Endothelial cell apoptosis in brown adipose tissue of rats induced by hyperinsulinaemia: the possible role of TNF-α

The aim of the present study was to investigate whether hyperinsulinaemia, which frequently precedes insulin resistance syndrome (obesity, diabetes), induces apoptosis of endothelial cells (ECs) in brown adipose tissue (BAT) and causes BAT atrophy and also, to investigate the possible mechanisms underlying ECs death. In order to induce hyperinsulinaemia, adult male rats of Wistar strain were treated with high dose of insulin (4 U/kg, intraperitonealy) for one or three days. Examinations at ultrastructural level showed apoptotic changes of ECs, allowing us to point out that changes mainly but not exclusively, occur in nuclei. Besides different stages of condensation and alterations of the chromatin, nuclear fragmentation was also observed. Higher number of ECs apoptotic nuclei in the BAT of hyperinsulinaemic rats was also confirmed by propidium iodide staining. Immunohistochemical localization of tumor necrosis factor-alpha (TNF-α) revealed increased expression in ECs of BAT of hyperinsulinaemic animals, indicating its possible role in insulin-induced apoptotic changes. These results suggest that BAT atrophy in hyperinsulinaemia is a result of endothelial and adipocyte apoptosis combined, rather than any of functional components alone.     

Do birds possess brown adipose tissue?

• 1.1. Fluorescence and electron microscopy were used to visualize differences between avian adipose tissue (AAT) collected from clavicular and abdominal regions of the great tit, the willow tit, the house sparrow and the Japanese quail, and interscapular brown adipose tissue (BAT) obtained from the Djungarian dwarf hamster.
• 2.2. Multilocular fat cells were found in AAT. The prerequisite for multilocularity, however, was not simply winter acclimatization [short photophase 4L:20D and low ambient temperature (< −20°C in January in Oulu)] or cold-acclimation (−25°C). Multilocular adipocytes were found during autumn and in unacclimated control birds as well. Mitochondria in the AAT were fewer and about one-sixth the length of those in BAT. This finding was associated with low cytochrome oxidase (COX) activity in the tissue homogenate and isolated mitochondrial fraction of the AAT (< 5.2% of that in BAT).
• 3.3. Catecholamine fluorescence was seen only around arteries in the AAT. Signs of sympathetic parenchymal innervation were found neither in winter- nor in cold-acclimated birds, but typically, sympathetic nerve fibers forming a basket-like network around every cell were seen in the brown fat of the hamster.
• 4.4. Our results show that AAT in the adult birds resembles white adipose tissue more than brown. Multilocularity of adipocytes may improve lipolysis to deliver fatty acids for muscle fuel of shivering or NST.

     

Metabolic effects of tumour necrosis factor-α on rat brown adipose tissue

Intravenous administration of a single dose (100 g/kg bw) of recombinant tumour necrosis factor- (TNF, cachectin) to rats increased the rate ofin vitro fatty acid synthesis in interscapular brown adipose tissue (IBAT) from both glucose and alanine, without changes in the oxidation of these substrates to¹⁴CO₂. Lactate production and glycerol release were also unaffected by treatment with the cytokine. Additionally, the presence of TNF in the incubation media did not affect fatty acid synthesis, suggesting an indirect effect of the cytokine. The activities of different enzymes of glucose and alanine metabolism such as hexokinase, phosphofructokinase, pyruvate kinase, glucose-6-phosphate dehydrogenase and alanine transaminase, did not suffer changes as a consequence of TNF administration. The same applied to the enzymatic activities involved in fatty acid synthesis such as fatty acid synthase, acetyl-CoA carboxylase and ATP-citrate lyase. Conversely, citrate levels in IBAT were increased in animals treated with TNF, suggesting that it could be the cause for the increased fatty acid synthesis in this tissue.     

Increased α1 receptor density in brown adipose tissue of cafeteria-fed rats

A possible general corollary between ₁-receptor density in brown adipose tissue and the degree of activation of the tissue was investigated. For this purpose, the effect of cafeteria feeding on ₁-adrenergic receptors in brown adipose tissue of seven-week-old female rats was studied by the use of the ₁-antagonist (³H)prazosin. In cafeteria-fed rats, the K_D of the ₁-receptor for (³H)prazosin was unchanged (about 0.35 nM), but the receptor density was doubled (up to 40 fmol per mg of membrane protein). This was also observed when the results were expressed per unit of a plasma-membrane marker (5-nucleotidase). It was concluded that an increased ₁-receptor density is seen not only in cold-acclimated rats, but also in other conditions where brown fat is activated, and a possible general physiological significance of ₁-adrenergic pathways in brown adipose tissue is discussed.     

Irbesartan increased PPARγ activity in vivo in white adipose tissue of atherosclerotic mice and improved adipose tissue dysfunction

The effect of the PPARγ agonistic action of an AT₁ receptor blocker, irbesartan, on adipose tissue dysfunction was explored using atherosclerotic model mice. Adult male apolipoprotein E-deficient (ApoEKO) mice at 9 weeks of age were treated with a high-cholesterol diet (HCD) with or without irbesartan at a dose of 50 mg/kg/day for 4 weeks. The weight of epididymal and retroperitoneal adipose tissue was decreased by irbesartan without changing food intake or body weight. Treatment with irbesartan increased the expression of PPARγ in white adipose tissue and the DNA-binding activity of PPARγ in nuclear extract prepared from adipose tissue. The expression of adiponectin, leptin and insulin receptor was also increased by irbesartan. These results suggest that irbesartan induced activation of PPARγ and improved adipose tissue dysfunction including insulin resistance.     

Major involvement of mTOR in the PPARγ-induced stimulation of adipose tissue lipid uptake and fat accretion

Evidence points to a role of the mammalian target of rapamycin (mTOR) signaling pathway as a regulator of adiposity, yet its involvement as a mediator of the positive actions of peroxisome proliferator-activated receptor (PPAR)γ agonism on lipemia, fat accretion, lipid uptake, and its major determinant lipoprotein lipase (LPL) remains to be elucidated. Herein we evaluated the plasma lipid profile, triacylglycerol (TAG) secretion rates, and adipose tissue LPL-dependent lipid uptake, LPL expression/activity, and expression profile of other lipid metabolism genes in rats treated with the PPARγ agonist rosiglitazone (15 mg/kg/day) in combination or not with the mTOR inhibitor rapamycin (2 mg/kg/day) for 15 days. Rosiglitazone stimulated adipose tissue mTOR complex 1 and AMPK and induced TAG-derived lipid uptake (136%), LPL mRNA/activity (2- to 6-fold), and fat accretion in subcutaneous (but not visceral) white adipose tissue (WAT; 50%) and in brown adipose tissue (BAT; 266%). Chronic mTOR inhibition attenuated the upregulation of lipid uptake, LPL expression/activity, and fat accretion induced by PPARγ activation in both subcutaneous WAT and BAT, which resulted in hyperlipidemia. In contrast, rapamycin did not affect most of the other WAT lipogenic genes upregulated by rosiglitazone. Together these findings demonstrate that mTOR is a major regulator of adipose tissue LPL-mediated lipid uptake and a critical mediator of the hypolipidemic and lipogenic actions of PPARγ activation.     

The effects of treadmill exercise on expression of UCP-2 of brown adipose tissue and TNF-α of soleus muscle in obese Zucker rats

Sorts of abnormal state, obesity and inflammation are involved in a number of serious disease occurring and both of them became important research topics among molecular biologists. UCP-2 and TNF-α respectively reflecting obese and inflammatory status have often been used to evaluate the effects of independent variable, such as exercise, on them. Because exercise has shown its potent control on obesity and inflammation, it is necessary to determine if exercise is working via same bioindices. The purpose of this study was to determine the effects of different treadmill exercise intensities on UCP-2 of brown adipose tissue and TNF-α of soleus muscle during 8 weeks in Zucker rat. Zucker rats were divided into four groups (n = 7 in each group): control group, low intensity exercise group, moderate intensity exercise group and high intensity exercise group. Zucker rats of the exercise groups were made to run on a motorized treadmill for 30 minutes once a day during 8 weeks. Rats were sacrificed 24 hours after the last bout of exercise. Blood glucose in Zucker rats were measured by Gluco-Card Ⅱ. Brown adipose tissue were extracted to analyze the level of UCP-2 and TNF-α, respectively. UCP-2 and TNF-α were analyzed using the Western Blotting technique. Statistical techniques for data analysis were repeated measure ANOVA and one way ANOVA to determine the difference between groups, and for post hoc test was Duncan'' test. The 5% level of significance was utilized as the critical level for acceptance of hypotheses for the study. The following results were obtained from this study; UCP-2 protein expression of brown adipose tissue in Zucker rats were increased significantly following exercise of the low and moderate intensities compared to those of control group after 8 weeks. It was shown that TNF-α protein expression of soleus muscle in Zucker rats were decreased significantly following exercise of the low and moderate intensities compared to those of control group after 8 weeks. But no significant differences in levels of fasting glucose were shown between groups. The present data suggested that low and moderate intensities treadmill exercise may improve glycometabolism control and fat oxidation by up-regulating UCP-2 expression. In addition, we found low and moderate intensities reduce damages on skeletal muscle by down-regulation the TNF-α in Zucker rats. Thus, the low and moderate intensity exercise are appropriate for anti-obesity and inflammatory effects.     

α- andβ-adrenergic control of thermogenin mRNA expression in brown adipose tissue

By the use of an earlier characterised cDNA clone, CIN-1, corresponding to a sequence of the mRNA coding for the brown-fat specific uncoupling protein, thermogenin, the amount of thermogenin mRNA found in the brown adipose tissue of mice was quantitatively investigated under different physiological and pharmacological conditions.It was found that a 4 hr cold stress led to a 7-fold increase in the amount of thermogenin mRNA; injection of norepinephrine had a significant but smaller effect. Most notably, isoprenaline (-agonist) and phenylephrine (-agonist) had in themselves no effect, but when injected together were able to increase the mRNA level synergistically. In 4 hr cold-stressed mice, norepinephrine, isoprenaline and cholera toxin could all further potentiate the effect of the cold stress itself on the mRNA level. Insulin and the glucocorticoid dexamethasone both had weak stimulatory effects on the mRNA level.It is concluded that an increase in intracellular cAMP levels is a necessary and perhaps sufficient stimulus for the increase in thermogenin gene expression. However, at least underin vivo conditions, this increase requires stimulation of both - and-adrenergic pathways.     

Human brown adipose tissue: Underestimated target in metabolic disease?

Active brown adipose tissue (BAT) has, since it rediscovery in adult humans in 2009, received much attention for its ability to increase energy expenditure when activated. By means of mitochondrial uncoupling activity BAT's main function is to produce heat instead of storing energy such as in white adipose tissue (WAT). Therefore, BAT is considered a new potential target to treat obesity and the metabolic syndrome. However, the contribution of this thermogenic tissue is still a matter of debate among researchers.The aim of this review is to give an overview of the differences between classical brown adipocytes and inducible beige adipocytes in humans, and the potential activators of BAT in humans. Furthermore newly described genetic markers for identification of these two types of brown adipocytes are examined. Finally, the potential of the current measurement techniques, and the contribution of BAT activity to whole body energy expenditure are discussed.     

Glycerokinase activity in brown adipose tissue: a sympathetic regulation?

The effect of brown adipose tissue (BAT) sympathetic hemidenervation on the activity of glycerokinase (GyK) was investigated in different physiological conditions. In rats fed a balanced diet, the activity of the enzyme was approximately 50% lower in BAT-denervated pads than in intact, innervated pads. In rats adapted to a high-protein, carbohydrate-free diet, norepinephrine turnover rates and BAT GyK activity were already reduced, and BAT denervation resulted in a further decrease in the activity of the enzyme. Cold acclimation of normally fed rats at 4 degrees C for 10 days markedly increased the activity of the enzyme. Cold exposure (4 degrees C) for 6 h was insufficient to stimulate BAT GyK, but the activity of the enzyme was already increased after 12 h of cold exposure. The cold-induced BAT GyK stimulation was completely blocked in BAT-denervated pads. The data indicate that an adequate sympathetic flow to BAT is required for the maintenance of normal levels of GyK activity and for the enzyme response to situations, such as cold exposure, which markedly increase BAT sympathetic flow.     

Hormone-sensitive lipase modulates adipose metabolism through PPARγ

Hormone-sensitive lipase (HSL) is rate limiting for diacylglycerol and cholesteryl ester hydrolysis in adipose tissue and essential for complete hormone-stimulated lipolysis. Gene expression profiling in HSL-/- mice suggests that HSL is important for modulating adipogenesis and adipose metabolism. To test whether HSL is required for the supply of intrinsic ligands for PPARγ for normal adipose differentiation, HSL-/- and wild-type (WT) littermates were fed normal chow (NC) and high-fat (HF) diets supplemented with or without rosiglitazone (200 mg/kg) for 16 weeks. Results show that supplementing rosiglitazone to an NC diet completely normalized the decreased body weight and adipose depots in HSL-/- mice. Additionally, rosiglitazone resulted in similar serum glucose, total cholesterol, FFA, and adiponectin values in WT and HSL-/- mice. Furthermore, rosiglitazone normalized the expression of genes involved in adipocyte differentiation, markers of adipocyte differentiation, and enzymes involved in triacylglycerol synthesis and metabolism, and cholesteryl ester homeostasis, in HSL-/- mice. Supplementing rosiglitazone to an HF diet resulted in improved glucose tolerance in both WT and HSL-/- animals and also partial normalization in HSL-/- mice of abnormal WAT gene expression, serum chemistries, organ and body weight changes. In vitro studies showed that adipocytes from WT animals can provide ligands for activation of PPARγ and that activation is further boosted following lipolytic stimulation, whereas adipocytes from HSL-/- mice displayed attenuated activation of PPARγ, with no change following lipolytic stimulation. These results suggest that one of the mechanisms by which HSL modulates adipose metabolism is by providing intrinsic ligands or pro-ligands for PPARγ.     

The contrasting roles of PPARδ and PPARγ in regulating the metabolic switch between oxidation and storage of fats in white adipose tissue

Background  

The nuclear receptors peroxisome proliferator-activated receptor γ (PPARγ) and peroxisome proliferator-activated receptor δ (PPARδ) play central roles in regulating metabolism in adipose tissue, as well as being targets for the treatment of insulin resistance. While the role of PPARγ in regulating insulin sensitivity has been well defined, research into PPARδ has been limited until recently due to a scarcity of selective PPARδ agonists.     

Iodothyronine 5′-deiodinase activity in rat brown adipose tissue during development

Iosothyronine 5′-deiodinase activity in rat brown adipose tissue has a characteristic pattern of developmental changes that is completely different from that of the liver. Fetal brown fat exhibits an extremely high iodothyronine 5′-deiodinase activity that is approx. 10-fold that in adult rats. Even though brown fat iodothyronine 5′-deiodinase activity falls suddenly at birth, there is a new peak in the activity around days 5–7 of life, whereas it remaines very low afterwards. Just after birth, brown adipose tissue iodothyronine 5′-deiodinase activity is already capable of stimulation by noradrenaline. The postnatal peak in brown fat iodothyronine 5′-deiodinase correlates with the known increase in the thermogenic activity of the tissue in the neonatal rat, thus reinforcing the suggestion that local 3′,3,5-triiodothyronine generation could be an important event related to thermogeneis in brown adispose tissue. However, the high fetal activity was only slightly related to the thermogenic activity of brown fat. Moreover, the increased iodothyronine 5′-deiodinase activity of brown adipose tissue during fetal and neonatal life suggests a substantial contribution by brown fat in the overall extrahydroidal 3′,3,5-triiodothyronine production in these physiological periods.     

Lack of activation of UCP1 in isolated brown adipose tissue mitochondria by glucose-O-ω-modified saturated fatty acids of various chain lengths

We previously demonstrated that uncoupling protein 1 activity, as measured in isolated brown adipose tissue mitochondria (and as a native protein reconstituted into liposome membranes), was not activated by the non-flippable modified saturated fatty acid, glucose-O-ω-palmitate, whereas activity was stimulated by palmitate alone (40 nM free final concentration). In this study, we investigated whether fatty acid chain length had any bearing on the ability of glucose-O-ω-fatty acids to activate uncoupling protein 1. Glucose-O-ω-saturated fatty acids of various chain lengths were synthesized and tested for their potential to activate GDP-inhibited uncoupling protein 1-dependent oxygen consumption in brown adipose tissue mitochondria, and the results were compared with equivalent non-modified fatty acid controls. Here we demonstrate that laurate (12C), palmitate (16C) and stearate (18C) could activate GDP-inhibited uncoupling protein 1-dependent oxygen consumption in brown adipose tissue mitochondria, whereas there was no activation with glucose-O-ω-laurate (12C), glucose-O-ω-palmitate (16C), glucose-O-ω-stearate (18C), glucose-O-ω-arachidate (20C) or arachidate alone. We conclude that non-flippable fatty acids cannot activate uncoupling protein 1 irrespective of chain length. Our data further undermine the cofactor activation model of uncoupling protein 1 function but are compatible with the model that uncoupling protein 1 functions by flipping long-chain fatty acid anions.     

PPARγ signal transduction pathway in the foam cell formation induced by visfatin

The aim of the present study was to investigate the role of peroxisome proliferator-activated receptor γ (PPARγ) signal transduction pathway in the expression of ATP binding cassette transporter A1 (ABCA1) and acyl-CoA:cholesterol acyltransferase 1 (ACAT1) induced by visfatin and to discuss the mechanism of foam cell formation induced by visfatin. THP-1 monocytes were induced into macrophages by 160 nmol/L phorbol myristate acetate (PMA) for 48 h, and then the macrophages were exposed to visfatin and PPARγ activator rosiglitazone, respectively. The expressions of PPARγ, ABCA1 and ACAT1 mRNA and protein were determined by RT-PCR and Western blot respectively. The contents of total cholesterol (TC) and free cholesterol (FC) were detected by enzyme fluorescence analysis. The content of cholesterol ester (CE) was calculated by the difference between TC and FC. The results showed that visfatin decreased the mRNA and protein expressions of PPARγ and ABCA1, increased the mRNA and protein expressions of ACAT1, and increased the contents of FC and CE in a concentration-dependent manner. These above effects of visfatin were inhibited by rosiglitazone in a concentration-dependent manner. These results suggest that visfatin may down-regulate the ABCA1 expression and up-regulate the ACAT1 expression via PPARγ signal transduction pathway, which decreases the outflow of FC, increases the content of CE, and then induces foam cell formation.