adipose tissue (WAT) than wild-type mice. This is a consequence of enhanced whole-body energy expenditure, due mostly to increased resting energy expenditure (REE). This is fuelled, in part, by increased fatty acid β-oxidation in skeletal muscle, which would contribute to depletion of lipid stores in WAT. The enhanced energy expenditure is attributed, in part, to an increased capacity for exercise. There is no evidence that the enhanced REE is due to increased adaptive thermogenesis; instead, our results are consistent with the operation in WAT of a futile energy cycle. In contrast to FMO2 and FMO4, FMO1 is highly expressed in metabolic tissues, including liver, kidney, WAT and BAT. This and other evidence implicates FMO1 as underlying the phenotype. The identification of a novel, previously unsuspected, role for FMO1 as a regulator of energy homeostasis establishes, for the first time, a role for a mammalian FMO in endogenous metabolism. Thus, FMO1 can no longer be considered to function exclusively as a xenobiotic-metabolizing enzyme. Consequently, chronic administration of drugs that are substrates for FMO1 would be expected to affect energy homeostasis, via competition for endogenous substrates, and, thus, have important implications for the general health of patients and their response to drug therapy.     

Expression of multiple Transient Receptor Potential channel genes in murine 3T3-L1 cell lines and adipose tissue

BackgroundCalcium and its signaling have a role in adipogenesis. Transient Receptor Potential (TRP) channels are non-selective cation channels with a high permeability to calcium.MethodsIn the present study the expression of multiple TRP channels on mouse 3T3-L1 preadipocyte and adipocyte cells, white (WAT) and brown (BAT) adipose tissues was investigated using real time PCR (RT-PCR).ResultsTRPV1, TRPV3, TRPM8, TRPC4, TRPC6 were differentially expressed in preadipocytes and adipocytes suggesting their significance in adipogenesis. Genes for multiple TRP channels were also expressed in murineWAT and BAT, out of which TRPV4, TRPV6 and TRPC6 showed differential expression.ConclusionPresent study demonstrates the expression of TRP channels in mouse cell lines and adipose tissues.     

Ablation of steroid receptor coactivator-3 in mice impairs adipogenesis and enhances energy expenditure

Obesity is a medical condition in which excess body fat has accumulated to an extent and may have an adverse effect on health, leading to reduced life expectancy, impaired energy homeostasis and increased health problems. The p160 steroid receptor coactivator (SRC) gene family members have been suggested to be involved in energy homeostasis, but the impact of SRC-3 ablation on white and brown adipose tissue needs to be elucidated. In the current study, we collected in vivo data and carried out morphological studies on the effect of SRC-3 deficiency on white adipose tissue (WAT) and brown adipose tissue (BAT). Primary cells were cultured to investigate the differentiation ability of both adipocytes. Western blot was applied to detect the expression of master genes governing adipogenesis and thermogenesis. We observed that SRC-3−/− mice were lean, with reduced WAT and decreased serum leptin levels, mainly due to the smaller white adipocyte size caused by impaired adipogenesis, presented by decreased peroxisome proliferator activated receptor γ (PPARγ) expression. In the BAT, the lipid droplets decreased significantly in SRC-3−/− mice as demonstrated by histological analysis and electron microscopic observation, which could be explained by enhanced thermogenesis. The expression of thermogenic marker gene PPARγ coactivator 1α and uncoupling protein-1 increased in BAT of SRC-3−/− mice, which proved our observations. Collectively, these results demonstrate that SRC-3 plays a key role in adipogenesis and energy expenditure.     

甲状腺素对大鼠脂肪组织解偶联蛋白 2基因表达的影响

目的：探讨甲状腺素对大鼠白色脂肪组织(WAT)解偶联蛋白 2(UCP2)基因表达的影响.方法：正常雄性Wistar大鼠30只,随机分为3组,对照组、甲状腺功能亢进(甲亢)组和甲状腺功能低下(甲低)组,每组10只,应用左甲状腺素钠和甲巯咪唑分别造成大鼠甲亢和甲低状态;用放射免疫法检测血清总三碘甲状腺原氨酸(T3)、甲状腺素(T4)的浓度;半定量RT PCR法检测WAT中UCP2 mRNA的表达水平.结果：与对照组相比,甲亢组大鼠WAT中UCP2 mRNA的水平增加1.4倍,甲低组大鼠WAT中UCP2 mRNA的水平降低20%(均P＜0.01).结论：甲状腺素对WAT中UCP2基因的表达具有上调作用     

Resistance to excessive bodyweight gain in risperidone-injected rats

1. The present study was carried out to explain the resistance of rats injected subcutaneously with risperidone, the atypical antipsychotic drug, for 21 consecutive days at 0.1 mg/kg per day (a dose equivalent to the one used for patients) to result in an excessive bodyweight despite the increase in diet-uptake in rats against risperidone-induced decrease in body temperature. 2. Rectal temperature measurements were made in 8-week-old male Sprague-Dawley rats maintained under standard laboratory conditions using a 12 h daylight cycle. A s.c. injection of risperidone (0.05 mg/kg) produced hypothermia in rats, which was observed during the daily injection for 21 consecutive days. 3. Sera, white and brown adipose tissues, skeletal muscle and liver were extracted from 8-week-old male Sprague-Dawley rats injected subcutaneously with risperidone (0.01 or 0.1 mg/kg per day) or a vehicle for 21 consecutive days. Serum levels of lipids, ketones and thyroid hormone were measured. The mRNA expression levels in these tissues and organs of the genes encoding the substances involved in heat production and/or lipid metabolism were investigated by using quantitative real-time polymerase chain reaction amplification. 4. Serum nonesterified fatty acid levels in risperidone 0.1 mg/kg per day s.c. injected rats were significantly lower than those in vehicle-injected ones. Serum beta-hydroxybutyrate levels in risperidone-injected rats tended to decrease compared with those in vehicle-injected ones. The serum level of neither triiodothyronine nor thyroxine was affected by risperidone s.c. injection at the doses examined, although their values were within normal limits. 5. Risperidone injection (0.1 mg/kg per day) for 21 consecutive days upregulated mRNA expressions in white adipose tissue of uncoupling protein 3 which dissipates energy as heat; peroxisome proliferator-activated receptor (PPAR) gamma coactivator 1alpha which activates mitochondrial biogenesis to expand the oxidative machinery; and PPARalpha which is necessary for the fat-depletion of adipocytes for thermogenesis. The mRNA of lipogenic enzymes (acetyl-CoA carboxylase alpha, fatty-acid synthase and glycerol-3-phosphate acyltransferase), hormone sensitive lipase and beta1-adrenoceptor were also enhanced in white adipose tissue by the injection of 0.1 mg/kg per day risperidone. 6. These findings suggest that the materials for heat generation in white adipose tissue would be readily supplied, which in turn would reduce a storage of lipids in white adipose tissue resulting in the lower rate of bodyweight gain of rats.     

ANTI-OBESITY AND ANTI-DIABETIC EFFECTS OF CARTEOLOL IN NON-INSULIN-DEPENDENT DIABETIC MICE

1. When carteolol, a β-adrenergic blocker, was administered to KK-A^y/Ta Jcl mice that are obese and develop spontaneously non-insulin dependent diabetes, their increase in bodyweight was arrested from the age of 16 weeks. Since their intake of food and water was not influenced by carteolol treatment, compared with the control KK-A^y/Ta Jcl mice, abolition of the weight gain might be attributed to increased energy metabolism. 2. Non-fasting serum glucose levels in carteolol-treated mice at the age of 17 weeks were within normal range (118±4 vs 186±12 mg/dL). An intraperitoneal glucose-tolerance test revealed that the carteolol treatment markedly restored glucose metabolism; fasting plasma glucose (88±6 mg/dL) was within normal range, and immunoreactive insulin (IRI; 5.8±0.8 vs 33.3 ± 10.5 ng/mL) and plasma glucose levels at 60 min post glucose (361±44 vs 541 ±32 mg/dL) were significantly lower in carteolol-treated mice than those in the control group at the age of 20 weeks. 3. From these findings, carteolol is considered to have little effect on the growth of mice but to correct the obesity that develops after age 16 weeks, when their growth terminates. In addition, the normalization of blood glucose and marked decrease in IRI levels suggests that carteolol improves glucose tolerance by increasing the insulin sensitivity. 4. Since brown adipose tissue (BAT) is closely associated with thermogenesis and energy consumption, we tested whether carteolol may affect BAT, When the regional blood flow was measured by radioactive microspheres in rats, blood flow in BAT and white adipose tissue was markedly increased by carteolol. 5. These findings indicate that carteolol blocks β₁- and β₂-adrenoceptors, but may stimulate β-receptors particularly in the adipose tissue to promote lipolysis and thermogenesis, and to consume excess energy in mice. Thus, carteolol does not influence mouse growth, but may prevent obesity leading to increases in insulin sensitivity.     

The tumor suppressors pRB and p53 as regulators of adipocyte differentiation and function

Background: The retinoblastoma protein (pRB) and p53 are crucial members of regulatory networks controlling the cell cycle and apoptosis, and a hallmark of virtually all cancers is dysregulation of expression or function of pRB or p53. Although they are best known for their role in cancer development, it is now evident that both are implicated in metabolism and cellular development. Objective/methods: To review the role of pRB and p53 in adipocyte differentiation and function emphasizing that pRB and p53, via their effects on adipocyte development and function, play a role in the regulation of energy metabolism and homeostasis. Results/conclusions: pRB is required for adipose conversion and also involved in determining its mitochondrial capacity. p53 inhibits adipogenesis and results suggest that it is involved in maintaining function of adipose tissue.     

棕色脂肪白色化研究进展

棕色脂肪组织可以通过产生热量来消耗储存的化学能。棕色脂肪白色化会损害它的产热功能造成肥胖和代谢紊乱相关的疾病,减缓或抑制棕色脂肪白色化进程具有重要意义。该文综述了棕色脂肪白色化的诱导因素及关键调控因子,以期为肥胖及代谢紊乱相关疾病的预防和治疗提供新思路。     

Cardiolipin profiles as a potential biomarker of mitochondrial health in diet‐induced obese mice subjected to exercise,diet‐restriction and ephedrine treatment

Cardiolipin (CL) is crucial for mitochondrial energy metabolism and structural integrity. Alterations in CL quantity or CL species have been associated with mitochondrial dysfunction in several pathological conditions and diseases, including mitochondrial dysfunction‐related compound attrition and post‐market withdrawal of promising drugs. Here we report alterations in the CL profiles in conjunction with morphology of soleus muscle (SM) and brown adipose tissue (BAT) in diet‐induced obese (DIO) mice, subjected to ephedrine treatment (EPH: 200 mg kg^–1 day^–1 orally), treadmill exercise (EX: 10 meters per min, 1 h per day), or dietary restriction (DR: 25% less of mean food consumed by the EX group) for 7 days. Mice from the DR and EPH groups had a significant decrease in percent body weight and reduced fat mass compared with DIO controls. Morphologic alterations in the BAT included brown adipocytes with reduced cytoplasmic lipid droplets and increased cytoplasmic eosinophilia in the EX, DR and EPH groups. Increased cytoplasmic eosinophilia in the BAT was ultrastructurally manifested by increased mitochondrial cristae, fenestration of mitochondrial cristae, increased electron density of mitochondrial matrix, and increased complexity of shape and elongation of mitochondria. Mitochondrial ultrastructural alterations in the SM of the EX and DR groups included increased mitochondrial cristae, cup‐shaped mitochondria and mitochondrial degeneration. All four CL species (tri‐linoleoyl‐mono‐docosahexaenoyl, tetralinoleoyl, tri‐linoleoyl‐mono‐oleoyl, and di‐linoleoyl‐di‐oleoyl) were increased in the BAT of the DR and EPH groups and in the SM of the EPH and EX groups. In conclusion, cardiolipin profiling supported standard methods for assessing mitochondrial biogenesis and health, and may serve as a potential marker of mitochondrial dysfunction in preclinical toxicity studies. Copyright © 2014 John Wiley & Sons, Ltd.     

Ovarian hormones influence brown adipose tissue

Adult female rats were ovariectomized (OVX) or sham-operated and 4-5 weeks later OVX groups were treated with estradiol benzoate (EB), progesterone, both hormones, or the oil vehicle. All rats were sacrificed on the 4th day of hormone treatment following an overnight fast and a terminal meal. Interscapular brown adipose tissue (BAT) pads of EB-treated groups were heavier and contained more lipid than those of the other OVX groups. Lipid content of adipose tissue differed according to site (BAT less than inguinal less than parametrial = retroperitoneal), but only BAT exhibited differential responsiveness to hormonal treatments. There was also a trend for increased oxygen consumption by BAT from EB-treated rats. It is concluded that BAT may be involved in the process of increased energy expenditure by estrogen-treated rats.