Induction Of Browning Research Articles

Moxibustion-Simulating Bipolar Radiofrequency Suppresses Weight Gain and Induces Adipose Tissue Browning via Activation of UCP1 and FGF21 in a Mouse Model of Diet-Induced Obesity.

Background Obesity is a pathological condition associated with various diseases including diabetes, stroke, arthritis, infertility, and heart disease. Moxibustion is widely used to prevent and manage obesity in traditional Asian medicine. We tested our hypothesis that moxibustion-simulating bipolar radiofrequency (M-RF) can suppress total body and white adipose tissue (WAT) weight gain via induction of WAT browning in a mouse model of diet-induced obesity (DIO). Methods We designed an M-RF device that could accurately adjust the depth and temperature at which heat stimulation was administered into the abdomen of DIO mice. High-fat-fed male C57BL/6 mice were treated with the M-RF device every two or three days for three weeks. We then harvested WAT and serum from the mice and measured total body and WAT weight, size of adipocytes, mitochondrial contents, features of the dead adipocyte environment, and levels of uncoupling protein 1 (UCP1) and fibroblast growth factor 21 (FGF21). Results Heat stimulation by M-RF in DIO mice resulted in precise temperature adjustment in the mice abdomen, with variance less than 1°C. Additionally, M-RF stimulation inhibited body and WAT weight gain, resulting in increased formation of beige adipocytes, increased mitochondrial content, and decreased formation of dead adipocytes in WAT. Moreover, treatment of M-RF induced expression of UCP1 and FGF21 in serum and/or epididymal WATs in DIO mice. Conclusion Heat stimulation by M-RF treatment induced upregulation of UCP1 and FGF21 expression in serum and/or WATs, which was correlated with reduced total body and WAT weight gain in DIO mice.

Correction to: Pioglitazone reduces cold-induced brown fat glucose uptake despite induction of browning in cultured human adipocytes: a randomised, controlled trial in humans.

The baseline insulin data given in Table 1 for the placebo group were incorrectly reported as 51 ± 10pmol/l instead of 48 ± 10pmol/l. This mistake also impacts on data reported in Table 4.

Nobiletin induces brown adipocyte-like phenotype and ameliorates stress in 3T3-L1 adipocytes

Browning of white adipocytes (beiging) is an attractive therapeutic strategy against obesity and its associated metabolic complications. Nobiletin (NOB) is a polymethoxylated flavone present in citrus fruits and has been reported to have anti-obesity effects. Here, we report that nobiletin exerts dual modulatory effects on adipocytes via induction of browning in 3T3-L1 white adipocytes and amelioration of stress in adipocytes. Nobiletin-induced beiging was investigated by determining expression levels of beige-specific genes and proteins by RT-PCR and immunoblot analysis, respectively. Nobiletin treatment rapidly elevated the expression levels of beige-specific genes such as Cd137, Cidea, Tbx1, and Tmem26. Further, nobiletin enhanced expression of the key transcription factors C/EBPβ, PPARδ, and PPARα, which are responsible for remodeling of white adipocytes. Nobiletin also strikingly activated HIB1B brown adipocytes and induced mitochondrial biogenesis in 3T3-L1 white adipocytes. In addition, nobiletin altered the expression of several lipid metabolism-related proteins such as ACOX1, CPT1, FAS, p-PLIN, SREBP and SIRT1. Moreover, nobiletin ameliorated stress in adipocytes by inhibiting expression levels of key stress molecules such as JNK and c-JUN. Nobiletin-induced browning could be mediated by tight regulation of kinases, as nobiletin induced PKA and p-AMPK at the protein expression level, and inhibition of PKA and p-AMPK by H-89 and dorsomorphin, respectively, abolished expression of the thermogenic markers PGC-1α and UCP1. Taken together, our findings suggest that nobiletin plays a modulatory role in adipocytes via induction of browning in 3T3-L1 white adipocytes and activation of HIB1B brown adipocytes combined with amelioration of stress in adipocytes, thereby exhibiting therapeutic potential against obesity.

Pioglitazone reduces cold-induced brown fat glucose uptake despite induction of browning in cultured human adipocytes: a randomised, controlled trial in humans.

Increasing brown adipose tissue (BAT) activity is a possible therapeutic strategy to increase energy expenditure and glucose and lipid clearance to ameliorate obesity and associated comorbidities. The thiazolidinedione (TZD) class of glucose-lowering drugs increase BAT browning in preclinical experimental models but whether these actions extend to humans in vivo is unknown. The aim of this study was to determine the effect of pioglitazone treatment on adipocyte browning and adaptive thermogenesis in humans. We first examined whether pioglitazone treatment of cultured human primary subacromioclavicular-derived adipocytes induced browning. Then, in a blinded, placebo-controlled, parallel trial, conducted within the Baker Institute clinical research laboratories, 14 lean male participants who were free of cardiometabolic disease were randomised to receive either placebo (lactose; n=7, age 22±1years) or pioglitazone (45mg/day, n=7, age 21±1years) for 28days. Participants were allocated to treatments by Alfred Hospital staff independent from the study via electronic generation of a random number sequence. Researchers conducting trials and analysing data were blind to treatment allocation. The change in cold-stimulated BAT activity, assessed before and after the intervention by [18F]fluorodeoxyglucose uptake via positron emission tomography/computed tomography in upper thoracic and cervical adipose tissue, was the primary outcome measure. Energy expenditure, cardiovascular responses, core temperature, blood metabolites and hormones were measured in response to acute cold exposure along with body composition before and after the intervention. Pioglitazone significantly increased in vitro browning and adipogenesis of adipocytes. In the clinical trial, cold-induced BAT maximum standardised uptake value was significantly reduced after pioglitazone compared with placebo (-57±6% vs -12±18%, respectively; p<0.05). BAT total glucose uptake followed a similar but non-significant trend (-50±10% vs -6±24%, respectively; p=0.097). Pioglitazone increased total and lean body mass compared with placebo (p<0.05). No other changes between groups were detected. The disparity in the actions of pioglitazone on BAT between preclinical experimental models and our in vivo human trial highlight the imperative to conduct human proof-of-concept studies as early as possible in BAT research programmes aimed at therapeutic development. Our clinical trial findings suggest that reduced BAT activity may contribute to weight gain associated with pioglitazone and other TZDs. ClinicalTrials.gov NCT02236962 FUNDING: This work was supported by the Diabetes Australia Research Program and OIS scheme from the Victorian State Government.

MCT1 and MCT4 Expression and Lactate Flux Activity Increase During White and Brown Adipogenesis and Impact Adipocyte Metabolism

Adipose tissue takes up glucose and releases lactate, thereby contributing significantly to systemic glucose and lactate homeostasis. This implies the necessity of upregulation of net acid and lactate flux capacity during adipocyte differentiation and function. However, the regulation of lactate- and acid/base transporters in adipocytes is poorly understood. Here, we tested the hypothesis that adipocyte thermogenesis, browning and differentiation are associated with an upregulation of plasma membrane lactate and acid/base transport capacity that in turn is important for adipocyte metabolism. The mRNA and protein levels of the lactate-H+ transporter MCT1 and the Na+,HCO3− cotransporter NBCe1 were upregulated in mouse interscapular brown and inguinal white adipose tissue upon cold induction of thermogenesis and browning. MCT1, MCT4, and NBCe1 were furthermore strongly upregulated at the mRNA and protein level upon differentiation of cultured pre-adipocytes. Adipocyte differentiation was accompanied by increased plasma membrane lactate flux capacity, which was reduced by MCT inhibition and by MCT1 knockdown. Finally, in differentiated brown adipocytes, glycolysis (assessed as ECAR), and after noradrenergic stimulation also oxidative metabolism (OCR), was decreased by MCT inhibition. We suggest that upregulation of MCT1- and MCT4-mediated lactate flux capacity and NBCe1-mediated HCO3−/pH homeostasis are important for the physiological function of mature adipocytes.

Honokiol exerts dual effects on browning and apoptosis of adipocytes.

Induction of brown adipocyte-like phenotype (browning) in white adipocytes and promotion of apoptosis by dietary and pharmacological compounds is considered a novel strategy against obesity. Here, we show that honokiol exerts dual modulatory effects on adipocytes via induction of browning in 3T3-L1 white adipocytes and apoptosis as well as activation of HIB1B brown adipocytes combined with inhibition of apoptosis. Honokiol-induced browning and apoptosis were investigated by determining expression levels of brown adipocyte-specific genes and proteins by RT-PCR and immunoblot analysis, respectively. Apoptotic data were validated by immunofluorescence and ROS levels were measured by FACS analysis. Honokiol treatment induced browning by elevating expression levels of brown adipocyte-specific genes such as Cidea, Cox8, Fgf21, Pgc-1α, and Ucp1. Honokiol promoted apoptosis of 3T3-L1 white adipocytes and inhibited apoptosis of HIB1B brown adipocytes via opposite regulation of the pro-apoptotic protein BAX and anti-apoptotic protein Bcl-2. Honokiol also significantly increased protein expression levels of ACOX1, CPT1, p-HSL, and p-PLIN and reduced ROS levels, suggesting its possible role in fat oxidation and lipid catabolism. Honokiol-induced browning could be mediated by activation of ERK, as inhibition of ERK by FR180204 abolished expression of PGC-1α and UCP1. Our findings suggest that honokiol exhibits a modulatory role in adipocytes via induction of browning and apoptosis in white adipocytes, promotion of catabolic lipid metabolism, as well as activation and inhibition of apoptosis in HIB1B brown adipocytes, thereby exhibiting therapeutic potential against obesity.

Independence of brown induction and brightness induction

Independence of brown induction and brightness induction

Loss of ADAMTS5 enhances brown adipose tissue mass and promotes browning of white adipose tissue via CREB signaling

ObjectiveA potential strategy to treat obesity – and the associated metabolic consequences – is to increase energy expenditure. This could be achieved by stimulating thermogenesis through activation of brown adipose tissue (BAT) and/or the induction of browning of white adipose tissue (WAT). Over the last years, it has become clear that several metalloproteinases play an important role in adipocyte biology. Here, we investigated the potential role of ADAMTS5. MethodsMice deficient in ADAMTS5 (Adamts5−/−) and wild-type (Adamts5+/+) littermates were kept on a standard of Western-type diet for 15 weeks. Energy expenditure and heat production was followed by indirect calorimetry. To activate thermogenesis, mice were treated with the β3-adrenergic receptor (β3-AR) agonist CL-316,243 or alternatively, exposed to cold for 2 weeks. ResultsCompared to Adamts5+/+ mice, Adamts5−/− mice have significantly more interscapular BAT and marked browning of their subcutaneous (SC) WAT. Thermogenic pathway analysis indicated, in the absence of ADAMTS5, enhanced β3-AR signaling via activation of the cAMP response element-binding protein (CREB). Additional β3-AR stimulation with CL-316,243 promoted browning of WAT in Adamts5+/+ mice but had no additive effect in Adamts5−/− mice. However, cold exposure induced more pronounced browning of WAT in Adamts5−/− mice. ConclusionsThese data indicate that ADAMTS5 plays a functional role in development of BAT and browning of WAT. Hence, selective targeting of ADAMTS5 could provide a novel therapeutic strategy for treatment/prevention of obesity and metabolic diseases.

Regulation of Systemic Energy Homeostasis by Peripheral Serotonin

Whole body energy balance is achieved through the coordinated regulation of energy intake and energy expenditure in various tissues including liver, muscle and adipose tissues. A positive energy imbalance by excessive energy intake or insufficient energy expenditure results in obesity and related metabolic diseases. Although there have been many obesity treatment trials aimed at the reduction of energy intake, these strategies have achieved only limited success because of their associated adverse effects. Serotonin is among those traditional pharmacological targets for anti-obesity treatment because central 5-HT functions as an anorexigenic neurotransmitter in the brain. Thus, there have been many trials aimed at increasing the activity of 5-HT in the central nervous system, and some of the developed methods are already used in the clinical setting as anti-obesity drugs. However, recent studies suggest the new functions of peripheral serotonin in energy homeostasis ranging from the endocrine regulation by gut-derived serotonin to the autocrine/paracrine regulation by adipocyte-derived serotonin. Pharmacological inhibition of 5-HT synthesis leads to inhibition of lipogenesis in epididymal white adipose tissue (WAT), induction of browning in inguinal WAT and activation of adaptive thermogenesis in brown adipose tissue (BAT). Fat specific Tph1 knock-out (Tph1 FKO) mice exhibit similar phenotypes as mice with pharmacological inhibition of 5-HT synthesis, suggesting the localized effects of 5-HT in adipose tissues. In addition, Htr3a KO mice exhibit increased energy expenditure in BAT and Htr2a KO mice exhibit the decreased lipid accumulation in WAT. These data suggest the clinical significance of the peripheral serotonergic system as a new therapeutic target for anti-obesity treatment.

METRNL decreases during adipogenesis and inhibits adipocyte differentiation leading to adipocyte hypertrophy in humans.

Meteorin-like (METRNL) is a recently described circulating protein shown to be highly expressed in white adipose tissue and to beneficially affect energy metabolism in mice. We systematically evaluated the role of METRNL for human adipogenesis and its association with obesity, browning and hyperinsulinemia in children. In addition, we assessed the functional relevance of METRNL for human adipogenesis. METRNL expression decreased during human adipocyte differentiation in vitro. Coherently, METRNL expression was lower in isolated adipocytes compared with stromal vascular fraction (SVF) cells in human samples. Withdrawal of the peroxisome proliferator-activated receptor-γ (PPARγ) agonist rosiglitazone from adipogenic media partially preserved the METRNL downregulation during adipogenesis. METRNL expression was higher in adipocytes of obese compared with lean children and correlated with adipocyte size, whereas in SVF METRNL expression correlated with proliferation capacity. Concordantly, overexpression of METRNL inhibited human adipocyte differentiation as shown by decreased lipogenesis and lower expression of PPARγ and markers of adipogenesis, whereas experimental downregulation promoted adipogenesis. Proliferation, in contrast, was advanced by METRNL overexpression. These interactions with adipose tissue dynamics may contribute to the clinically observed body mass index-independent association of METRNL expression with hyperinsulinemia and adipose tissue inflammation in human samples. METRNL was not associated with UCP1 expression or induction of browning in white adipocytes. Taken together, the downregulation of METRNL during adipogenesis and functional induction of increased proliferation in SVF cells with concomitant inhibition of adipocyte differentiation may result in hypertrophic AT accumulation. This may also explain our observations of increased METRNL expression in adipocytes but not SVF cells in obese children compared with lean children and the subsequent hyperinsulinemia.

Reduction of Hypothalamic Endoplasmic Reticulum Stress Activates Browning of White Fat and Ameliorates Obesity.

The chaperone GRP78/BiP (glucose-regulated protein 78 kDa/binding immunoglobulin protein) modulates protein folding in reply to cellular insults that lead to endoplasmic reticulum (ER) stress. This study investigated the role of hypothalamic GRP78 on energy balance, with particular interest in thermogenesis and browning of white adipose tissue (WAT). For this purpose, we used diet-induced obese rats and rats administered thapsigargin, and by combining metabolic, histologic, physiologic, pharmacologic, thermographic, and molecular techniques, we studied the effect of genetic manipulation of hypothalamic GRP78. Our data showed that rats fed a high-fat diet or that were centrally administered thapsigargin displayed hypothalamic ER stress, whereas genetic overexpression of GRP78 specifically in the ventromedial nucleus of the hypothalamus was sufficient to alleviate ER stress and to revert the obese and metabolic phenotype. Those effects were independent of feeding and leptin but were related to increased thermogenic activation of brown adipose tissue and induction of browning in WAT and could be reversed by antagonism of β3 adrenergic receptors. This evidence indicates that modulation of hypothalamic GRP78 activity may be a potential strategy against obesity and associated comorbidities.

EndoG Knockout Mice Show Increased Brown Adipocyte Recruitment in White Adipose Tissue and Improved Glucose Homeostasis

Brown adipose tissue (BAT) plays a central role in the regulation of whole-body energy and glucose homeostasis owing to its elevated capacity for lipid and glucose oxidation. The BAT thermogenic function, which is essential for the defense of body temperature against exposure to low environmental temperatures, relies on the expression in the inner membrane of brown adipocyte's mitochondria of uncoupling protein-1, a protein that uncouples substrate oxidation from oxidative phosphorylation and leads to the production of heat instead of ATP. BAT thermogenesis depends on proper mitochondrial biogenesis during the differentiation of brown adipocytes. Despite the data that support a role for Endonuclease G (EndoG) in the process of mitochondrial biogenesis, its function in BAT has not been explored. Here, using an EndoG knockout mouse model, we demonstrate that EndoG is not essential for the expression of mitochondrial genes involved in substrate oxidation or for the induction of thermogenic genes in BAT in response to cold exposure. We also show that a lack of EndoG is associated with an increased expression of thermogenic genes (ie, uncoupling protein-1, peroxisome proliferator-activated receptor-γ coactivator-1α) in white adipose tissue (WAT) that correlates with the appearance of brown adipocyte-like cells interspersed among white adipocytes. Interestingly, the increased browning of WAT elicited by the lack of EndoG was associated with a better glucose tolerance and reduced fat mass. Our results suggest that the induction of browning in WAT by means of inhibiting EndoG activity appears as a potential therapeutic strategy to prevent obesity and ameliorate glucose intolerance.

Cannabidiol promotes browning in 3T3-L1 adipocytes.

Recruitment of the brown-like phenotype in white adipocytes (browning) and activation of existing brown adipocytes are currently being investigated as a means to combat obesity. Thus, a wide variety of dietary agents that contribute to browning of white adipocytes have been identified. The present study was designed to investigate the effects of cannabidiol (CBD), a major nonpsychotropic phytocannabinoid of Cannabis sativa, on induction of browning in 3T3-L1 adipocytes. CBD enhanced expression of a core set of brown fat-specific marker genes (Ucp1, Cited1, Tmem26, Prdm16, Cidea, Tbx1, Fgf21, and Pgc-1α) and proteins (UCP1, PRDM16, and PGC-1α). Increased expression of UCP1 and other brown fat-specific markers contributed to the browning of 3T3-L1 adipocytes possibly via activation of PPARγ and PI3K. In addition, CBD increased protein expression levels of CPT1, ACSL, SIRT1, and PLIN while down-regulating JNK2, SREBP1, and LPL. These data suggest possible roles for CBD in browning of white adipocytes, augmentation of lipolysis, thermogenesis, and reduction of lipogenesis. In conclusion, the current data suggest that CBD plays dual modulatory roles in the form of inducing the brown-like phenotype as well as promoting lipid metabolism. Thus, CBD may be explored as a potentially promising therapeutic agent for the prevention of obesity.

Induction of Browning of White Adipocyte Formation by Ganoderma tsugae Extract

Ganoderma, also known as Lingzhi or Reishi, is the top grade of Traditional Chinese Medicine (TCM) and has gained considerable attention in oriental countries for its potential functions of promoting the health benefit, regulating the vital energy, postponing the old and feeble, and extending life span that were mentioned in the TCM books. However, the metabolic health benefits by regulating the vital energy from Ganoderma remain to be experimentally confirmed. Adipose tissue is viewed as one major lipid metabolic and energy‐modulated organ, and is metabolically adaptive to external stimuli. Recently, browning of white adipocyte upon stimulation has been considered as a key aspect in energy metabolism due to its energy expenditure characteristics. The current study aims to investigate the direct role of Ganoderma extracts on adipocyte formation and lipid metabolism as well as regulation of energy homeostasis. To examine the relevance of these effects mediated by Ganoderma, in vitro and in vivo assays were performed after treatment with Ganoderma extracts. Here, we demonstrate that a quality assured extract of Ganoderma tsugae (GTE), a popular species of Lingzhi locally cultivated in Taiwan, had no obviously cytotoxic effect but contributed to adipogenesis accompanied with the changes of morphological features (lipid droplet and mitochondria) in 3T3‐L1 adipocytes. Moreover, the histological analysis of white fat‐pad tissues from mice during the development of obesity showed browning characteristics after treatment with GTE. Subsequently, western blotting further confirmed the browning of white adipocyte formation, including up‐regulation of adipogenic‐, mitochondrial‐ and energy‐related proteins induced by GTE. In addition, GTE treatment not only reduced lipogenesis and lipid droplet fusion but also promoted lipid mobilization. Taken together, our findings reveal for the first time that GTE promotes metabolic health benefit and regulates the vital energy, as mentioned in the traditional Chinese Materia Medica, by modulating intracellular lipid homeostasis and induction of browning of adipocyte formation. This study provides new perspectives to elucidate the role and physiological effects of Ganoderma via current biotechnological techniques.Support or Funding InformationThis study was supported by the Ministry of Science and Technology (MOST 104‐2320‐B‐039‐024), R.O.C.

Is Brown Induction Just Brightness Induction?

Is Brown Induction Just Brightness Induction?

Dichoptic Presentation Separates Brown Induction from Red/Green Balance Change

Dichoptic Presentation Separates Brown Induction from Red/Green Balance Change

Dichoptic perception of brown.

Two experiments assessed mechanisms underlying brown induction by presenting a foveal target disk and concentric annular surround stimuli that varied in contrast relative to larger backgrounds. Stimuli were presented under monocular, binocular, and dichoptic viewing conditions. Observers adjusted the luminance of the target disk to a criterion brown level. We found evidence for at least two separate mechanisms for brown induction: one mechanism that is dependent on physically contiguous contrast and operates in monocular pathways and another mechanism that responds to high luminance contrast anywhere in the visual field and can operate after convergence of signals from the two eyes.

No effects of surround complexity on brown induction.

A yellow stimulus turns brown when it is made sufficiently darker than its surroundings. Most previous studies have used simple contiguous surround stimuli to induce brown, so we know little about how brown induction may be controlled by more distant and more complex surround features. We begin to address this issue by varying the complexity of two configurations of achromatic surround stimuli. It was shown that the area most immediately contiguous to the test stimulus has strong effects on brown induction. More importantly, we found that neither the number of surround features nor the distribution of light in the surround region had an effect on brown induction, as long as the overall size of the surround region remained constant. Instead, we found that brown induction depended on the total amount of light in the constant-size surround region, regardless of how that light was distributed. This potentially distinguishes the mechanisms of brown induction from those of brightness induction.

Influence of surround proximity on induction of brown and darkness.

A bright white surround makes a yellow long-wavelength target look both browner and darker. We explored the parallel between these two types of induction by examining their dependence on the proximity of the bright surround to the target at two different time scales with 27ms and 1s stimulus durations. We assessed (a)brown induction by adjustment of target luminance to perceptual brown and yellow boundaries and (b)darkness induction by a successive matching procedure. We found that brown induction is a quick process that is robust even for 27ms stimuli. For darkness induction, there was a strong, spatially localized surround proximity effect for the 27ms stimuli and much weaker proximity effect for the 1s stimuli. For brown induction, proximity effects were generally weaker but still showed relatively stronger localized proximity effects for 27ms stimuli than for 1s stimuli. For these stimuli, darkness induction predicts the relative pattern but not the magnitudes of brown induction. Both brown and darkness inductions show the operation of quick, spatially localized processes that are apparently superseded by other processes for extended stimulus presentations.

Skeletal muscle mitochondrial uncoupling prevents diabetes but not obesity in NZO mice, a model for polygenic diabesity.

Induction of skeletal muscle (SM) mitochondrial stress by expression of uncoupling protein 1 (UCP1) in mice results in a healthy metabolic phenotype associated with increased secretion of FGF21 from SM. Here, we investigated whether SM mitochondrial uncoupling can compensate obesity and insulin resistance in the NZO mouse, a polygenic diabesity model. Male NZO mice were crossed with heterozygous UCP1 transgenic (tg) mice (mixed C57BL/6/CBA background) and further backcrossed to obtain F1 and N2 offspring with 50 and 75% NZO background, respectively. Male F1 and N2 progeny were fed a high-fat diet ad libitum for 20weeks from weaning. Blood glucose was reduced, and diabetes (severe hyperglycemia >300mg/dl) was fully prevented in both F1- and N2-tg progeny compared to a diabetes prevalence of 15% in F1 and 42% in N2 wild type. In contrast, relative body fat content and plasma insulin were decreased, and glucose tolerance was improved, in F1-tg only. Both F1 and N2-tg showed decreased lean body mass. Accordingly, induction of SM stress response including FGF21 expression and secretion was similar in both F1 and N2-tg mice. In white adipose tissue, expression of FGF21 target genes was enhanced in F1 and N2-tg mice, whereas lipid metabolism genes were induced in F1-tg only. There was no evidence for induction of browning in either UCP1 backcross. We conclude that SM mitochondrial uncoupling induces FGF21 expression and prevents diabetes in mice with a 50-75% NZO background independent of its effects on adipose tissue.