Uncoupling Protein mRNA Expression Research Articles

Lycopene supplementation of maternal and weanling high-fat diets influences adipose tissue development and metabolic outcomes of Sprague-Dawley offspring.

AbstractDietary patterns high in fat contribute to the onset of cardiometabolic disease through the accrual of adipose tissue (AT). Lycopene, a carotenoid shown to exert multiple health benefits, may disrupt these metabolic perturbations. The purpose of the present study was to evaluate AT development and obesity-associated metabolic outcomes in the neonate and weanling offspring of Sprague-Dawley mothers fed a high-fat diet (HFD = 50 % fat) with and without lycopene supplementation. Sprague-Dawley rats consumed either a normal fat diet (NFD; 25 % fat) or HFD throughout gestation. Upon delivery, half of HFD mothers were transitioned to an HFD supplemented with 1 % lycopene (HFDL). At postnatal day 14 (P14), P25, and P35, pups were euthanised, body weight was recorded, and visceral white AT (WAT) and brown AT (BAT) mass were determined. Serum redox status, adipokines, glucose and inflammatory biomarkers were evaluated, as well as BAT mRNA expression of uncoupling protein 1 (UCP1). The HFD was effective in inducing weight gain as evident by significantly greater BW and WAT in the HFD group compared to the NFD group across all time points. Compared to HFD, the HFDL group exhibited significantly greater BAT with concomitant reductions in WAT mass, serum lipid peroxides and serum glucose. No significant differences were observed in serum adipokines, inflammatory markers or UCP1 expression despite the aforementioned alterations in AT development. Results suggest that dietary lycopene supplementation may influence metabolic outcomes during the weaning and post-weaning periods. Additional research is warranted to elucidate molecular mechanisms by which lycopene influences AT biology.

Distinctive Features of Orbital Adipose Tissue (OAT) in Graves’ Orbitopathy

Depot specific expansion of orbital-adipose-tissue (OAT) in Graves’ Orbitopathy (GO) is associated with lipid metabolism signaling defects. We hypothesize that the unique adipocyte biology of OAT facilitates its expansion in GO. A comprehensive comparison of OAT and white-adipose-tissue (WAT) was performed by light/electron-microscopy, lipidomic and transcriptional analysis using ex vivo WAT, healthy OAT (OAT-H) and OAT from GO (OAT-GO). OAT-H/OAT-GO have a single lipid-vacuole and low mitochondrial number. Lower lipolytic activity and smaller adipocytes of OAT-H/OAT-GO, accompanied by similar essential linoleic fatty acid (FA) and (low) FA synthesis to WAT, revealed a hyperplastic OAT expansion through external FA-uptake via abundant SLC27A6 (FA-transporter) expression. Mitochondrial dysfunction of OAT in GO was apparent, as evidenced by the increased mRNA expression of uncoupling protein 1 (UCP1) and mitofusin-2 (MFN2) in OAT-GO compared to OAT-H. Transcriptional profiles of OAT-H revealed high expression of Iroquois homeobox-family (IRX-3&5), and low expression in HOX-family/TBX5 (essential for WAT/BAT (brown-adipose-tissue)/BRITE (BRown-in-whITE) development). We demonstrated unique features of OAT not presented in either WAT or BAT/BRITE. This study reveals that the pathologically enhanced FA-uptake driven hyperplastic expansion of OAT in GO is associated with a depot specific mechanism (the SLC27A6 FA-transporter) and mitochondrial dysfunction. We uncovered that OAT functions as a distinctive fat depot, providing novel insights into adipocyte biology and the pathological development of OAT expansion in GO.

Can eosinophils in adipose tissue add fuel to the fire?

In inguinal adipose tissue, beige adipocytes are interspersed among white adipocytes and in close communication with dynamic populations of immune cells. Recent data have demonstrated that anti-inflammatory macrophages (M2) increase thermogenic activity of beige adipocytes, although the mechanism is currently under debate. ILC2, cells via secretion, of methionine enkephalin peptide were found to be able to increase beige thermogenesis. Knights et al. have recently generated a whole-body knock-out of KLF3 (KLF3-/-) to explore its contribution to thermogenesis and weight gain in a diet-induced obesity animal model.

Effects of corticosterone within the hypothalamic arcuate nucleus on food intake and body weight in male rats

ObjectiveObesity is a major cause of morbidity and mortality. Few weight-reducing medications are available, and these have limited efficacy. Cushing's Syndrome (caused by elevated glucocorticoid levels) and obesity have similar metabolic features. Though circulating glucocorticoid levels are not elevated in obesity, tissue-specific glucocorticoid levels have been implicated in the development of the metabolic phenotype of obesity. Tissue glucocorticoid levels are regulated by 11β-hydroxysteroid dehydrogenase type1 (11βHSD1), which increases the local concentration of active glucocorticoids by the production of corticosterone from 11-dehydrocorticosterone. 11βHSD1 is expressed in the hypothalamic arcuate nucleus (ARC), a major weight and appetite-regulating centre, and therefore represents a target for novel anti-obesity therapeutic agents. Thus, we sought to investigate the effect of chronic alterations of ARC corticosterone levels (mediated by 11βHSD1) on food intake and body weight in adult male rats. MethodsRecombinant adeno-associated virus particles bearing sense 11βHSD1 (rAAV-S11βHSD1) and small interfering 11βHSD1 (rAAV-si11βHSD1), respectively, were stereotactically injected into the ARC (bilaterally) of adult male Wistar rats. rAAV-GFP was injected into control groups of male Wistar rats. Food intake and body weight were measured three times a week for 70 days. Terminal brain, plasma and intrascapular brown adipose tissue (iBAT) samples were taken for measurement of mRNA expression and hormone levels. ResultsCompared to controls, rAAV-S11βHSD1 injection resulted in higher ARC corticosterone levels, hyperphagia and increased weight gain. Conversely, rAAV-si11βHSD1 injection (compared to controls) resulted in lower ARC corticosterone levels, higher iBAT uncoupling protein-1 mRNA expression and less weight gain despite similar food intake. ConclusionsTherefore ARC corticosterone, regulated by 11βHSD1, may play a role in food intake and body weight regulation. These data have important implications for the development of centrally-acting 11βHSD1 inhibitors, which are currently being developed for the treatment of obesity, metabolic disorders, and other conditions.

Differences in Neuregulin 4 Expression in Children: Effects of Fat Depots and Obese Status

ABSTRACT Purpose To observe the expression of Nrg4, uncoupling protein-1 (UCP1), tumor necrosis factor α (TNFα), CD31, VE-cadherin/CDH5 and vascular endothelial growth factor A (VEGF-A) mRNA in abdominal subcutaneous (SC), omental (OM) adipose tissue in children with relation to anthropometric parameters. Further to verify the effect of inflammatory mediators on Nrg4 and UCP1 mRNA expression in adipocytes. Methods Paired SC and OM adipose tissues were obtained from 58 children. In vitro, the adipocytes isolated from primary inguinal adipose tissue of mice were treated with TNFα (50 ng/ml) for 12–48 h. mRNA levels of Nrg4, UCP1 and TNFα were determined by real-time PCR. Results Nrg4, UCP1, VEGF-A and CDH5 mRNA levels in SC were significantly higher than those in OM adipose tissue and the mRNA level of TNFα showed the opposite result. Moreover, Nrg4 and UCP1 mRNA in SC were significantly lower in overweight children compared to normal weight children. Nrg4 in SC and OM was negatively associated with BMISDS, WHtR. CDH55 mRNA in OM was negatively associated with WHR. VEGF-A was positively correlated with Nrg4 in SC. In vitro, Nrg4 and UCP1 mRNA levels in adipocytes were dose- and time-dependently decreased under TNFα treatment. Conclusions Nrg4, UCP1, VEGF-A and CDH5 mRNA expression in adipose tissues display a depot-specific pattern. Nrg4 mRNA levels in adipose tissue are decreased with obesity and associated with WAT browning and angiogenesis. TNFα may be involved in the regulation of Nrg4 level in adipose tissue, which may be one of the causes of the down-regulation of Nrg4 expression in obesity with chronic inflammatory response.

Depot-specific UCP1 expression in human white adipose tissue and its association with obesity-related markers.

This study investigated depot-specific mRNA expression of uncoupling protein 1 (UCP1) in human white adipose tissue (WAT) and its association with obesity-related markers. We recruited 39 normal-weight, 41 nondiabetic obese, and 22 diabetic obese women. We measured UCP1 mRNA expression in abdominal visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT), and investigated the associations between UCP1 mRNA expression in VAT and SAT, and obesity-related markers including mRNA expression of leptin, adiponectin, CCAAT-enhancer-binding protein homologous protein (CHOP), and positive regulatory domain-containing protein 16 (PRDM16). We also evaluated UCP1 mRNA expression in differentiated human white adipocytes after treatment with various stressors and metabolic improvement agents in vitro. UCP1 mRNA in VAT was significantly higher than in SAT in all groups. UCP1 mRNA in SAT was negatively correlated with BMI, total abdominal fat area, visceral fat area, blood pressure, fasting glucose, insulin, HOMA-IR score, triglyceride, hsCRP, fasting leptin levels, and adipocyte size. UCP1 mRNA in SAT was positively correlated with fasting adiponectin levels. UCP1 mRNA in VAT was negatively correlated with visceral-to-subcutaneous fat ratio (VSR), fasting glucose, and triglyceride levels. In SAT, UCP1 mRNA was negatively correlated with mRNA expression of leptin and CHOP, and positively correlated with mRNA expression of adiponectin. The expression of PRDM16 was positively correlated with UCP1 mRNA in both VAT and SAT. UCP1 mRNA expression in differentiated human white adipocytes was significantly reduced after incubation with thapsigargin, tunicamycin, homocysteine, TNF-α, or IL-β, and significantly increased after incubation with exendin 4, dapagliflozin, and telmisartan. This study demonstrated depot-specific mRNA expression of UCP1 and its association with obesity-related markers in human WAT. UCP1 mRNA in human white adipocytes was suppressed by inflammatory agents and enhanced by metabolic improvement agents. UCP1 in human WAT might participate in the pathogenesis of obesity-related metabolic diseases.

Postnatal leptin surge is critical for the transient induction of the developmental beige adipocytes in mice.

Beige adipocytes have become a promising therapeutic target to combat obesity. Our senior author Dr. B. Xue previously discovered a transient but significant induction of beige adipocytes in mice during early postnatal development, which peaked at postnatal day (P) 20 and then disappeared thereafter. However, the physiological mechanism underlying the transient induction of the developmental beige cells remains mystery. Interestingly, there exists a postnatal surge of leptin in mice at P10 before the appearance of the developmental beige adipocytes. Given the neurotropic effect of leptin during neuronal development and its role in activating the sympathetic nervous system (SNS), we tested the hypothesis that postnatal leptin surge is required for the transient induction of developmental beige adipocytes through sympathetic innervation. Unlike wild-type (WT) mice that were able to acquire the developmentally induced beige adipocytes at P20, ob/ob mice had much less uncoupling protein 1 (UCP1)-positive multilocular cells in inguinal white adipose tissue at the same age. This was consistent with reduced expression of UCP1 mRNA and protein levels in white fat of ob/ob mice. In contrast, daily injection of ob/ob mice with leptin between P8 and P16, mimicking the postnatal leptin surge, largely rescued the ability of these mice to acquire the developmentally induced beige adipocytes at P20, which was associated with enhanced sympathetic nerve innervation assessed by whole mount adipose tissue immunostaining of tyrosine hydroxylase. Our data demonstrate that the postnatal leptin surge is essential for the developmentally induced beige adipocyte formation in mice, possibly through increasing sympathetic nerve innervation.

Beta-1 and Not Beta-3 Adrenergic Receptors May Be the Primary Regulator of Human Brown Adipocyte Metabolism.

Brown adipose tissue (BAT) activation in humans has gained interest as a potential target for treatment of obesity and insulin resistance. In rodents, BAT is primarily induced through beta-3 adrenergic receptor (ADRB3) stimulation, whereas the primary beta adrenergic receptors (ADRBs) involved in human BAT activation are debated. We evaluated the importance of different ADRB subtypes for uncoupling protein 1 (UCP1) induction in human brown adipocytes. A human BAT cell model (TERT-hBA) was investigated for subtype-specific ADRB agonists and receptor knockdown on UCP1 mRNA levels and lipolysis (glycerol release). In addition, fresh human BAT biopsies and TERT-hBA were evaluated for expression of ADRB1, ADRB2, and ADRB3 using RT-qPCR. The predominant ADRB subtype in TERT-hBA adipocytes and BAT biopsies was ADRB1. In TERT-hBA, UCP1 mRNA expression was stimulated 11.0-fold by dibutyryl cAMP (dbcAMP), 8.0-fold to 8.4-fold by isoproterenol (ISO; a pan-ADRB agonist), and 6.1-fold to 12.7-fold by dobutamine (ADRB1 agonist), whereas neither procaterol (ADRB2 agonist), CL314.432, or Mirabegron (ADRB3 agonists) affected UCP1. Similarly, dbcAMP, ISO, and dobutamine stimulated glycerol release, whereas lipolysis was unaffected by ADRB2 and ADRB3 agonists. Selective knockdown of ADRB1 significantly attenuated ISO-induced UCP1 expression. The adrenergic stimulation of UCP1 and lipolysis may mainly be mediated through ADRB1. Moreover, ADRB1 is the predominant ADRB in both TERT-hBA and human BAT biopsies. Thus, UCP1 expression in human BAT may, unlike in rodents, primarily be regulated by ADRB1. These findings may have implications for ADRB agonists as future therapeutic compounds for human BAT activation.

The anti-obesity effect of FGF19 does not require UCP1-dependent thermogenesis

ObjectiveFibroblast growth factor 19 (FGF19) is a postprandial hormone which plays diverse roles in the regulation of bile acid, glucose, and lipid metabolism. Administration of FGF19 to obese/diabetic mice lowers body weight, improves insulin sensitivity, and enhances glycemic control. The primary target organ of FGF19 is the liver, where it regulates bile acid homeostasis in response to nutrient absorption. In contrast, the broader pharmacologic actions of FGF19 are proposed to be driven, in part, by the recruitment of the thermogenic protein uncoupling protein 1 (UCP1) in white and brown adipose tissue. However, the precise contribution of UCP1-dependent thermogenesis to the therapeutic actions of FGF19 has not been critically evaluated. MethodsUsing WT and germline UCP1 knockout mice, the primary objective of the current investigation was to determine the in vivo pharmacology of FGF19, focusing on its thermogenic and anti-obesity activity. ResultsWe report that FGF19 induced mRNA expression of UCP1 in adipose tissue and show that this effect is required for FGF19 to increase caloric expenditure. However, we demonstrate that neither UCP1 induction nor an elevation in caloric expenditure are necessary for FGF19 to induce weight loss in obese mice. In contrast, the anti-obesity action of FGF19 appeared to be associated with its known physiological role. In mice treated with FGF19, there was a significant reduction in the mRNA expression of genes associated with hepatic bile acid synthesis enzymes, lowered levels of hepatic bile acid species, and a significant increase in fecal energy content, all indicative of reduced lipid absorption in animals treated with FGF19. ConclusionTaken together, we report that the anti-obesity effect of FGF19 occurs in the absence of UCP1. Our data suggest that the primary way in which exogenous FGF19 lowers body weight in mice may be through the inhibition of bile acid synthesis and subsequently a reduction of dietary lipid absorption.

Down regulation of UCP2 expression in retinal pigment epithelium cells under oxidative stress: an in vitro study.

To evaluate the expression of uncoupling protein 2 (UCP2) in a retinal pigment epithelium cell line (ARPE-19), under oxidative stress (OS). ARPE-19 cells were divided into groups treated with various concentrations of hydrogen peroxide (H2O2; 0, 150, 300, 500, 700, and 900 µmol/L) for 24h, to induce oxidative damage and cell viability was assessed by MTT assay. UCP2 mRNA expression in cells treated with H2O2 was investigated by reverse transcription-polymerase chain reaction (RT-PCR). UCP2 protein expression was assessed by Western blotting and ROS levels analyzed by flow cytometry (FCM). Further, UCP2-siRNA treated cultures were exposed to H2O2 (0, 75, 150, and 300 µmol/L) for 2h and cell viability determined by MTT assay. Cells treated with higher concentrations of H2O2 appeared shrunken; their adhesion to adjacent cells was disrupted, and the number of dead cells increased. The results of cell viability assays demonstrated that the numbers of cells were decreased in a dose-dependent manner following treatment with H2O2. Compared with untreated controls, cell viability was significantly reduced after treatment with >300 µmol/L H2O2 (P<0.05). Cell metabolic activity was decreased with increased concentrations of H2O2 as detected by MTT assay. Levels of OS were further decreased in cells treated with UCP2-siRNA compared with those treated with H2O2 alone (P<0.05). The results of RT-PCR and Western blotting demonstrated that UCP2 expression was reduced in H2O2-treated groups compared with controls (P<0.05). FCM analysis showed that cell reactive oxygen species (ROS) levels were increased in H2O2-treated groups and further upregulated by UCP2-siRNA treatment (P<0.05). Expression levels of UCP2 are decreased in ARPE-19 cells treated with H2O2. ROS levels are further increased in cells treated with UCP2-siRNA relative to those treated with H2O2 alone. UCP2 may have a protective role in ARPE-19 cells during oxidative injury.

Dietary Fiber in Bilberry Ameliorates Pre-Obesity Events in Rats by Regulating Lipid Depot, Cecal Short-Chain Fatty Acid Formation and Microbiota Composition.

Obesity is linked to non-alcoholic fatty liver disease and risk factors associated to metabolic syndrome. Bilberry (Vaccinium myrtillus) that contains easily fermentable fiber may strengthen the intestinal barrier function, attenuate inflammation and modulate gut microbiota composition, thereby prevent obesity development. In the current study, liver lipid metabolism, fat depot, cecal and serum short-chain fatty acids (SCFAs) and gut microbiome were evaluated in rats fed bilberries in a high-fat (HFD + BB) or low-fat (LFD + BB) setting for 8 weeks and compared with diets containing equal amount of fiber resistant to fermentation (cellulose, HFD and LFD). HFD fed rats did not obtain an obese phenotype but underwent pre-obesity events including increased liver index, lipid accumulation and increased serum cholesterol levels. This was linked to shifts of cecal bacterial community and reduction of major SCFAs. Bilberry inclusion improved liver metabolism and serum lipid levels. Bilberry inclusion under either LFD or HFD, maintained microbiota homeostasis, stimulated interscapular-brown adipose tissue depot associated with increased mRNA expression of uncoupling protein-1; enhanced SCFAs in the cecum and circulation; and promoted butyric acid and butyrate-producing bacteria. These findings suggest that bilberry may serve as a preventative dietary measure to optimize microbiome and associated lipid metabolism during or prior to HFD.

2161-P: Overexpression of UCP2 in Pancreatic Beta Cells Caused Impaired Insulin Secretion through Mitochondrial Dysfunction and Aldolase B Expression

Uncoupling protein 2 (UCP2), a mitochondrial protein, is known to be upregulated in pancreatic islets of patients with type 2 diabetes (T2DM); however, the pathological significance of the increase in UCP2 expression remained unclear. In this study, we investigated the regulatory mechanisms of UCP2 expression and its effects on β-cell functions. The mRNA expression of UCP2 in mouse islets was upregulated by the treatment with glucose, glucokinase activator, or insulin (p&amp;lt;0.05), while metformin downregulated it (p&amp;lt;0.01). Islets from db/db mice and insulin receptor substrate-2 knockout (IRS-2KO) mice showed the increment in the UCP2 expression. Transgenic mouse with specific UCP2 overexpression in β-cells (βUCP2Tg) demonstrated impaired glucose tolerance due to reduced insulin secretion. Adenoviral expression of UCP2 in MIN6-M9 β-cell line also attenuated glucose-induced insulin secretion. The ATP production in response to glucose and mitochondrial Complex I protein level were reduced (p&amp;lt;0.01) in βUCP2Tg islets. Dilatation (1.8-fold, p&amp;lt;0.05) and morphological alteration of mitochondria were observed in βUCP2Tg β-cells. Gene expression microarray analysis revealed 18 upregulated genes (1.5-fold, p&amp;lt;0.05) and 2 downregulated genes (&amp;lt;67%, p&amp;lt;0.05) in βUCP2Tg islets. The most upregulated mRNA by UCP2 expression encoded the aldolase B (Aldob) (77-fold), whose expression was reported to be negatively associated with insulin secretion in human islets (J Clin Endocrinol Metab. 103(12):4373-4383, 2018). We validated increased Aldob expression in islets from βUCP2Tg mice, db/db mice, and IRS-2KO mice. Taken together, UCP2 expression seemed to be increased under T2DM conditions including hyperglycemia or hyperinsulinemia. Our data also suggested that the increase in UCP2 expression in T2DM β-cells cell-autonomously attenuated glucose-induced insulin secretion, possibly via mitochondrial abnormality and Aldob expression. Disclosure R. Inoue: None. J. Shirakawa: None. Y. Togashi: None. Y. Terauchi: Advisory Panel; Self; AstraZeneca, Daiichi Sankyo Company, Limited, Eli Lilly and Company, Merck Sharp &amp; Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Novo Nordisk A/S, Sanofi. Research Support; Self; Daiichi Sankyo Company, Limited, Eli Lilly and Company, Merck Sharp &amp; Dohme Corp., Novartis Pharmaceuticals Corporation, Novo Nordisk A/S, Ono Pharmaceutical Co., Ltd., Sanofi, Shionogi &amp; Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd. Speaker's Bureau; Self; Astellas Pharma Inc., AstraZeneca, Bayer Yakuhin, Ltd., Daiichi Sankyo Company, Limited, Eli Lilly and Company, Merck Sharp &amp; Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Novartis Pharmaceuticals Corporation, Novo Nordisk A/S, Ono Pharmaceutical Co., Ltd., Sanofi, Sanwa Kagaku Kenkyusho, Shionogi &amp; Co., Ltd., Sumitomo Dainippon Pharma Co., Ltd.

Role of epicardial adipose tissue NPR-C in acute coronary syndrome

Background and aimsIt has been suggested that epicardial adipose tissue (EAT) thermogenesis plays a role in coronary artery disease (CAD). Recent evidence indicates that natriuretic peptide receptors (NPRs) are critical for thermogenesis. We determined the expression and signaling of NPRs in EAT in the context of CAD progression and their association with brown fat-related genes, such as uncoupling protein 1 (UCP1) and peroxisome proliferator-activated receptor gamma coactivator alpha (PGC1α). MethodsNPR-A, NPR-B and NPR-C mRNA and protein expression levels were analyzed in EAT and thoracic subcutaneous adipose tissue (SAT) from non-CAD (NCAD), stable CAD and acute coronary syndrome (ACS) patients. The associations of NPRs with thermogenic genes were also evaluated. ResultsThe EAT of ACS patients showed lower NPR-C gene and protein expression levels compared with that of stable CAD or NCAD patients. NPR-C mRNA expression in EAT also decreased as the number of injured arteries rose, and correlated positively with left ventricular ejection fraction and EAT PGC1α mRNA expression. EAT PGC1α and UCP1 gene expression levels also decreased in the ACS group. Linear and logistic regression models showed associations of EAT NPR-C mRNA levels with EAT PGC1α mRNA levels and the presence of ACS. Furthermore, the EAT of ACS patients showed reduced p38 mitogen-activated protein kinase (p38 MAPK) phosphorylation levels, which correlated positively with NPR-C protein levels. ConclusionsThe EAT of patients with ACS is characterized by decreased NPR-C, reduced UCP1 and PGC1α mRNA expression levels and reduced activation of the p38 MAPK pathway. The associations among the expression of EAT NPR-C and ACS, and brown fat markers suggest that NPR-C may play a role in ACS and in the regulation of EAT brown-like fat features in humans.

Role of ketone signaling in the hepatic response to fasting.

Ketosis is a metabolic adaptation to fasting, nonalcoholic fatty liver disease (NAFLD), and prolonged exercise. β-OH butyrate acts as a transcriptional regulator and at G protein-coupled receptors to modulate cellular signaling pathways in a hormone-like manner. While physiological ketosis is often adaptive, chronic hyperketonemia may contribute to the metabolic dysfunction of NAFLD. To understand how β-OH butyrate signaling affects hepatic metabolism, we compared the hepatic fasting response in control and 3-hydroxy-3-methylglutaryl-CoA synthase II (HMGCS2) knockdown mice that are unable to elevate β-OH butyrate production. To establish that rescue of ketone metabolic/endocrine signaling would restore the normal hepatic fasting response, we gave intraperitoneal injections of β-OH butyrate (5.7 mmol/kg) to HMGCS2 knockdown and control mice every 2 h for the final 9 h of a 16-h fast. In hypoketonemic, HMGCS2 knockdown mice, fasting more robustly increased mRNA expression of uncoupling protein 2 (UCP2), a protein critical for supporting fatty acid oxidation and ketogenesis. In turn, exogenous β-OH butyrate administration to HMGCS2 knockdown mice decreased fasting UCP2 mRNA expression to that observed in control mice. Also supporting feedback at the transcriptional level, β-OH butyrate lowered the fasting-induced expression of HMGCS2 mRNA in control mice. β-OH butyrate also regulates the glycemic response to fasting. The fast-induced fall in serum glucose was absent in HMGCS2 knockdown mice but was restored by β-OH butyrate administration. These data propose that endogenous β-OH butyrate signaling transcriptionally regulates hepatic fatty acid oxidation and ketogenesis, while modulating glucose tolerance. NEW & NOTEWORTHY Ketogenesis regulates whole body glucose metabolism and β-OH butyrate produced by the liver feeds back to inhibit hepatic β-oxidation and ketogenesis during fasting.

Induction of beige adipocytes by naturally occurring β3-adrenoceptor agonist p-synephrine

The prevalence of obesity and its associated diseases is increasing worldwide, and the therapeutic potential of increasing energy expenditure through differentiation or activation of beige adipocytes has attracted much interest. Therefore, we explored naturally occurring compounds that induce beige adipocytes by screening for activity to induce mRNA expression of uncoupling protein 1 (UCP1) in stromal vascular fraction (SVF) cells cultured in beige adipocyte differentiation medium. Through screening, p-synephrine, a compound isolated from Citrus unshiu Marcov., was found to be an active compound that increased UCP1 mRNA expression in a dose-dependent manner from a concentration of 3.12 µM, which induced morphological changes specific for beige adipocytes. Similar effects were also observed in SVF cells prepared from db/db obese mice. While investigating the underlying mechanism of p-synephrine-induced beige adipocyte differentiation, we found that the effects of p-synephrine were abolished by the β3-adrenoceptor antagonist SR58894. Intriguingly, p-synephrine increased UCP1 mRNA levels in SVF cells cultured in beige adipocyte differentiation medium lacking insulin to an extent different from those by the β-agonist isoprenaline. Furthermore, phosphatidylinositol 3-kinase inhibitor LY294002 decreased isoprenaline-induced UCP1 mRNA levels in the early phase of beige adipocyte differentiation and p-synephrine-induced UCP1 mRNA levels in fully differentiated beige adipocytes. Thus, p-synephrine appears to elicit signals via β3-adrenoceptor combined with some part of the insulin signaling pathway, finally resulting in efficient stimulation of beige adipocyte differentiation with the support of certain beige adipocyte differentiation-inducing factors. The present results suggest the potential of p-synephrine for prophylaxis and treatment of obesity and its associated diseases.

Effects of visfatin on brown adipose tissue energy regulation using T37i cells

The role of brown adipose tissue (BAT) in pathological states of energy homeostasis and impaired adipocyte function, such as obesity has been a major area of research interest in recent years. Herein, we sought to determine the direct effects of adipokines, visfatin and leptin on BAT thermogenesis.The effects of mouse recombinant visfatin, nicotinamide mononucleotide (NMN) and leptin with or without FK866 were studied on differentiated T37i cells. Treated cells were analyzed for key genes and proteins regulating BAT [UCP-1, PRD1-BF1-RIZ1 homologous domain-containing 16 (PRDM-16), PPARgamma-coactivator-1alpha (PGC-1α) and receptor-interacting protein 140 (RIP-140)] using quantitative PCR and western blot analysis. Data is presented as mean P-values.Both visfatin and leptin had significant concentration dependent effects on thermogenesis in brown pre-adipocytes and at physiological levels, increased uncoupling protein-1 (UCP-1) levels in brown adipocytes. These effects of visfatin were similar to that of nicotinamide mononucleotide (NMN), further strengthening the enzymatic role of visfatin. We also showed that leptin induced UCP-1 mRNA expression and protein production appears to be mediated by visfatin. High concentrations of both visfatin and leptin led to a dramatic decrease in UCP-1 protein levels, supporting the notion that visfatin levels are raised in obesity and that obese people have reduced BAT activity, plausibly through a reduction in UCP-1 levels. Additionally, we found differential regulation of key brown adipogenic genes, specifically, PRD1-BF1-RIZ1 homologous domain-containing 16 (PRDM-16), PPARgamma-coactivator-1alpha (PGC-1α) and receptor-interacting protein 140 (RIP-140) by visfatin. Our observations provide novel insights in the potential actions of visfatin in BAT.

Consumption of the edible sea urchin Mesocentrotus nudus attenuates body weight gain and hepatic lipid accumulation in mice

The beneficial health effects of edible sea urchin consumption in mice fed a normal (ND) or high-fat diet (HFD) were investigated in this study. Notably, sea urchin-administered (250, 500, 1000 mg/kg) mice exhibited lower body, liver, and visceral fat weights, lower plasma levels of aspartate aminotransferase and alanine aminotransferase, and lower hepatic triacylglycerol levels than those fed carboxymethyl cellulose (CMC). Despite the high levels of cholesterol found in sea urchins, intake of these organisms had no effect on plasma cholesterol levels among the mice tested. Additionally, sea urchin consumption resulted in enhanced levels of arachidonic acid and docosahexaenoic acid within mouse livers. Lastly, mice fed HFD with sea urchin (500 mg/kg) exhibited increased mRNA expression of uncoupling protein-1 within brown adipose tissue, compared with those fed HFD with CMC. In conclusion, consumption of sea urchin might provide a protective effect against the development of obesity and/or nonalcoholic fatty liver disease.

Role of LncRNA ROR in Ad36-induced browning of human adipose-derived stem cells

Objective To investigate the role of LncRNA ROR in Ad36-induced browning of human adipose-derived stem cells(hADSC). Methods After hADSC was induced by cocktail and Ad36 for 2, 4, 6, and 8 days, Oil red O staining was performed for observing the adipogenic status. The mRNA expressions of LncRNA ROR, uncoupling protein 1(UCP1), and PRDM16 were detected by real- time PCR and the protein expressions of UCP1 and PRDM16 were detected by Western-blot. After LncRNA ROR was knocked down by siRNA, UCP1 and PRDM16 mRNA and protein expression levels in the process of Ad36-induced adipocyte differentiation were detected by real- time PCR and Western-blot. Results Oil red O staining showed that fat droplets in the cocktail-induced group were larger than those in the Ad36-induced group. Compared with the cocktail group, the mRNA expressions of LncRNA ROR, UCP1 and PRDM16, and the protein expressions of UCP1 and PRDM16 in Ad36 group were significantly increased(P<0.05). The mRNA and protein expressions of UCP1 and PRDM16 in LncRNA ROR knockdown group were significantly lower than those in control group(P<0.05). Conclusion In the process of Ad36-induced hADSC differentiation, the up-regulation of LncRNA ROR may stimulate UCP1 and PRDM16 expression, and thus promote the browning of hADSC. (Chin J Endocrinol Metab, 2018, 34: 318-324) Key words: Human adipose-derived stem cells; Browning; Cocktail; Ad36; Long noncoding RNA

Effects of α-(prazosin and yohimbine) and β-receptors activity on cAMP generation and UCP1 gene expression in brown adipocytes.

Abstract Background Brown adipose tissue (BAT) contains both α- and β-adrenergic receptors. In the literature, the activity of α-adrenoreceptors is less documented, and their functions still remain puzzling. The present investigation has been undertaken to understand α-adrenoreceptors’ activity and their relation between uncoupling protein 1 (UCP1) mRNA expression and cyclic AMP (cAMP) generation in BAT. Methods BAT precursor cells from young mice were grown in culture. Cells were exposed to norepinephrine (NE) and other agents. RNA was isolated after harvesting the cells, and northern blot was performed. Filters were exposed to film after hybridization with nick-translated complementary DNA probes, and results were evaluated by scanning. Amersham assay kit was used for cAMP measurement. Results Treatment of prazosin and yohimbine separately with 1 μM of NE shows stimulation of UCP1 mRNA expression 106% and 154%, respectively, whereas with that of both drugs shows only 76%. cAMP generation occurs 282% with prazosin, 100% with yohimbine, and 382% with both drugs with 1 μM of NE, whereas it is 310%, 40%, and 358%, respectively, with 10 μM of NE. Conclusions Stimulation of thermogenesis after treatment of prazosin and NE may be due to the inhibition of phosphodiesterase enzyme and with yohimbine and NE indicates the possibility of inhibition of the inhibitory effect of α2- and stimulation of α1-receptors. Increase of cAMP concentration with yohimbine and both drugs with NE are not correlated to UCP1 mRNA expression. This indicates that the relationship between cAMP elevation and stimulation of thermogenesis is not simple. This study clearly shows the interaction between β- and α-adrenoreceptor activities.

Indomethacin Enhances Brown Fat Activity.

Indomethacin, a nonsteroidal anti-inflammatory drug, has been shown to induce white adipocyte differentiation; however, its roles in brown adipocyte differentiation and activation in brown adipose tissue (BAT) and obesity are unknown. To address this issue, we treated mouse brown preadipocytes with different doses of indomethacin, and delivered indomethacin to interscapular BAT (iBAT) of obese mice using implanted osmotic pumps. Indomethacin dose dependently increased brown preadipocyte differentiation and upregulated both mRNA and protein expression of uncoupling protein 1 (UCP1) and peroxisome proliferator-activated receptor (PPAR) γ coactivator 1-alpha. The mechanistic study showed that indomethacin significantly activated the reporter driven by the PPAR response element, indicating that indomethacin may work as a PPARγ agonist in this cell line. Consistently, indomethacin significantly decreased iBAT mass and fasting blood glucose levels in high-fat diet-induced obesity (DIO) mice. Histologic analysis showed that brown adipocytes of indomethacin-treated mice contained smaller lipid droplets compared with control mice, suggesting that indomethacin alleviated the whitening of BAT induced by the high-fat diet. Moreover, indomethacin significantly increased UCP1 mRNA expression in iBAT. Taken together, this study indicates that indomethacin can promote mouse brown adipocyte differentiation, and might increase brown fat and glucose oxidation capacity in DIO mice.