UCP1 Protein Research Articles

Combined Phyllostachys pubescens and Scutellaria baicalensis Prevent High-Fat Diet-Induced Obesity via Upregulating Thermogenesis and Energy Expenditure by UCP1 in Male C57BL/6J Mice

This study examined the anti-obesity effects of a Phyllostachys pubescens (leaf) and Scutellaria baicalensis root mixture (BS21), and its underlying mechanisms of action, in high-fat diet (HFD)-induced obese mice. Mice were fed a HFD with BS21 (100, 200, or 400 mg/kg) for 9 weeks. BS21 reduced body weight, white adipose tissue (WAT) and liver weights, liver lipid accumulation, and adipocyte size. Additionally, BS21 reduced serum concentrations of non-esterified fatty acid, triglyceride, glucose, lactate dehydrogenase, low-density lipoprotein cholesterol, total cholesterol, leptin, and insulin growth factor 1, but elevated the adiponectin concentrations. Furthermore, BS21 suppressed the mRNA levels of lipogenesis-related proteins, such as peroxisome proliferator–activated receptor (PPAR) γ, SREBP-1c, C/EBP-α, fatty acid synthase, and leptin, but increased the mRNA gene expression of lipolysis-related proteins, such as PPAR-α, uncoupling protein (UCP) 2, adiponectin, and CPT1b, in WAT. In addition, BS21 increased the cold-stimulated adaptive thermogenesis and UCP1 protein expression with AMPK activation in adipose tissue. Furthermore, BS21 increased the WAT and mRNA expression of energy metabolism-related proteins SIRT1, PGC-1α, and FNDC5/irisin in the quadriceps femoris muscle. These results suggest that BS21 exerts anti-obesity and antihyperlipidemic activities in HFD-induced obese mice by increasing the thermogenesis and energy expenditure, and regulating lipid metabolism. Therefore, BS21 could be useful for preventing and treating obesity and its related metabolic diseases.

Specific adipose tissue Lbp gene knockdown prevents diet-induced body weight gain, impacting fat accretion-related gene and protein expression

Lipopolysaccharide binding protein (Lbp) has been recently identified as a relevant component of innate immunity response associated to adiposity. Here, we aimed to investigate the impact of adipose tissue Lbp on weight gain and white adipose tissue (WAT) in male and female mice fed an obesogenic diet. Specific adipose tissue Lbp gene knockdown was achieved through lentiviral particles containing shRNA-Lbp injected through surgery intervention. In males, WAT Lbp mRNA levels increased in parallel to fat accretion, and specific WAT Lbp gene knockdown led to reduced body weight gain, decreased fat accretion-related gene and protein expression, and increased inguinal WAT basal lipase activity, in parallel to lowered plasma free fatty acids, leptin, triglycerides but higher glycerol levels, resulting in slightly improved insulin action in the insulin tolerance test. In both males and females, inguinal WAT Lbp gene knockdown resulted in increased Ucp1 and Ppargc1a mRNA and Ucp1 protein levels, confirming adipose Lbp as a WAT browning repressor. In perigonadal WAT, Lbp gene knockdown also resulted in increased Ucp1 mRNA levels, but only in female mice, in which it was 500-fold increased. These data suggest specific adipose tissue Lbp gene knockdown as a possible therapeutic approach in the prevention of obesity-associated fat accretion.

Liraglutide Activates Type 2 Deiodinase and Enhances β3-Adrenergic-Induced Thermogenesis in Mouse Adipose Tissue.

AimsLiraglutide is a long-acting glucagon-like peptide 1 (GLP-1) receptor agonist used as an anti-hyperglycemic agent in type 2 diabetes treatment and recently approved for obesity management. Weight loss is attributed to appetite suppression, but therapy may also increase energy expenditure. To further investigate the effect of GLP-1 signaling in thermogenic fat, we assessed adipose tissue oxygen consumption and type 2 deiodinase (D2) activity in mice treated with liraglutide, both basally and after β3-adrenergic treatment.MethodsMale C57BL/6J mice were randomly assigned to receive liraglutide (400 μg/kg, n=12) or vehicle (n=12). After 16 days, mice in each group were co-treated with the selective β3-adrenergic agonist CL316,243 (1 mg/kg, n=6) or vehicle (n=6) for 5 days. Adipose tissue depots were assessed for gene and protein expression, oxygen consumption, and D2 activity.ResultsLiraglutide increased interscapular brown adipose tissue (iBAT) oxygen consumption and enhanced β3-adrenergic-induced oxygen consumption in iBAT and inguinal white adipose tissue (ingWAT). These effects were accompanied by upregulation of UCP-1 protein levels in iBAT and ingWAT. Notably, liraglutide increased D2 activity without significantly upregulating its mRNA levels in iBAT and exhibited additive effects to β3-adrenergic stimulation in inducing D2 activity in ingWAT.ConclusionsLiraglutide exhibits additive effects to those of β3-adrenergic stimulation in thermogenic fat and increases D2 activity in BAT, implying that it may activate this adipose tissue depot by increasing intracellular thyroid activation, adding to the currently known mechanisms of GLP-1A-induced weight loss.

Inhibitory effect of androgens on white adipose tissue thermogenic capacity

White adipose tissue (WAT) browning has gained interest due to its impact in obesity. Here, we evaluated the effect of androgens on the Ucp1-dependent thermogenic process from inguinal (IAT) and retroperitoneal (RPAT) WAT. Surgically androgens depleted rats (ODX) showed basal thermogenic activation (room temperature) in both WAT depots, which expressed higher levels of Ucp1, Prdm16 and Pgc1a. WAT pads from ODX cold-exposed rats (ODX-C) expressed increased levels of Ucp1 and Pgc1a and showed high UCP1 protein content. In primary beige adipocyte cultures, testosterone decreased the mitochondrial marker Cox8b and mitochondrial content. Finally, testosterone and dihydrotestosterone (DHT) decreased the expression of Ucp1, Pcg1a and Prdm16 in forskolin-stimulated beige adipocytes, an effect that was prevented by the antiandrogen flutamide. In conclusion, androgen deficient rats developed WAT depots with enhanced basal and cold-stimulated thermogenic activity. Additionally, in vitro androgen treatments inhibited the thermogenic program, effect which was mediated by the androgen receptor pathway.

Integrin αV Mediates the Effects of Irisin on Human Mature Adipocytes

Introdution and Aims: The myokine irisin is critical to modulating adipocytes thermogenesis and influence whole-body metabolism. However, whether there is difference in the effects of irisin on adipocytes derived from different depots remains unknown, and the receptor of irisin on adipocytes is still unclear. In this study, we determine the browning effect of irisin on adipocytes of subcutaneous and visceral human adipose tissue and explore the possibility that integrin αV was the receptor of irisin on human adipocytes. Methods: Human adipose-derived stem cells were isolated from human subcutaneous and visceral white adipose tissues and induced to differentiate into mature adipocytes, and the expression of UCP1 and thermogenic genes in mature adipocytes were examined with or without irisin treatment and compared between groups of different adiposity and different spots. Immunoprecipitation analysis was used to detect the interaction between irisin and integrin αV on adipocytes, and the protein expression of integrin αV in adipocytes was also compared between groups of different adiposity and anatomic position. Results: Irisin treatment could increase the expression level of beige adipocyte marker protein UCP1 and specific thermogenic genes in mature adipocytes derived from subcutaneous white adipose tissue but not in visceral adipose tissue. The results of immunoprecipitation showed that irisin could be attached to integrin αV on mature adipocytes, and there was no significant difference in the gene and protein expression of integrin αV in adipocytes, either derived from subcutaneous and visceral adipose tissue, or derived from obese and normal-weight individuals. Conclusion: The results of the present study indicated that irisin contributed to the transformation of mature white adipocytes to beige adipocytes in human subcutaneous adipose tissue but not in visceral adipose tissue. Integrin αV may mediate the browning effects of irisin on human mature adipocytes, which could provide the potential therapeutic targets for obesity and metabolic syndrome by promoting human brown adipose tissue activity.

The Effect of a Sustained High-Fat Diet on the Metabolism of White and Brown Adipose Tissue and Its Impact on Insulin Resistance: A Selected Time Point Cross-Sectional Study.

(1) Background: studies on the long-term dynamic changes in fat depot metabolism in response to a high-fat diet (HFD) on hepatic lipid deposition and insulin resistance are sparse. This study investigated the dynamic changes produced by HFD and the production of dysfunctional fat depots on insulin resistance and liver lipid metabolism. (2) Methods: mice fed a chow or HFD (45% kcal fat) diet had three fat depots, liver, and blood collected at 6, 10, 20, and 30 weeks. Anthropometric changes and gene markers for adipogenesis, thermogenesis, ECM remodeling, inflammation, and tissue insulin resistance were measured. (3) Results: early responses to the HFD were increased body weight, minor deposition of lipid in liver, increased adipocyte size, and adipogenesis. Later changes were dysfunctional adipose depots, increased liver fat, insulin resistance (shown by changes in ITT) accompanied by increased inflammatory markers, increased fibrosis (fibrosis > 2-fold, p < 0.05 from week 6), and the presence of crown cells in white fat depots. Later, changes did not increase thermogenic markers in response to the increased calories and decreased UCP1 and PRDM16 proteins in WAT. (4) Conclusions: HFD feeding initially increased adipocyte diameter and number, but later changes caused adipose depots to become dysfunctional, restricting adipose tissue expansion, changing the brown/beige ratios in adipose depots, and causing ectopic lipid deposition and insulin resistance.

Cyanidin-3-O-glucoside Regulates the Expression of Ucp1 in Brown Adipose Tissue by Activating Prdm16 Gene.

(1) Background: Brown adipose tissue (BAT) burns energy to produce heat. Cyanidin-3-O-glucoside (C3G) can then enhance the thermogenic ability of BAT in vivo. However, the mechanism by which C3G regulates Ucp1 protein expression remains unclear. (2) Methods: In this study, C3H10T12 brown adipose cells and db/db mice and mice with high-fat, high-fructose, diet-induced obesity were used as the model to explore the effect of C3G on the expression of the Ucp1 gene. Furthermore, the 293T cell line was used for an in vitro cell transgene, a double luciferase reporting system, and yeast single hybridization to explore the mechanism of C3G in regulating Ucp1 protein. (3) Results: we identified that, under the influence of C3G, Prdm16 directly binds to the −500 to −150 bp promoter region of Ucp1 to activate its transcription and, thus, facilitate BAT programming. (4) Conclusions: This study clarified the mechanism by which C3G regulates the expression of the Ucp1 gene of brown fat to a certain extent.

Mitochondrial uncoupling protein 1 antagonizes atherosclerosis by blocking NLRP3 inflammasome-dependent interleukin-1β production.

Mitochondrial uncoupling protein 1 (UCP1) is the hallmark of brown adipocytes responsible for cold- and diet-induced thermogenesis. Here, we report a previously unidentified role of UCP1 in maintaining vascular health through its anti-inflammatory actions possibly in perivascular adipose tissue. UCP1 deficiency exacerbates dietary obesity-induced endothelial dysfunction, vascular inflammation, and atherogenesis in mice, which was not rectified by reconstitution of UCP1 in interscapular brown adipose tissue. Mechanistically, lack of UCP1 augments mitochondrial membrane potential and mitochondrial superoxide, leading to hyperactivation of the NLRP3-inflammasome and caspase-1–mediated maturation of interleukin-1β (IL-1β). UCP1 deficiency–evoked deterioration of vascular dysfunction and atherogenesis is reversed by IL-1β neutralization or a chemical mitochondrial uncoupler. Furthermore, UCP1 knockin pigs (which lack endogenous UCP1) are refractory to vascular inflammation and coronary atherosclerosis. Thus, UCP1 acts as a gatekeeper to prevent NLRP3 inflammasome activation and IL-1β production in the vasculature, thereby conferring a protective effect against cardiovascular diseases.

Macrophages expressing uncoupling protein 1 increase in adipose tissue in response to cold in humans

Acute cold induces beige adipocyte protein marker expression in human subcutaneous white adipose tissue (SC WAT) from both the cold treated and contralateral leg, and the immune system regulates SC WAT beiging in mice. Cold treatment significantly increased the gene expression of the macrophage markers CD68 and 86 in SC WAT. Therefore, we comprehensively investigated the involvement of macrophages in SC WAT beiging in lean and obese humans by immunohistochemistry. Cold treatment significantly increased CD163/CD68 macrophages in SC WAT from the cold treated and contralateral legs of lean and obese subjects, and had similar effects on CD206/CD68 macrophages, whereas the effects on CD86/CD68 macrophages were inconsistent between lean and obese. However, linear regression analysis did not find significant relationships between the change in macrophage numbers and the change in UCP1 protein abundance. A high percentage of CD163 macrophages in SC WAT expressed UCP1, and these UCP1 expressing CD163 macrophages were significantly increased by cold treatment in SC WAT of lean subjects. In conclusion, our results suggest that CD163 macrophages are involved in some aspect of the tissue remodeling that occurs during SC WAT beiging in humans after cold treatment, but they are likely not direct mediators of the beiging process.

Effects of In Vitro Muscle Contraction on Thermogenic Protein Levels in Co-Cultured Adipocytes.

The crosstalk between the exercising muscle and the adipose tissue, mediated by myokines and metabolites, derived from both tissues during exercise has created a controversy between animal and human studies with respect to the impact of exercise on the browning process. The aim of this study was to investigate whether co-culturing of C2C12 myotubes and 3T3-L1 adipocytes under the stimuli of electrical pulse stimulation (EPS) mimicking muscle contraction can impact the expression of UCP1, PGC-1a, and IL-6 in adipocytes, therefore providing evidence on the direct crosstalk between adipocytes and stimulated muscle cells. In the co-cultured C2C12 cells, EPS increased the expression of PGC-1a (p = 0.129; d = 0.73) and IL-6 (p = 0.09; d = 1.13) protein levels. When EPS was applied, we found that co-culturing led to increases in UCP1 (p = 0.044; d = 1.29) and IL-6 (p = 0.097; d = 1.13) protein expression in the 3T3-L1 adipocytes. The expression of PGC-1a increased by EPS but was not significantly elevated after co-culturing (p = 0.448; d = 0.08). In vitro co-culturing of C2C12 myotubes and 3T3-L1 adipocytes under the stimuli of EPS leads to increased expression of thermogenic proteins. These findings indicate changes in the expression pattern of proteins related to browning of adipose tissue, supporting the use of this in vitro model to study the crosstalk between adipocytes and contracting muscle.

Neuroprotection of retinal cells by Caffeic Acid Phenylethyl Ester（CAPE) is mediated by mitochondrial uncoupling protein UCP2

Oxidative stress due to mitochondrial produced reactive oxygen species is a major cause of damage seen in many retinal degenerative diseases. Caffeic acid phenylethyl ester （CAPE） is protective agent in multiple tissues and is reported to have anti-oxidant properties. Systemically applied CAPE protected retinal ganglion cells from ischemic injury induced by increased intraocular pressure. CAPE provided complete protection for ARPE19 retinal pigment epithelial cells against tert-butyl hydrogen peroxide and reduced both basal and LPS-stimulated ROS production. The major effect of CAPE was mediated by the mitochondrial uncoupling protein UCP2 since both pharmacological inhibition of UCP2 and siRNA-induced knockdown removed the ability of CAPE to block ROS production. Based on common structural features, CAPE may be acting as a mimetic of the natural UCP2 homeostatic regulator 4-hydroxy-2-nonenal. CAPE may provide a valuable tool to treat oxidative stress-related damage in retinal and other degenerative diseases.

Maternal high fructose diet exacerbates white adipose tissue thermogenic process in offspring upon exposure to cold temperature

AimAn adverse endogenous environment during early life predisposes to metabolic disorder development. We previously reported adverse metabolic and adipose tissue effects in adult male rats born to dams fed with a fructose-rich diet (FRD). The aim of this work was to determine the effect of a FRD consumed by the pregnant mother on the white adipose tissue (WAT) browning capacity of male offspring at adulthood. Main methodsAdult SD male offspring from control (C) and FRD-fed mothers were exposed during one week to a cold stimulus. WAT browning capacity was studied through in vivo and in vitro approaches. Key findingsAfter cold exposure, WAT browning was higher in fructose-programmed animals as evidenced by an increase in ucp-1 gene expression, protein levels, and higher UCP-1 positive foci. Moreover, pgc1-α gene expression was increased. In vitro studies showed a lower adipogenic capacity in cells of prenatally fructose-exposed animals differentiated with a white differentiation cocktail, while a higher ucp-1 expression was noted when their cells were treated with a pro-beige differentiation cocktail. SignificanceFor the first time we demonstrate that pre-natal fructose exposure predisposes programmed male rats to a higher WAT browning-induced response, under stimulated conditions, despite an apparent lower basal thermogenic capacity. These results should be considered in future studies to generate new therapeutic approaches to deal with adverse programming malnutrition effects.

Exploratory Study on Expression of Fatty Tissue in Gestational Diabetes Mouse.

Objective To investigate the changes of UCP1 protein in brown fat by establishing a model of gestational diabetes mellitus through intervention of high fat and carbohydrate diet. To explore the changes of UCP1 protein in brown fat in gestational diabetic mice, and analyze the characteristics of abnormal glucose metabolism in gestational diabetic mice and its relationship with changes in adipocyte morphology and insulin resistance. Methods Eighty C57BL/6J pregnant mice were randomly divided into a normal control group and a high-fat and high-sugar feeding group. The normal control group was fed a normal diet, while the high-fat and high-sugar group was fed a high-fat and high-sugar diet to establish a gestational diabetes mellitus (GDM) model. On the pregnancy days 0, 10, and 18, the weight and fasting glucose were measured. On the pregnancy day 18, the triglycerides (TG), the total cholesterol (TC), free fatty acid (FFA), insulin levels, and insulin resistance index (HOMA-IR) were measured or calculated. At the same time, brown fat UCP1 protein of the two groups of pregnant mice were measured using Western blot Observed the Adipose tissue pathological changes by staining HE. The adipocyte was observed, and the correlation was analyzed. Results Twenty-one pregnant mice reach the level of gestational diabetes diagnosis (FBG ≥ 5.1 mmol/L) in the high-fat and high-sugar diet group. On the pregnancy day 10 and 18, the fasting plasma glucose and the body weight significantly increased (P < 0.05). The total cholesterol, triglycerides, free fatty acid, insulin level, and insulin resistance index of the GDM group were also higher compared with that of the control group (P < 0.05). The adipocyte size significantly increased in the GDM mice. TG, TC, FFA, and body weight at 18 days of gestation were significantly correlated with HOMA-IR and single-adipocyte area in the GDM mice. HOMA-IR was significantly correlated with a single-adipocyte area. Compared with the normal control group, the UCP1 proteins of GDM mice decreased significantly and negatively correlated with body weight increase. Conclusion Feeding C57BL/6J pregnant mice with high fat and high sugar to establish a gestational diabetes mouse model has good stability and is similar to human gestational diabetes. The reduction of brown fat UCP1 protein in GDM mice has a certain correlation with obesity tendency and obvious insulin resistance.

Transcriptional response of key metabolic and stress response genes of a nuculanid bivalve, Lembulus bicuspidatus from an oxygen minimum zone exposed to hypoxia-reoxygenation.

Benthic animals inhabiting the edges of marine oxygen minimum zones (OMZ) are exposed to unpredictable large fluctuations of oxygen levels. Sessile organisms including bivalves must depend on physiological adaptations to withstand these conditions. However, as habitats are rather inaccessible, physiological adaptations of the OMZ margin inhabitants to oxygen fluctuations are not well understood. We therefore investigated the transcriptional responses of selected key genes involved in energy metabolism and stress protection in a dominant benthic species of the northern edge of the Namibian OMZ, the nuculanid clam Lembulus bicuspidatus,. We exposed clams to normoxia (~5.8ml O2 l-1), severe hypoxia (36h at ~0.01ml O2 l-1) and post-hypoxic recovery (24h of normoxia following 36h of severe hypoxia). Using newly identified gene sequences, we determined the transcriptional responses to hypoxia and reoxygenation of the mitochondrial aerobic energy metabolism (pyruvate dehydrogenase E1 complex, cytochrome c oxidase, citrate synthase, and adenine nucleotide translocator), anaerobic glycolysis (hexokinase (HK), phosphoenolpyruvate carboxykinase (PEPCK), phosphofructokinase, and aldolase), mitochondrial antioxidants (glutaredoxin, peroxiredoxin, and uncoupling protein UCP2) and stress protection mechanisms (a molecular chaperone HSP70 and a mitochondrial quality control protein MIEAP) in the gills and the labial palps of L. bicuspidatus. Exposure to severe hypoxia transcriptionally stimulated anaerobic glycolysis (including HK and PEPCK), antioxidant protection (UCP2), and quality control mechanisms (HSP70 and MIEAP) in the gills of L. bicuspidatus. Unlike UCP2, mRNA levels of the thiol-dependent mitochondrial antioxidants were not affected by hypoxia-reoxygenation stress. Transcript levels of marker genes for aerobic energy metabolism were not responsive to oxygen fluctuations in L. bicuspidatus. Our findings highlight the probable importance of anaerobic succinate production (via PEPCK) and mitochondrial and proteome quality control mechanisms in responses to oxygen fluctuations of the OMZ bivalve L.bicuspidatus. The reaction of L.bicuspidatus to oxygen fluctuations implies parallels to that of other hypoxia-tolerant bivalves, such as intertidal species.

Isothermal microcalorimetry measures UCP1-mediated thermogenesis in mature brite adipocytes

The activation of thermogenesis in adipose tissue has emerged as an important target for the development of novel anti-obesity therapies. Using multi-well isothermal microcalorimetry, we have demonstrated that mature murine brown and brite adipocytes produce quantifiable heat upon β3-AR stimulation, independently of any anaerobic mechanisms. Additionally, in brite adipocytes lacking UCP1 protein, β3-AR stimulation still induces heat production, albeit to a much lower extent than in their wildtype counterparts, suggesting that UCP1 is an essential component of adrenergic induced thermogenesis in murine brite adipocytes exvivo. Similarly, we could observe an increase in heat production in human-derived adipocytes (hMADS) upon β-AR stimulation. Collectively, these results establish the use of isothermal microcalorimetry as a sensitive and accurate technique for measuring thermogenic responses in intact mature brite adipocytes from murine and human origin.

Oncostatin M suppresses browning of white adipocytes via gp130-STAT3 signaling

ObjectiveObesity is associated with low-grade adipose tissue inflammation and locally elevated levels of several glycoprotein 130 (gp130) cytokines. The conversion of white into brown-like adipocytes (browning) may increase energy expenditure and revert the positive energy balance that underlies obesity. Although different gp130 cytokines and their downstream targets were shown to regulate expression of the key browning marker uncoupling protein 1 (Ucp1), it remains largely unknown how this contributes to the development and maintenance of obesity. Herein, we aim to study the role of gp130 cytokine signaling in white adipose tissue (WAT) browning in the obese state. MethodsProtein and gene expression levels of UCP1 and other thermogenic markers were assessed in a subcutaneous adipocyte cell line, adipose tissue depots from control or adipocyte-specific gp130 knockout (gp130Δadipo) mice fed either chow or a high-fat diet (HFD), or subcutaneous WAT biopsies from a human cohort of lean and obese subjects. WAT browning was modeled in vitro by exposing mature adipocytes to isoproterenol after stimulation with gp130 cytokines. ERK and JAK-STAT signaling were blocked using the inhibitors U0126 and Tofacitinib, respectively. ResultsInguinal WAT of HFD-fed gp130Δadipo mice exhibited significantly elevated levels of UCP1 and other browning markers such as Cidea and Pgc-1α. In vitro, treatment with the gp130 cytokine oncostatin M (OSM) lowered isoproterenol-induced UCP1 protein and gene expression levels in a dose-dependent manner. Mechanistically, OSM mediated the inhibition of Ucp1 via the JAK-STAT but not the ERK pathway. As with mouse data, OSM gene expression in human WAT positively correlated with BMI (r = 0.284, p = 0.021, n = 66) and negatively with UCP1 expression (r = −0.413, p < 0.001, n = 66). ConclusionsOur data support the notion that OSM negatively regulates thermogenesis in WAT and thus may be an attractive target for treating obesity.

Melatonin Enhances the Mitochondrial Functionality of Brown Adipose Tissue in Obese—Diabetic Rats

Developing novel drugs/targets remains a major effort toward controlling obesity-related type 2 diabetes (diabesity). Melatonin controls obesity and improves glucose homeostasis in rodents, mainly via the thermogenic effects of increasing the amount of brown adipose tissue (BAT) and increases in mitochondrial mass, amount of UCP1 protein, and thermogenic capacity. Importantly, mitochondria are widely known as a therapeutic target of melatonin; however, direct evidence of melatonin on the function of mitochondria from BAT and the mechanistic pathways underlying these effects remains lacking. This study investigated the effects of melatonin on mitochondrial functions in BAT of Zücker diabetic fatty (ZDF) rats, which are considered a model of obesity-related type 2 diabetes mellitus (T2DM). At five weeks of age, Zücker lean (ZL) and ZDF rats were subdivided into two groups, consisting of control and treated with oral melatonin for six weeks. Mitochondria were isolated from BAT of animals from both groups, using subcellular fractionation techniques, followed by measurement of several mitochondrial parameters, including respiratory control ratio (RCR), phosphorylation coefficient (ADP/O ratio), ATP production, level of mitochondrial nitrites, superoxide dismutase activity, and alteration in the mitochondrial permeability transition pore (mPTP). Interestingly, melatonin increased RCR in mitochondria from brown fat of both ZL and ZDF rats through the reduction of the proton leak component of respiration (state 4). In addition, melatonin improved the ADP/O ratio in obese rats and augmented ATP production in lean rats. Further, melatonin reduced mitochondrial nitrosative and oxidative status by decreasing nitrite levels and increasing superoxide dismutase activity in both groups, as well as inhibited mPTP in mitochondria isolated from brown fat. Taken together, the present data revealed that chronic oral administration of melatonin improved mitochondrial respiration in brown adipocytes, while decreasing oxidative and nitrosative stress and susceptibility of adipocytes to apoptosis in ZDF rats, suggesting a beneficial use in the treatment of diabesity. Further research regarding the molecular mechanisms underlying the effects of melatonin on diabesity is warranted.

Breastfeeding undernutrition changes iBAT-involved thermogenesis protein expression and leads to a lean phenotype in adult rat offspring

Nutritional insults early in life have been associated with metabolic diseases in adulthood. We aimed to evaluate the effects of maternal food restriction during the suckling period on metabolism and interscapular brown adipose tissue (iBAT) thermogenically involved proteins in adult rat offspring. Wistar rats underwent food restriction by 50% during the first two-thirds of lactation (FR50 group). Control rats were fed ad libitum throughout lactation (CONT group). At birth, the litter size was adjusted to eight pups, and weaning was performed at 22 days old. Body weight and food and water intake were assessed every two days. High- (HCD, 4,589 cal) and normal-caloric diet (NCD, 3,860 cal) preferences, as well as food intake during the dark part of the cycle, were assessed. At 100 days old, the rats were euthanized, and blood and tissues were removed for further analyses. Adult FR50 rats, although hyperphagic and preferring to eat HCD (P<.001), were leaner (P<.001) than the CONT group. The FR50 rats, were normoglycemic (P=.962) and had hypertriglyceridemia (P<.01). In addition, the FR50 rats were dyslipidemic (P<.01), presenting with a high atherogenic risk by the Castelli indexes (P<.01), had a higher iBAT mass (P<.01), fewer β3 adrenergic receptors (β3-AR, P<.05) and higher iBAT expression of uncoupled protein 1 (UCP1, P<.05) and peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α, P<.001) than the CONT rats. In conclusion, maternal food restriction during early breastfeeding programs rat offspring to have a lean phenotype, despite hyperphagia, and increased iBAT UCP1 and PGC-1α protein expression.

Obeticholic acid ameliorates obesity and hepatic steatosis by activating brown fat.

Obeticholic acid (OCA) is exemplified as a potent drug for treating primary biliary cirrhosis and nonalcoholic fatty liver disease by inhibiting bile acid synthesis. However, it remains unclear whether the effect of OCA is mediated by the function of brown adipose tissue (BAT). In the present study, brown adipogenesis differentiation in vitro and db/db mouse model treated with OCA were used to assess the anti-obesity function by body weight tracking, O2 consumption, food intake, physical activity, glucose tolerance tests. In addition, uncoupling protein 1 (Ucp1) protein expression in brown adipose tissue was measured by western blotting, morphometry of brown adipose tissue was analyzed by hematoxylin and eosin staining. Hepatic steatosis was detected by Oil-Red O staining and serological analysis was performed to assess the effect of OCA on hyperlipidemia. OCA treatment enhanced brown adipocyte cell differentiation and upregulated the expression of the BAT-specific gene Ucp1) in C3H10T1/2 cells in vitro. Consistent with these findings, OCA increased whole-body energy metabolism and glucose homeostasis by enhancing BAT activity in vivo, and ultimately decreased body weight gain in db/db mice. In addition, the results demonstrated that spontaneous hepatic steatosis in db/db mice was ameliorated following OCA treatment. In summary, OCA functioned as a BAT activator to help ameliorate obesity and maintain glucose homeostasis in db/db mice. The present results may provide a novel potential therapeutic approach to activate brown fat in patients with obesity and other metabolic disorders.

34-OR: Role of Arid5b in Thermogenesis and Adipose Tissue Browning

Background: Adipose tissue considerably influence metabolic homeostasis and play a central role in regulating whole-body energy and glucose metabolism. Previously, we have reported that mice with the global deletion of Arid5b are lean and have less fat in white and brown adipose tissues. However, this role is unknown in adipocyte specific Arid5b knockout (5bFKO) mice. Aim: We investigated the effects of adipose-specific deficiency of Arid5b on systemic glucose homeostasis and adipose tissue metabolism. Result: Deletion of Arid5b in adipose tissue using adiponectin promoter-driven Cre recombinase significantly improved glucose tolerance with significant reduction of insulin level. Body weight of 5bFKO mice was significantly reduced at the later age. Core body temperature of 5bFKO mice was significantly higher compared to the control mice. Robust increase in the protein expression of uncoupling protein 1 (Ucp1) was observed in inguinal white adipose tissue (iWAT) of 5bFKO mice housed at ambient temperature. This phenotype was further confirmed by prolonged cold exposure where Ucp1 and mitochondrial proteins such as pyruvate dehydrogenase (PDH), mitochondrially encoded cytochrome c oxidase (MTCO1) and ATP5A were substantially increased in iWAT of adipocyte specific Arid5b deficient mice. Gene expression of beige adipogenesis markers such as cell death inducing DFFA like effector A (Cidea) and iodothyronine deiodinase 2 (Dio2) were also significantly increased indicating browning of iWAT. Furthermore, loss of Arid5B in fat led to significant reduction in macrophages and total immune cells in iWAT. All together these results show that the deletion of Arid5b could promote browning of iWAT. Conclusion: These findings demonstrate that Arid5b regulates thermogenic genes expression to promote beige adipocytes formation and controls adipose inflammation. Inhibition of Arid5b may provide a novel therapeutic approach for obesity and diabetes. Disclosure M. Shukare: None. R. Huang: None. A. Ehsani: None. G. Zhang: None. J. Chalise: None. R. H. Whitson: None. K. Itakura: None.