Brown Adipose Tissue Function Research Articles

PGC-1\u03b2-expressing POMC neurons mediate the effect of leptin on thermoregulation in the mouse

The arcuate nucleus (ARC) of the hypothalamus is a key regulator of food intake, brown adipose tissue (BAT) thermogenesis, and locomotor activity. Whole-body deficiency of the transcriptional coactivator peroxisome proliferator-activated receptor γ (PPARγ) coactivator-1β (PGC-1β) disrupts mouse circadian locomotor activity and BAT-regulated thermogenesis, in association with altered gene expression at the central level. We examined whether PGC-1β expression in the ARC is required for proper energy balance and locomotor behavior by generating mice lacking the PGC-1β gene specifically in pro-opiomelanocortin (POMC) neurons. POMC neuron-specific deletion of PGC-1β did not impact locomotor behavior, food intake, body composition, energy fuel utilization and metabolic rate in fed, 24-h fasted and 24-h refed conditions. In contrast, in the fed state, deletion of PGC-1β in POMC cells elevated core body temperature during the nighttime period. Importantly, this higher body temperature is not associated with changes in BAT function and gene expression. Conversely, we provide evidence that mice lacking PGC-1β in POMC neurons are more sensitive to the effect of leptin on heat dissipation. Our data indicate that PGC-1β-expressing POMC neurons are part of a circuit controlling body temperature homeostasis and that PGC-1β function in these neurons is involved in the thermoregulatory effect of leptin.

Mild-cold water swimming does not exacerbate white adipose tissue browning and brown adipose tissue activation in mice.

The present study investigated the effects of swimming physical training either thermoneutral or below thermoneutral water temperature on white (WAT) and brown (BAT) adipose tissue metabolism, morphology, and function. C57BL/6J male mice (n = 40; weight 25.3 ± 0.1g) were divided into control (CT30), cold control (CT20), trained (TR30), and cold trained (TR20) groups. Swimming training consisted of 30-min exercise at 30°C (control) or 20°C (cold) water temperature. After 8-week training, adipose tissues were excised and inguinal (ingWAT) and BAT were processed for histology, lipolysis, and protein contents of total OXPHOS, PGC1α, and UCP1 by western blotting analysis. Swimming training reduced body weight gain independently of water temperature (P < 0.05). ingWAT mass was decreased for TR30 in comparison to other groups (P < 0.05), while for BAT, there was a significant increase in CT20 in relation to CT30, and both trained groups were significantly increased in relation to control groups (P < 0.05). ingWAT mean adipocyte area was smaller for trained groups, and seemed to present multilocular adipocytes. Lipolytic activity and protein content of UCP1, PGC1α, and mitochondrial markers were increased in trained groups for ingWAT (P < 0.05), independent of water temperature (P > 0.05), and these patterns were not observed for BAT (P > 0.05). Our findings suggest that mild-cold water exposure and swimming physical exercise seem to, independently, promote browning in ingWAT with no effects on BAT; however, the association of exercise and mild-cold water did not exacerbate these effects.

Moderate SIRT1 overexpression protects against brown adipose tissue inflammation

ObjectiveMetainflammation is a chronic low-grade inflammatory state induced by obesity and associated comorbidities, including peripheral insulin resistance. Brown adipose tissue (BAT), a therapeutic target against obesity, is an insulin target tissue sensitive to inflammation. Therefore, it is necessary to find strategies to protect BAT against the effects of inflammation in energy balance. In this study, we explored the impact of moderate sirtuin 1 (SIRT1) overexpression on insulin sensitivity and β-adrenergic responses in BAT and brown adipocytes (BA) under pro-inflammatory conditions. MethodsThe effect of inflammation on BAT functionality was studied in obese db/db mice and lean wild-type (WT) mice or mice with moderate overexpression of SIRT1 (SIRT1Tg+) injected with a low dose of bacterial lipopolysaccharide (LPS) to mimic endotoxemia. We also conducted studies on differentiated BA (BA-WT and BA-SIRT1Tg+) exposed to a macrophage-derived pro-inflammatory conditioned medium (CM) to evaluate the protection of SIRT1 overexpression in insulin signaling and glucose uptake, mitochondrial respiration, fatty acid oxidation (FAO), and norepinephrine (NE)-mediated-modulation of uncoupling protein-1 (UCP-1) expression. ResultsBAT from the db/db mice was susceptible to metabolic inflammation manifested by the activation of pro-inflammatory signaling cascades, increased pro-inflammatory gene expression, tissue-specific insulin resistance, and reduced UCP-1 expression. Impairment of insulin and noradrenergic responses were also found in the lean WT mice upon LPS injection. In contrast, BAT from the mice with moderate overexpression of SIRT1 (SIRT1Tg+) was protected against LPS-induced activation of pro-inflammatory signaling, insulin resistance, and defective thermogenic-related responses upon cold exposure. Importantly, the decline in triiodothyronine (T3) levels in the circulation and intra-BAT after exposure of the WT mice to LPS and cold was markedly attenuated in the SIRT1Tg+ mice. In vitro BA experiments in the two genotypes revealed that upon differentiation with a T3-enriched medium and subsequent exposure to a macrophage-derived pro-inflammatory CM, only BA-SIRT1Tg+ fully recovered insulin and noradrenergic responses. ConclusionsThis study has ascertained the benefit of the moderate overexpression of SIRT1 to confer protection against defective insulin and β-adrenergic responses caused by BAT inflammation. Our results have potential therapeutic value in combinatorial therapies for BAT-specific thyromimetics and SIRT1 activators to combat metainflammation in this tissue.

Substrate Utilization by Brown Adipose Tissue: What's Hot and What's Not?

Our understanding of brown adipose tissue (BAT) function in humans has increased rapidly over the past 10 years. This is predominantly due to the development of powerful non-invasive imaging techniques such as positron emission tomography that can quantify BAT mass and function using metabolic tracers. Activation of BAT during cold–induced thermogenesis is an effective way to dissipate energy to generate heat and requires utilization of multiple energy substrates for optimal function. This has led to interest in the activation of BAT as a potential therapeutic target for type 2 diabetes, dyslipidaemia, and obesity. Here, we provide an overview of the current understanding of BAT substrate utilization in humans and highlight additional mechanisms found in rodents, where BAT more prominently contributes to energy expenditure. During thermogenesis, BAT demonstrates substantially increased glucose uptake which appears to be critical for BAT function. However, glucose is not fully oxidized, with a large proportion converted to lactate. The primary energy substrate for thermogenesis is fatty acids, released from brown adipocyte triglyceride stores. Active BAT also sequesters circulating lipids to sustain optimal thermogenesis. Recent evidence reveals that metabolic intermediates from the tricarboxylic acid cycle and glycolytic pathways also play a critical role in BAT function. Understanding the role of these metabolites in regulating thermogenesis and whole body substrate utilization may elucidate novel strategies for therapeutic BAT activation.

Studying Brown Adipose Tissue in a Human in vitro Context.

New treatments for obesity and associated metabolic disease are increasingly warranted with the growth of the obesity pandemic. Brown adipose tissue (BAT) may represent a promising therapeutic target to treat obesity, as this tissue has been shown to regulate energy expenditure through non-shivering thermogenesis. Three different strategies could be employed for therapeutic targeting of human thermogenic adipocytes: increasing BAT mass through stimulation of BAT progenitors, increasing BAT function through regulatory pathways, and increasing WAT browning through promotion of beige adipocyte formation. However, these strategies require deeper understanding of human brown and beige adipocytes. While murine studies have greatly increased our understanding of BAT, it is becoming clear that human BAT does not exactly resemble that of the mouse, highlighting the need for human in vitro models of brown adipocytes. Several different human brown adipocyte models will be discussed here, along with the potential to improve brown adipocyte culture through recreation of the BAT microenvironment.

Adipose-derived exosomal miR-210/92a cluster inhibits adipose browning via the FGFR-1 signaling pathway in high-altitude hypoxia

Cold and hypoxia are critical drivers of adaptation to high altitudes. Organisms at high altitudes have adapted to maximize the efficiency of oxygen utilization and are less prone to obesity and diabetes than those at low altitudes. Brown adipose tissue (BAT) dissipates energy in the form of heat in both humans and rodents; it also serves to regulate metabolism to curb obesity. However, the role of BAT in high-altitude populations is poorly understood. Serum exosomes can be easily obtained, enabling the study of BAT functions and identification of biomarkers in serum exosomes, both of which contribute to understanding the role of BAT in high-altitude populations. 18F-Fluorodeoxyglucose (18F-FDG) positron emission tomography integrated with computed tomography (PET/CT) is the gold standard for studying BAT in human adults. Here, we studied BAT in healthy high-altitude populations via PET/CT and serum exosomal microRNAs (miRNAs). The observations were validated in mouse tissues and demonstrated that high-altitude hypoxia activated BAT through attenuated white adipose tissue (WAT) secreted exosomal miR-210/92a, which enhanced the FGFR-1 expression in BAT.

Magnetic Resonance Imaging Techniques for Brown Adipose Tissue Detection.

Magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS) methods can non-invasively assess brown adipose tissue (BAT) structure and function. Recently, MRI and MRS have been proposed as a means to differentiate BAT from white adipose tissue (WAT) and to extract morphological and functional information on BAT inaccessible by other means. Specifically, proton MR (1H) techniques, such as proton density fat fraction mapping, diffusion imaging, and intermolecular multiple quantum coherence imaging, have been employed to access BAT microstructure; MR thermometry, relaxometry, and MRI and MRS with 31P, 2H, 13C, and 129Xe have shown to provide complementary information on BAT function. The purpose of the present review is to provide a comprehensive overview of MR imaging and spectroscopy techniques used to detect BAT in rodents and in humans. The present work discusses common challenges of current methods and provides an outlook on possible future directions of using MRI and MRS in BAT studies.

Brown adipose tissue ameliorates autoimmune arthritis via inhibition of Th17 cells

The functions of adipose tissue are associated with autoimmune diseases, such as rheumatoid arthritis (RA). Some studies have shown that the three compositions of adipose tissue (white, brown, and beige) have different functions. Brown adipose tissue (BAT) is known to secrete several factors that differ from those in white adipose tissue. This suggests that BAT might have potential positive advantages in the physiology of autoimmune diseases. We compared the functions of collagen-induced arthritis mice-derived BAT (CIA BAT) with normal mice-derived BAT. DBA/1J mice (6–7 weeks of age) were immunized by intradermal injection at the base of the tail with 100 μg of bovine type II collagen (CII) emulsified in complete Freund’s adjuvant. Immunized mice then received booster immunizations by intraperitoneal injection with 100 μg of CII in incomplete Freund’s adjuvant. We transplanted CIA BAT and normal BAT into CIA recipient mice. After transplantation, we measured the functions of CIA BAT and normal BAT in mice. Normal BAT-transplanted mice showed significantly lower scores of bone damage, inflammation, and cartilage damage. The proinflammatory cytokines in normal BAT-transplanted mice, such as IL-12, IL-17, IL-6, and tumor necrosis factor-α (TNF-α), tended to decrease. Microarray analysis showed that the PI3K-AKT signaling pathway and IL-17 levels of CIA BAT tissues were significantly higher than those of normal BAT tissues. These results suggest that the transplantation of normal brown fat may have a therapeutic effect in RA patients.

Innervation of supraclavicular adipose tissue: A human cadaveric study.

Functional brown adipose tissue (BAT) was identified in adult humans only in 2007 with the use of fluorodeoxyglucose positron emission tomography imaging. Previous studies have demonstrated a negative correlation between obesity and BAT presence in humans. It is proposed that BAT possesses the capacity to increase metabolism and aid weight loss. In rodents it is well established that BAT is stimulated by the sympathetic nervous system with the interscapular BAT being innervated via branches of intercostal nerves. Whilst there is evidence to suggest that BAT possesses beta-3 adrenoceptors, no studies have identified the specific nerve branch that carries sympathetic innervation to BAT in humans. The aim of this study was to identify and trace the peripheral nerve or nerves that innervate human BAT in the supraclavicular region. The posterior triangle region of the neck of cadaveric specimens were dissected in order to identify any peripheral nerve branches piercing and/or terminating in supraclavicular BAT. A previously undescribed branch of the cervical plexus terminating in a supraclavicular adipose depot was identified in all specimens. This was typically an independent branch of the plexus, from the third cervical spinal nerve, but in one specimen was a branch of the supraclavicular nerve. Histological analysis revealed the supraclavicular adipose depot contained tyrosine hydroxylase immunoreactive structures, which likely represent sympathetic axons. This is the first study that identifies a nerve branch to supraclavicular BAT-like tissue. This finding opens new avenues for the investigation of neural regulation of fat metabolism in humans.

NCLX prevents cell death during adrenergic activation of the brown adipose tissue

A sharp increase in mitochondrial Ca2+ marks the activation of brown adipose tissue (BAT) thermogenesis, yet the mechanisms preventing Ca2+ deleterious effects are poorly understood. Here, we show that adrenergic stimulation of BAT activates a PKA-dependent mitochondrial Ca2+ extrusion via the mitochondrial Na+/Ca2+ exchanger, NCLX. Adrenergic stimulation of NCLX-null brown adipocytes (BA) induces a profound mitochondrial Ca2+ overload and impaired uncoupled respiration. Core body temperature, PET imaging of glucose uptake and VO2 measurements confirm a thermogenic defect in NCLX-null mice. We show that Ca2+ overload induced by adrenergic stimulation of NCLX-null BAT, triggers the mitochondrial permeability transition pore (mPTP) opening, leading to a remarkable mitochondrial swelling and cell death. Treatment with mPTP inhibitors rescue mitochondrial function and thermogenesis in NCLX-null BAT, while calcium overload persists. Our findings identify a key pathway through which BA evade apoptosis during adrenergic stimulation of uncoupling. NCLX deletion transforms the adrenergic pathway responsible for thermogenesis activation into a death pathway.

Effects of Royal Jelly and Tocotrienol Rich Fraction in obesity treatment of calorie-restricted obese rats: a focus on white fat browning properties and thermogenic capacity

BackgroundObesity has reached an alarming rate worldwide. Promoting thermogenesis via increasing the function of brown adipose tissue (BAT) or white adipose tissue (WAT) browning has been proposed as a new protective approach against obesity. The goal of this study was to evaluate the effects of Royal Jelly (RJ) and tocotrienol rich fraction (TRF) on BAT activation and WAT browning during calorie restriction diet (CRD) in obesity model.MethodsIn this experimental study, 50 obese Wistar rats were randomly divided into 5 groups and then received one of the following treatments for a period of 8-week: High-fat diet (HFD), CRD, RJ + CRD, TRF + CRD, and RJ + TRF + CRD. Effects of RJ and TRF, individually and in combination on body weight and the expression of key thermoregulatory genes in WAT and BAT were examined by quantitative real-time (qRT-PCR). Also, morphological alterations were assessed by hematoxylin and eosin staining.ResultsRJ (− 67.21 g ±4.84 g) and RJ + TRF (− 73.29 g ±4.51 g) significantly reduced weight gain relative to the CRD group (− 40.70 g ±6.50 g, P < 0.001). In comparison with the CRD group, RJ and RJ + TRF remarkably enhanced the uncoupling protein1 (UCP1) expression in WAT (5.81, 4.72 fold, P < 0.001) and BAT (4.99, 4.75 fold, P < 0.001). The expression of PR domain containing 16(PRDM 16), cAMP response element-binding protein1 (CREB1), P38 mitogen-activated protein kinases (P38MAPK), and Bone morphogenetic protein8B (BMP8B) have significantly increased following RJ and RJ + TRF treatments (P < 0.001). However, the expression levels of CCAAT/enhancer-binding protein beta (CEBPβ) and Bone morphogenetic protein7 (BMP7) did not remarkably change. Multilocular beige cells in WAT and compacted dense adipocytes were also observed in BAT of RJ and RJ + TRF received groups. TRF showed no substantial effects on the expression of the mentioned thermoregulatory genes and brown fat-like phenotype.ConclusionOur results suggest that, Royal Jelly promotes thermogenesis and browning of WAT, contributing to an increase in energy expenditure. Thus, Royal Jelly may give rise to a novel dietary choice to attenuate obesity.

Omega-3 Fatty Acids and Adipose Tissue: Inflammation and Browning.

White adipose tissue (WAT) and brown adipose tissue (BAT) are involved in whole-body energy homeostasis and metabolic regulation. Changes to mass and function of these tissues impact glucose homeostasis and whole-body energy balance during development of obesity, weight loss, and subsequent weight regain. Omega-3 polyunsaturated fatty acids (ω-3 PUFAs), which have known hypotriglyceridemic and cardioprotective effects, can also impact WAT and BAT function. In rodent models, these fatty acids alleviate obesity-associated WAT inflammation, improve energy metabolism, and increase thermogenic markers in BAT. Emerging evidence suggests that ω-3 PUFAs can also modulate gut microbiota impacting WAT function and adiposity. This review discusses molecular mechanisms, implications of these findings, translation to humans, and future work, especially with reference to the potential of these fatty acids in weight loss maintenance.

Wu-Mei-Wan prevents high-fat diet-induced obesity by reducing white adipose tissue and enhancing brown adipose tissue function

BackgroundWu-Mei-Wan, a classic traditional Chinese herb medicine, is one of the most important formulations to treat digestive diseases from ancient times to the present. Our previous study showed that WMW treatment can prevent T2DM in db/db mice, which motivating the application of WMW on metabolic disorders. PurposeObesity and its comorbid diseases have increased dramatically and are now a worldwide health problem. There is still a lack of satisfactory treatment strategies for obesity. This work was designed to assess the effect and related mechanism of WMW on high fat diet (HFD)-induced obese mice model. MethodsObese mice were induced by HFD. Thetherapeutic effect of WMW were analyzed by examining body and adipose tissue weight, metabolic profile and energy expenditure. Adipose tissue phenotype was determined by histological staining and the mitochondrial content was examined by transmission electron microscopy (TEM). Immunohistochemical and immunofluorescence staining, RT-qPCR and Western blot analysis were used to evaluate expression of key molecules in adipose tissue. ResultsWMW treatment significantly protects HFD-induced obesity. Here we showed that WMW limits weight gain, improves metabolic profile and increases energy expenditure. WMW inhibits the hypertrophy and hyperplasia of white adipocytes, the mechanism involving the inhibition of TLR3/IL-6/JAK1/STAT3 pathway. In brown adipose tissue (BAT), WMW promotes thermogenicprogramme without affecting cell proliferation. The activated BMP7/ Smad1/5/9 pathway is considered to be one of the explanations for the effect of WMW on BAT. ConclusionOur results suggested that WMW can prevent obesity and its underlying mechanisms are associated with reducing white adipose tissue and enhancing brown adipose tissue function.

1701-P: The Role of UGT1a in Regulating Brown Adipose Tissue and Thermogenic Function

Increasing biological thermogenesis (the conversion of chemical energy to heat) has been proposed as an approach towards lowering blood glucose, improving insulin sensitivity and mitigating type 2 diabetes. Non-shivering thermogenesis occurs primarily in adipocytes that express uncoupling protein 1 (UCP1) including brown adipose tissue (BAT) and beige adipose tissue but does not substantially occur in white adipose tissue (WAT). To identify genes that regulate thermogenesis and BAT function, we recently compared genes expressed in paired human patient samples (n = 10) of the deep neck BAT located next to the carotid sheath and superficial subcutaneous WAT in the neck. We found UDP-glucuronide transferase 1A (UGT1A) is co-expressed in individuals with a high amount of the thermogenic protein uncoupling protein 1 (UCP1). This enzyme is known to glucuronidate metabolites, including bilirubin and xenobiotics, which increases their water solubility and excretion. Based on this function, we hypothesize UGT1A regulates BAT not through direct action of the enzyme but through modulating the availability of thermogenic molecules. Here, we characterized expression of different UGT1A isoforms in vivo and in vitro. Cold exposure is an effective method for activating BAT and increasing thermogenesis in mice. Interestingly, expression of multiple UGT1A isoforms, including UGT1A6a and UGT1A6b, are decreased in BAT of mice exposed to a cold environment (5°C) for 7 days. This finding is consistent with data in multiple publicly available datasets which have shown UGT1A6a/6b are decreased in cold exposure and other thermogenic stimuli. In vitro, most UGT1A isoforms were increased during the differentiation of murine brown adipocytes but only certain isoforms were differentially expressed during in vitro differentiation in human brown adipocytes. Based on these data, we propose that UGT1A may have a function related to thermogenesis and understanding its function could lead to novel approaches for activating BAT. Disclosure S. Kodani: None. M. Lynes: None. F. Shamsi: None. C. Wang: None. S. Sugimoto: None. R. Xue: None. T. Huang: None. A.P. White: None. A.M. Cypess: None. Y. Tseng: Consultant; Self; Boehringer Ingelheim International GmbH. Funding National Institutes of Health (R01DK07709, R01DK102898, T32DK007260)

42-OR: The Impact of GSNOR-Mediated Nitroso-Redox Signaling on Immuno-Metabolic Interaction in the Brown Adipose Tissue

The thermogenic function of brown adipose tissue (BAT) is tightly regulated by cellular redox status, however, the molecular mechanism underlying this regulation is largely unknown. S-nitrosoglutathione reductase (GSNOR, Adh5) is a major denitrosylase that balances intracellular nitroso-redox status. Although GSNOR dysregulation has been implicated in the pathogenesis of many chronic diseases, the pathophysiological role of GSNOR in the BAT is completely unknown. We found cold exposure increased murine BAT GSNOR while diet-induced obesity (DIO) suppressed it, indicating a potential role for GSNOR in metabolic adaptation in the BAT. Mechanistically, we found that the transcription factor heat shock factor 1 (HSF1) occupies the Adh5 promoter activating Adh5 expression, while DIO inhibited the HSF1-GSNOR axis in the BAT. Notably, administration of a HSF1 enhancer to mice with DIO significantly improved glucose intolerance and BAT mitochondrial dysfunciton. Furthermore, GSNOR deletion in the BAT (GSNORBKO) suppressed UCP1-dependent respiration under both thermoneutral and cold acclimation conditions, as well as elevated immune cell infiltration in the BAT. This was in part mediated by increased BAT nitrosative stress and UCP1 S-nitrosylation induced by GSNOR deletion. Moreover, GSNORBKO mice have worsened DIO-induced glucose intolerance and decreased UCP1-mediated oxygen consumption. In contrast, gain-of GSNOR function in the BAT improved DIO-associated glucose intolerance, increased UCP1-dependent oxygen consumption and decreased DIO-induced inflammation. These data provide novel evidence that GSNOR plays a protective role in BAT against metabolic stressors. Ultimately, insights into the mechanisms by which dysregulation of nitroso-redox signaling impairs BAT metabolic homoestasis and drives the pathologies associated with obesity are expected to accelerate the development of novel therapeutic targets for metabolic diseases. Disclosure S. Sebag: None. Q. Qian: None. Z. Zhang: None. M. Li: None. V.A. Lira: None. M.J. Potthoff: None. L. Yang: None. Funding American Diabetes Association (1-18-IBS-149 to L.Y.)

1945-P: Pioglitazone Promotes the Function of White and Brown Adipose Tissue in Diet-Induced Obese Mice

PPARγ agonist pioglitazone is an insulin sensitizer in the treatment of type 2 diabetes. Here, we investigate the effects of pioglitazone on white and brown fat in diet-induced obese mice. After 16-week diet intervention, C57BL/6 mice were divided randomly into three groups: normal chow diet group (NC group), high-fat diet group (HFD group) and high-fat diet + pioglitazone group (HFDP group). Pioglitazone (30-40mg/kg/day) was administered in drinking water for 6 weeks. After 6-week pioglitazone treatment, there was no significant difference in both body weight and fat mass between the HFDP group and the HFD group. However, fasting blood glucose, glucose tolerance and insulin sensitivity were significantly improved in HFDP group. What’s more, the weight of liver and epididymal adipose tissue (eWAT) were not reduced by pioglitazone treatment. However, the weight of brown adipose tissue (BAT) in the HFDP group was significantly higher than HFD group. Histological staining showed that the hepatic steatosis was ameliorated by pioglitazone. Quantitative real-time PCR revealed that mRNA levels of heat-producing genes (PRDM16, FGF-21, UCP1) and mitochondrial biosynthesis and functional genes (Tfam, cidea, cox7a1, Fabp4) were upregulated in eWAT by pioglitazone. Western blotting results also showed that the protein expression of UCP1 and PGC1α was upregulated by pioglitazone both in eWAT and in BAT when compared with the HFD group. To sum up, our study revealed that pioglitazone could promote the browning of WAT and activate the function of BAT with improved hepatic steatosis in diet-induced obese mice. Disclosure F. Xu: None. W. Wang: None. X. Zheng: None. W. Guo: None. Y. Shen: None.

1498-P: A Feedback Loop between miRNA Processing Complex and miR-3975 in Western Diet–Induced Brown Adipose Tissue Dysfunction

Brown adipose tissue (BAT) is considered as promising therapeutic targets for obesity and metabolic disorder through modulation of energy homeostasis. microRNAs (miRNAs) are established as essential modulators of biological processes in many tissues, including the differentiation and function of BAT. Here we demonstrated that high-fat/high fructose associated dysfunction of BAT coincides with impaired miRNA-processing pathway in a feedback loop with miR-3975 activation. Eight-week-old male C57BL/6 mice were fed a low-fat diet (10% fat and 17% sucrose; LFD) and high fat-high fructose (45% fat, 17% sucrose, and 17% fructose; HFHFD) for 4, 12, and 20 weeks (n=6-9 per group). We evaluated expression levels of apoptosis markers, thermogenesis markers, miRNA biogenesis regulators, and miRNAs in BAT. Mice fed an HFHFD showed a gradually significance increased body weight and brown adipose tissue weight when compared to the mice fed with LFD. BAT of HFHFD fed mice has a pale appearance versus feeding period matched LFD fed mice. The protein expression of thermogenesis markers, UCP1 (P &amp;lt;0.05), and PGC1α (P &amp;lt;0.05) were shown to decrease after 12 weeks of HFHFD diet feeding. The apoptosis marker, the Bax/Bcl-2 ratio, was upregulated at 12 weeks (P &amp;lt; 0.01). Mice fed HFHFD for 12 and 20 weeks showed a significant decrease in Dicer (P &amp;lt; 0.01 for 12 and 20 weeks) and TRBP2 (P &amp;lt; 0.01 for 12 weeks and P &amp;lt; 0.0001 for 20 weeks). miR-3975, which is predicted miRNA to regulate both Dicer and TRBP2 by IPA was activated at 12 weeks (P &amp;lt; 0.01), while inactivated at 20 weeks (P &amp;lt; 0.001). Our results suggest that miR-3975 acts as a component of a negative feedback loop that blocks miRNA biogenesis machinery through the inhibition of Dicer and TRBP2 expression. Therefore, long-term HFHFD may disturb miRNA processing and contribute to brown adipose tissue dysfunction and apoptosis by modulation of miRNA expression. We are currently investigating the novel feedback loop in brown adipocyte primary cells. Disclosure M. Park: None. S. Meruvu: None. J. Zhang: None. M. Choudhury: None.

1709-P: Increased Adiponectin Levels Are Associated with Impaired Brown Adipose Tissue in Mice

Brown adipose tissue (BAT) thermogenesis affects energy expenditure, as well as glucose and lipid clearance. Thus, making BAT an attractive target in obesity/type-2 diabetes. Adiponectin (Apn) regulates whole-body metabolism and lowered Apn levels are associated with obesity and increased type-2 diabetes risk. Apn has been suggested to be required for maintaining normal body temperature in cold environment, but the role of Apn in BAT functionality is not clear. Here, we have studied BAT in adiponectin overexpressing (Apn-Tg) mice. During unchallenged chow-diet conditions, BAT of Apn-Tg mice was enlarged, fibrotic and more whitish. This phenotype was, as judged by tracer experiments, associated with lower lipid and glucose uptake, decreased de novo lipogenesis and decreased fatty acid oxidation in BAT. High-fat diet (HFD)-induced obesity increased these differences between genotypes. The BAT mRNA expression of thermogenesis markers such as UCP-1 and DIO-2 were down-regulated in BAT of obese, but not lean, Apn-Tg mice. Moreover, Seahorse-based analysis revealed reduced mitochondrial function in Apn-Tg BAT in ex vivo-differentiated brown adipocytes. To test whether increased Apn levels affect cold-induced BAT activation, we exposed Apn-tg and littermate control mice to cold temperature (6°C) for four weeks. Under this setting, the BAT lipid and glucose uptake, and body temperature were similar between genotypes. Accordingly, UCP-1 and DIO-2 mRNA levels were not different between Apn-Tg and WT mice, and highly induced by chronic cold exposure. Nevertheless, Seahorse data from isolated BAT mitochondria from cold-exposed animals still indicate mitochondrial dysfunction in Apn-Tg mice. Thus, our data suggest that increased adiponectin levels are associated with metabolic dysfunction in BAT. We propose that the increased BAT expansion in Apn-Tg mice occurs at the expense of the mitochondrial function. Disclosure B. Chanclon: None. M. Bauza-Thorbrugge: None. A. Benrick: None. I. Wernstedt-Asterholm: None.

41-OR: Single Cell RNA-Sequencing Reveals the Developmental Heterogeneity of Brown Adipocytes

Brown adipose tissue (BAT) is the major thermogenic organ, which promotes energy expenditure through a process called adaptive thermogenesis. Central to the inherent thermogenic activity of brown adipocytes is uncoupling protein 1(UCP1). Other than mature adipocytes, several other cell types form the adipose niche and play important roles in BAT function and remodeling. Prolonged cold exposure increases BAT mass and activity through de novo recruitment of brown adipocytes as well as coordinated expansion of other cells within the adipose niche to enable maximal thermogenic activity. However, the source of cold-induced brown adipocytes and the molecular mechanism regulating BAT expansion is not known. To determine the cellular composition of BAT and its remodeling in response to ambient temperature, we performed single-cell RNA-sequencing (scRNA-seq) of the stromal vascular fraction of BAT from mice housed at different temperatures. The unsupervised clustering identified 19 major cell types, including several sub-populations of adipose progenitors, endothelial cells, immune cells, vascular smooth muscles, and Schwann cells. The transcriptional profile of each cell type in mice housed at different temperatures revealed the dynamic remodeling of brown adipocyte niche in response to environmental challenges. Among them, we identified a novel population of brown adipose progenitors derived from BAT endothelium. These endothelial-derived adipose progenitors have a distinct gene expression profile, which distinguishes them from the previously known Pdgfra-expressing adipose progenitors. Using an inducible genetic reporter, we validated the induction of UCP1 and other thermogenic gene program in the endothelial-derived adipose progenitors upon cold exposure. In summary, scRNA-seq of BAT provides a high-resolution map of brown adipose niche and suggests the critical contribution of each cell type to BAT functions, homeostasis, and turn-over. Disclosure F. Shamsi: None. M. Piper: None. L. Ho: None. T. Huang: None. Y. Tseng: Consultant; Self; Boehringer Ingelheim International GmbH. Funding American Diabetes Association (1-18-PDF-169 to F.S.); National Institutes of Health (R01DK077097, R01DK102898, P30DK036836); Harvard Stem Cell Institute

46-OR: The Transcriptional Coregulator Ldb1 Directly Regulates Brown Adipose Thermogenic Function

Brown adipose tissue (BAT) is critical for mammalian thermogenesis. BAT utilizes fatty acids and glucose for heat production via mitochondrial uncoupling and is thus an attractive therapeutic target for combatting obesity. Exploiting the energy uncoupling capacity of this tissue will require a greater understanding of underlying BAT transcriptional mechanisms. Our prior work using a global deficiency model supported a novel role for the LIM domain-binding protein 1 (Ldb1) transcriptional co-regulator in regulating BAT function. However, direct roles for Ldb1 expressed in brown adipocytes has not been tested. Ldb1 acts as a scaffold to promote the assembly of transcriptional complexes and is important for the development and function of several metabolic tissues. We hypothesized that Ldb1 maintains BAT thermogenic function through direct regulation of thermogenic genes, like Ucp1. Loss of Ldb1 in primary brown adipocytes resulted in reduced expression of BAT-selective genes including Ucp1 and Elovl3 and the inability to induce Ucp1 expression upon Beta-3 adrenergic receptor agonism. This result was replicated using a siLdb1-transfected Ucp1-expressing cell line. To test Ldb1 roles on systemic thermogenesis, we developed a mouse model, termed Ldb1ΔBAT, where Ldb1 was reduced in thermogenic adipocytes using a Ucp1-driven Cre recombinase. Similar to our in vitro observations, Ldb1ΔBAT mice had reduced mRNAs of key BAT genes, including Ucp1 and Cidea. Through indirect calorimetry and a cold challenge, we uncovered significant impairments in energy expenditure and cold intolerance in Ldb1ΔBAT mice. To assess the transcriptional mechanisms underlying Ldb1 impacts on BAT thermogenesis, we conducted chromatin immunoprecipitation and found Ldb1 occupied upstream regulatory domains of Ucp1 in chromatin prepared from a BAT cell line and primary tissue. Overall, this work supports that Ldb1 is required for the thermogenic function of brown adipocytes. Disclosure J. Kepple: None. Y. Liu: None. T. Kim: None. K.M. Habegger: Consultant; Self; Glyscend, Inc. Research Support; Self; Daiichi Sankyo, Novo Nordisk Inc. M. Young: None. C.S. Hunter: None. Funding National Institutes of Health (R01DK111483, F31DK121414, T32GM109780)