UCP2 mRNA Research Articles

Tissue oxidative stress and expression of chicken UCP and ANT mRNA in laying hens exposed to acute cold stress

ABSTRACT 1. Exposure to stress alters normal homoeostasis and, hence, the antioxidant defence system. The aim of this study was to examine the effect of acute cold temperature on the antioxidant defence system in hens. 2. Hy-line grey commercial layers (80 40-week-old) were randomly assigned to one of eight groups. In groups 1 to 5, hens were exposed to low temperature at −8.68°C (cool stressed) for 2, 4, 6, 8 and 10 h, respectively. In groups 6 and 7, post 10 h cool stressed, hens were quickly transferred to room at 21°C to recovery for 2 h and 4h, respectively. In treatment groups 6 and 7, post 10 h cool stressed, hens were quickly transferred to room at 21°C for 2 h and 4 h, respectively. Group 8 was the control, where hens were housed under regular condition at 21°C as controls. 3. Antioxidant enzymes (T-AOC), superoxide dismutase (SOD), glutathione peroxidase (GPx) and malondialdehyde (MDA), in skeletal muscle, the kidney, liver and pancreas were measured. The transcription of avUCP and ANT mRNA was tested by RT-PCR. 4. The T-AOC activity was increased in the skeletal muscle of hens cold stressed for 2, 4, 6, 8 and 10 h and the 2 h recovery groups compared with control hens (p < 0.05). The GPx activity was increased in the liver and skeletal muscle after cold stress 4 h and in the pancreas of cold stress 2 h compared with the control group (p < 0.05). Antioxidant SOD activity was increased in the kidney after cold stress 6 h and in the liver after cold stress 10 h compared to the control group (p < 0.05). Measured MDA activity was increased in the pancreas after 2 h cold stress (p < 0.05). 5. UCP mRNA expression level was increased in the pectoral muscle for 2 h and 4 h recovery groups compared with the control hens (p < 0.05) and avian uncoupling protein (UPC), adenine nucleotide translocator (ANT) expression level was increased in the leg muscle of hens cold stress for 2, 6, 8 h and recovery 2 and 4 h. 6. The observed changes in the antioxidant defence system were tissue specific. Increments in levels of ANT (leg muscle) and UCP (pectoral and leg muscle) mRNA expression may be involved in the regulation of thermogenesis in skeletal muscle.

Translation complex profile sequencing allows discrimination of leaky scanning and reinitiation in upstream open reading frame-controlled translation.

Upstream open reading frames (uORFs) are a class of translated regions (translons) in mRNA 5' leaders. uORFs are believed to be pervasive regulators of the translation of mammalian mRNAs. Some uORFs are highly repressive but others have little or no impact on downstream mRNA translation either due to inefficient recognition of their start codon(s) or/and due to efficient reinitiation after uORF translation. While experiments with uORF reporter constructs proved to be instrumental in the investigation of uORF-mediated mechanisms of translation control, they can have serious limitations as manipulations with uORF sequences can yield various artefacts. Here we propose a general approach for using translation complex profiling (TCP-seq) data for exploring uORF regulatory characteristics. Using several examples, we show how TCP-seq could be used to estimate both repressiveness and modes of action of individual uORFs. We demonstrate how this approach could be used to assess the mechanisms of uORF-mediated translation control in the mRNA of several human genes, including EIF5, IFRD1, MDM2, MIEF1, PPP1R15B, TAF7, and UCP2.

Responses to Exercise with Low Carbohydrate Availability on Muscle Glycogen and Cell Signaling: A Systematic Review and Meta-analysis.

The growing interest in how exercise and carbohydrate (CHO) restriction may modify molecular responses that promote endurance adaptations has led to many interesting controversies. We conducted a systematic review and a meta-analysis regarding the effect of low-carbohydrate availability (LOW) pre-, during, or post-exercise, on the mRNA content of commonly measured genes involved in mitochondrial biogenesis (PGC-1α, TFAM mRNA) and metabolism (PDK4, UCP3 and GLUT4 mRNA), and on muscle glycogen levels, compared with a high-CHO (CON) condition. MEDLINE, Scopus, and Web of Science databases were searched following the PRISMA 2020 guidelines (with an end date of November 2023). In total, 19 randomized-controlled studies were considered for inclusion. We evaluated the methodological quality of all studies using the Cochrane Risk of Bias tool for randomized clinical studies. A meta-analysis was performed using a random effects model to calculate the standardized mean difference (SMD), estimated by Hedges' g, and 95% confidence intervals (CIs). The LOW condition was associated with an increased mRNA content of several genes during the early recovery period post-exercise, such as PDK4 (SMD 1.61; 95% CI 0.80-2.42), GLUT4 (SMD 1.38; 95% CI 0.46-2.30), and UCP3 (SMD 2.05; 95% CI 0.40-3.69). However, overall, there was no significant effect on the mRNA content of PGC-1α or TFAM. Finally, CHO restriction and exercise significantly reduced muscle glycogen levels (SMD 3.69; 95% CI 2.82-5.09). A meta-analysis of subgroups from studies with a difference in muscle glycogen concentration of > 200mmolkgdw-1 between the LOW and CON conditions showed an increase in exercise-induced PGC-1α mRNA (SMD 2.08; 95% CI 0.64-3.52; p = 0.005; I2 = 75%) and a greater effect in PDK4 and GLUT4 mRNA. The meta-analysis results show that CHO restriction was associated with an increase in the exercise-induced mRNA content of PDK4, UCP3, and GLUT4, but not the exercise-induced mRNA content of PGC-1ɑ and TFAM. However, when there were substantial differences in glycogen depletion between CON and LOW CHO conditions (> 200mmolkgdw-1), there was a greater effect of CHO restriction on the exercise-induced mRNA content of metabolic genes, and an increase in exercise-induced PGC-1α mRNA.

Myricetin and myricitrin indirectly and directly increases uncoupling protein-1 mRNA expression in C3H10T1/2 beige adipocytes

In thermogenic brown and beige adipocytes, the proton gradient formed by energy derived from nutrients such as lipids and carbohydrates is consumed by uncoupling protein-1 (UCP-1), resulting in thermogenesis without ATP production in the mitochondria. Accordingly, increased UCP-1 expression represents a crucial aspect of dietary management for individuals with overweight and obesity. Myricetin and its glycoside, myricitrin, are food-derived flavonoids that possess various beneficial effects. This is the first study to examine the effects of myricetin and myricitrin on the inflammation-inhibited expression of Ucp-1 using a modified cell-based assay with conditioned medium (CM). The CM derived from lipopolysaccharide (LPS)-activated RAW264.7 macrophages was observed to inhibit the Ucp-1 expression induced by adrenergic stimulation in 10T1/2 adipocytes. Conversely, the CM derived from activated macrophages treated with myricetin or myricitrin reversed this inhibition of Ucp-1 expression. Subsequently, the direct effects of both the compounds on basal and adrenaline-induced expression of Ucp-1 were investigated. In contrast to a previous report, myricetin and myricitrin did not increase the basal Ucp-1 mRNA expression in 10T1/2 adipocytes when treated during the differentiation-promoting period. However, we have found for the first time that both compounds enhanced the adrenergic sensitivity of 10T1/2 adipocytes when treated during the differentiation-inducing period. These results indicate that myricetin and myricitrin have indirect effects on inflammation-induced suppression and direct effects on adrenergic sensitivity, suggesting a novel mechanism that both compounds increase Ucp-1 expression in vivo by both indirect and direct effects, rather than by affecting basal expression.

Characterization of Ucp1-iCre knockin mice reveals the recombination activity in male germ cells.

Ucp1 promoter-driven Cre transgenic mice are useful in the manipulation of gene expression specifically in thermogenic adipose tissues. However, the wildly used Ucp1-Cre line was generated by random insertion into the genome and showed ectopic activity in some tissues beyond adipose tissues. Here, we characterized a knockin mouse line Ucp1-iCre generated by targeting IRES-Cre cassette immediately downstream the stop codon of the Ucp1 gene. The Cre insertion had little to no effect on uncoupling protein 1 (UCP1) levels in brown adipose tissue. Ucp1-iCre mice of both genders exhibited normal thermogenesis and cold tolerance. When crossed with Rosa-tdTomato reporter mice, Ucp1-iCre mice showed robust Cre activity in thermogenic adipose tissues. In addition, limited Cre activity was sparsely present in the ventromedial hypothalamus (VMH), choroid plexus, kidney, adrenal glands, ovary, and testis in Ucp1-iCre mice, albeit to a much lesser extent and with reduced intensity compared with the conventional Ucp1-Cre line. Single-cell transcriptome analysis revealed Ucp1 mRNA expression in male spermatocytes. Moreover, male Ucp1-iCre mice displayed a high frequency of Cre-mediated recombination in the germline, whereas no such effect was observed in female Ucp1-iCre mice. These findings suggest that Ucp1-iCre mice offer promising utility in the context of conditional gene manipulation in thermogenic adipose tissues, while also highlighting the need for caution in mouse mating and genotyping procedures.NEW & NOTEWORTHY Ucp1 promoter-driven Cre transgenic mice are useful in the manipulation of gene expression specifically in thermogenic adipose tissues. The widely used Ucp1-Cre mouse line (Ucp1-CreEvdr), which was generated using the bacterial artificial chromosome (BAC) strategy, exhibits major brown and white fat transcriptomic dysregulation and ectopic activity beyond adipose tissues. Here, we comprehensively validate Ucp1-iCre knockin mice, which serve as another optional model besides Ucp1-CreEvdr mice for specific genetic manipulation in thermogenic tissue.

Timing of acute cold exposure determines UCP1 and FGF21 expression - Possible interactions between the thermal environment, thermoregulatory responses, and peripheral clocks

Thermoregulation is synchronized across the circadian cycle to uphold thermal homeostasis. To test if time-of-day matters for the response to environmental cold exposure, mice were acclimated to thermoneutrality (27 °C) for 2 months were subjected acutely (8 h) to cold ambient conditions (15 °C), whereas controls were maintained at thermoneutral conditions. The thermal exposure was tested in separate groups (N = 8) at three distinct time-of-day periods: in the LIGHT phase (L); the DARK phase (D); and a mix of the two (D + L). The magnitude of UCP1 protein and mRNA induction in brown adipose tissue (BAT) in response to acute cold exposure was time-of-day sensitive, peaking in LIGHT, whereas lower induction levels were observed in D + L, and DARK. Plasma levels of FGF21 were induced 3-fold by acute cold exposure at LIGHT and D + L, compared to the time-matched thermoneutral controls, whereas cold in DARK did not cause a significant increase of FGF21 plasma levels. Cold exposure affected, in BAT, the temporal mRNA expression patterns of core circadian clock components: Bmal1, Clock, Per1, Per3, Cry1, Cry2 Nr1d1, and Nr1d2, but in the liver, none of the transcripts were modified. Behavioral assessment using the Thermal Gradient Test (TGT) showed that acute cold exposure reduced cold sensitivity in D + L, but not in DARK. RNA-seq analyses of somatosensory neurons in DRG highlighted the role of the core circadian components in these cells, as well as transcriptional changes due to acute cold exposure. This elucidates the sensory system as a gauge and potential regulator of thermoregulatory responses based on circadian physiology. In conclusion, acute cold exposure elicits time-of-day specific effects on thermoregulatory pathways, which may involve underlying changes in thermal perception. These results have implications for efforts aimed at reducing risks associated with the organization of shift work in cold environments.

Lycopene and Garcinia cambogia Induce White-to-Brown Adipose Differentiation: An Innovative Strategy to Curb Obesity.

Obesity is considered one of the main risk factors for cardiovascular diseases. The browning process has been recently recognized as a promising anti-obesity therapy. Lycopene (LYC) and Garcinia cambogia fruit extract (GE) might be important resources for anti-obesity drugs; therefore, the aim of this study was to investigate the anti-obesity effects of LYC and GE on 3T3-L1 adipocytes and Zucker rats. Mouse 3T3-L1 pre-adipocytes were differentiated in mature adipocytes and then treated with LYC (0.5 μM), GE (30 mg/mL) or LYC + GE for 24 h. Moreover, male Zucker Crl:ZUC-Leprfa rats were randomly assigned to 5 groups of 10 animals to orally receive Vehicle (Ctrl), Orlistat (20 mg/kg), LYC (5 mg/kg), GE (1000 mg/kg) or LYC + GE for 28 days. LYC, GC extracts and even more LYC + GE stimulated the mRNA and protein expression of thermogenic genes UCP1, CIDEA and DIO2, significantly reduced lipid droplet size and increased lipid droplet number in adipocytes. UCP1 mRNA and protein expression was also increased in the visceral adipose tissue of the rats that received the dietary intake of LYC, GE and even more LYC + GE. Moreover, LYC + GE induced the reorganization of visceral fat depots that showed a great number of small adipocytes and a significant reduction in weight gain and food intake compared to the control group. The obtained results demonstrated that LYC + GE might be used as new approaches for obesity management in order to induce the browning process and achieve a metabolically active tissue instead of a tissue characterized by lipid depot accumulation.

Activin E upregulates uncoupling protein 1 and fibroblast growth factor 21 in brown adipocytes

Activin E activates brown and beige adipocytes and has been controversially implicated as a factor that induces obesity and fatty liver. Here, we sought to address this controversial issue by producing recombinant human activin E to evaluate its effects on HB2 brown adipocytes in vitro. Activin E increased uncoupling protein 1 (Ucp1) and fibroblast growth factor 21 (Fgf21) mRNA expression in the adipocytes. This upregulation was suppressed by SB431542, an inhibitor of activin receptor-like kinase (Alk) TGF-β type I receptors. SB431542 also inhibited the activin E-induced phosphorylation of Smad2/3. A promoter assay using a CAGA-Luc reporter and Alk expression vectors revealed that activin E activated the TGF-β/activin pathway via Alk7. The upregulation of Ucp1 and Fgf21 mRNA might be mediated through Alk7 and Smad2/3 phosphorylation. Activin E is a potential stimulator of energy expenditure by activating brown adipocytes and highlights its potential as a therapeutic target for treating obesity.

Mouse vascularized adipose spheroids: an organotypic model for thermogenic adipocytes.

Adipose tissues, particularly beige and brown adipose tissue, play crucial roles in energy metabolism. Brown adipose tissues' thermogenic capacity and the appearance of beige cells within white adipose tissue have spurred interest in their metabolic impact and therapeutic potential. Brown and beige fat cells, activated by environmental factors like cold exposure or by pharmacology, share metabolic mechanisms that drive non-shivering thermogenesis. Understanding these two cell types requires advanced, yet broadly applicable in vitro models that reflect the complex microenvironment and vasculature of adipose tissues. Here we present mouse vascularized adipose spheroids of the stromal vascular microenvironment from inguinal white adipose tissue, a tissue with 'beiging' capacity in mice and humans. We show that adding a scaffold improves vascular sprouting, enhances spheroid growth, and upregulates adipogenic markers, thus reflecting increased adipocyte maturity. Transcriptional profiling via RNA sequencing revealed distinct metabolic pathways upregulated in our vascularized adipose spheroids, with increased expression of genes involved in glucose metabolism, lipid metabolism, and thermogenesis. Functional assessment demonstrated increased oxygen consumption in vascularized adipose spheroids compared to classical 2D cultures, which was enhanced by β-adrenergic receptor stimulation correlating with elevated β-adrenergic receptor expression. Moreover, stimulation with the naturally occurring adipokine, FGF21, induced Ucp1 mRNA expression in the vascularized adipose spheroids. In conclusion, vascularized inguinal white adipose tissue spheroids provide a physiologically relevant platform to study how the stromal vascular microenvironment shapes adipocyte responses and influence activated thermogenesis in beige adipocytes.

Amyloid-β neuropathology induces bone loss in male mice by suppressing bone formation and enhancing bone resorption

Alzheimer's disease (AD) and osteoporosis often coexist in the elderly. Although observational studies suggest an association between these two diseases, the pathophysiologic link between AD and skeletal health has been poorly defined. We examined the skeletal phenotype of 5xFAD mice, an AD model with accelerated neuron-specific amyloid-β accumulation causing full-blown AD phenotype by the age of 8 months. Micro-computed tomography indicated significantly lower trabecular and cortical bone parameters in 8-month-old male, but not female, 5xFAD mice than sex-matched wild-type littermates. Dynamic histomorphometry revealed reduced bone formation and increased bone resorption, and quantitative RT-PCR showed elevated skeletal RANKL gene expression in 5xFAD males. These mice also had diminished body fat percentage with unaltered lean mass, as determined by dual-energy X-ray absorptiometry (DXA), and elevated Ucp1 mRNA levels in brown adipose tissue, consistent with increased sympathetic tone, which may contribute to the osteopenia observed in 5xFAD males. Nevertheless, no significant changes could be detected between male 5xFAD and wild-type littermates regarding the serum and skeletal concentrations of norepinephrine. Thus, brain-specific amyloid-β pathology is associated with osteopenia and appears to affect both bone formation and bone resorption. Our findings shed new light on the pathophysiologic link between Alzheimer's disease and osteoporosis.

Dexamethasone Inhibits White Adipose Tissue Browning.

White adipose tissue (WAT) regulates energy balance through energy storage, adipokines secretion and the thermogenesis process. Beige adipocytes are responsible for WAT thermogenesis. They are generated by adipogenesis or transdifferentiation during cold or β3-adrenergic agonist stimulus through a process called browning. Browning has gained significant interest for to its preventive effect on obesity. Glucocorticoids (GCs) have several functions in WAT biology; however, their role in beige adipocyte generation and WAT browning is not fully understood. The aim of our study was to determine the effect of dexamethasone (DXM) on WAT thermogenesis. For this purpose, rats were treated with DXM at room temperature (RT) or cold conditions to determine different thermogenic markers. Furthermore, the effects of DXM on the adipogenic potential of beige precursors and on mature beige adipocytes were evaluated in vitro. Our results showed that DXM decreased UCP-1 mRNA and protein levels, mainly after cold exposure. In vitro studies showed that DXM decreased the expression of a beige precursor marker (Ebf2), affecting their ability to differentiate into beige adipocytes, and inhibited the thermogenic response of mature beige adipocytes (Ucp-1, Dio2 and Pgc1α gene expressions and mitochondrial respiration). Overall, our data strongly suggest that DXM can inhibit the thermogenic program of both retroperitoneal and inguinal WAT depots, an effect that could be exerted, at least partially, by inhibiting de novo cell generation and the thermogenic response in beige adipocytes.

Fat accumulation in striped hamsters (Cricetulus barabensis) reflects the temperature of prior cold acclimation

BackgroundProper adjustments of metabolic thermogenesis play an important role in thermoregulation in endotherm to cope with cold and/or warm ambient temperatures, however its roles in energy balance and fat accumulation remain uncertain. Our study aimed to investigate the effect of previous cold exposure (10 and 0 °C) on the energy budgets and fat accumulation in the striped hamsters (Cricetulus barabensis) in response to warm acclimation. The body mass, energy intake, resting metabolic rate (RMR) and nonshivering thermogenesis (NST), serum thyroid hormone levels (THs: T3 and T4), and the activity of brown adipose tissue (BAT), indicated by cytochrome c oxidase (COX) activity and uncoupling protein 1 (ucp1) expression, were measured following exposure to the cold (10 °C and 0 °C) and transition to the warm temperature (30 °C).ResultsThe hamsters at 10 °C and 0 °C showed significant increases in energy intake, RMR and NST, and a considerable reduction in body fat than their counterparts kept at 21 °C. After being transferred from cold to warm temperature, the hamsters consumed less food, and decreased RMR and NST, but they significantly increased body fat content. Interestingly, the hamsters that were previously exposed to the colder temperature showed significantly more fat accumulation after transition to the warm. Serum T3 levels, BAT COX activity and ucp1 mRNA expression were significantly increased following cold exposure, and were considerably decreased after transition to the warm. Furthermore, body fat content was negatively correlated with serum T3 levels, BAT COX activity and UCP1 expression.ConclusionThe data suggest that the positive energy balance resulting from the decreased RMR and NST in BAT under the transition from the cold to the warm plays important roles in inducing fat accumulation. The extent of fat accumulation in the warm appears to reflect the temperature of the previous cold acclimation.

NG ,NG -Dimethylarginine Dimethylaminohydrolase 1 Expression Is Dispensable for Cold- or Diet-Induced Thermogenesis.

The strategy to activate thermogenic adipocytes has therapeutic potential to overcome obesity as they dissipate surplus energy as heat through various mechanisms. NG,NG-dimethylarginine dimethylaminohydrolases (DDAHs) are enzymes involved in the nitric oxide-protein kinase G signaling axis which increases thermogenic gene expression. However, the role of DDAHs in thermogenic adipocytes has not been elucidated. The adipocyte-specific Ddah1 knockout mice are generated by crossing Ddah1fl/fl mice with adiponectin Cre recombinase mice. Adipocyte-specific DDAH1 overexpressing mice are generated using adeno-associated virus-double-floxed inverse open reading frame (AAV-DIO) system. These mice are analyzed under basal, cold exposure, or high-fat diet (HFD) conditions. Primary inguinal white adipose tissue cells from adipocyte-specific Ddah1 knockout mice expressed comparable amounts of Ucp1 mRNA. Adipocyte-specific DDAH1 overexpressing mice do not exhibit enhanced activation of thermogenic adipocytes. In addition, when these mice are exposed to cold environment or fed an HFD, their body temperature/weight and thermogenesis-related gene and protein expressions are unchanged. These findings indicate that DDAH1 does not play a role in either cold- or diet-induced thermogenesis. Therefore, adipocyte targeting DDAH1 gene therapy for the treatment of obesity is unlikely to be effective.

(-)-epicatechin treatment did not modify the thermogenic pathway in the gastrocnemius muscle of male rat offspring obeses by programming.

The aim of this study was to analyse the expression of genes related to the regulation of energy metabolism in skeletal muscle tissue by comparing male offspring in two age groups [at 110 and 245 postnatal days (pnd)] from a mother with obesity induced by a high-fat diet and (-)-epicatechin (Epi) administration. Four groups of six male offspring from different litters were randomly selected for the control groups [C and offspring of mothers with maternal obesity (MO)] or Epi intervention groups. We evaluated the effect of Epi on gastrocnemius tissue by analysing the mRNA and protein expression levels of Fndc5/irisin, Pgc-1α, Ucp3, and Sln. Epi significantly increased the Pgc-1α protein in the MO group of offspring at 110 pnd (p < 0.036, MO vs. MO+Epi), while at 245 pnd, Epi increased Fndc5/irisin mRNA expression in the MO+Epi group versus the MO group (p = 0.006).No differences were detected in Fndc5/irisin, Ucp3 or Sln mRNA or protein levels (including Pgc-1α mRNA) in the offspring at 110 pnd or in Pgc-1α, Ucp3, or Sln mRNA or protein levels (including Fndc5/irisin protein) at 245 pnd among the experimental groups. In conclusion, (-)-epicatechin treatment increased Fndc5/irisin mRNA expression and Pgc-α protein levels in the gastrocnemius muscle of offspring at postnatal days 110 and 245. Furthermore, it is suggested that the flavonoid effect in a model of obesity and its impact on thermogenesis in skeletal muscle are regulated by a different pathway than Fndc5/irisin.

Nuciferine prevents obesity by activating brown adipose tissue.

Increasing evidence suggests that brown adipose tissue (BAT) plays an important role in obesity and related diseases. Increasing the amount or activity of BAT could prevent obesity. Therefore, a safe and effective method of activating BAT is urgently required. Here, we evaluated the potential effects of lotus leaf extract (LLE) on BAT function. We found that LLE substantially increased UCP1 mRNA and protein levels as well as thermogenic protein expression in primary brown adipocytes. Additionally, LLE treatment reduced diet-induced obesity and improved glucose homeostasis owing to BAT activation and increased energy expenditure. We found that nuciferine, an active ingredient of LLE, could dose-dependently activate BAT in vitro and in vivo, alleviate diet-induced obesity, and improve glucose homeostasis by increasing energy expenditure. Mechanistically, we found that nuciferine induced PPARG coactivator 1 alpha (PGC1-α) expression, which is a key gene involved in mitochondrial biogenesis promoter activity, by directly binding to RXRA. Furthermore, RXRA knockdown abolished expression of the nuciferine-induced mitochondrial and thermogenesis-related gene in primary brown adipocytes. In summary, we found that LLE and nuciferine have a notable effect on BAT activation and highlight the potential applications of the main component of LLE in preventing obesity and treating metabolic disorders.

The effect of endurance training combined with adenosine on the gene expression of UCP-1 and MAPK p38 in subcutaneous adipose tissue of male Wistar rats fed a high -fat diet

Background: Strategies to increase energy expenditure are an attractive approach to reduce excess fat storage and body weight to improve metabolic health. The aim of the present study was to investigate the effects of endurance training combined with adenosine injection on the gene expression of UCP-1 and MAPK p38 in the subcutaneous adipose tissue of male rats fed a high-fat diet. Materials and Methods: Forty rats were randomly divided into 4 groups: 1. normal control, 2. high-fat diet (HFD) control, 3. HFD + adenosine, and 4. HFD + endurance training + adenosine. After 13 weeks of HFD, 12 weeks of endurance training on a moderate-intensity treadmill was performed. UCP-1 and MAPK p38 mRNA levels were measured by RT-PCR. Results: A significant increase in UCP-1 was observed with in HFD + endurance training + adenosine and HFD + adenosine compared to normal and HFD controls. A significant decrease in MAPK p38 was also observed with HFD + endurance training + adenosine and HFD + adenosine compared to HFD. Conclusion: Endurance training and adenosine are likely activators of UCP-1 gene expression and can be used as effective lipolytic agents in obesity. The MAPK p38 pathway increases glucose uptake by insulin and also induces oxidative phosphorylation in mitochondria following a healthy diet and aerobic activity.

FRI025 Succinate Receptor Activation Increases Respiration In Human But Not Murine Brown Adipocytes

Abstract Disclosure: B.T. McNeill: None. A. Kelman: None. L.E. Ramage: None. K.J. Suchacki: None. S.J. Wakelin: None. N.M. Morton: None. R.H. Stimson: None. Brown adipose tissue (BAT) is a thermogenic tissue that generates heat to maintain body temperature. BAT activation is a potential therapeutic target for treating obesity-related cardiometabolic disease. We performed RNA sequencing of human primary brown and white adipocytes and identified SUCNR1, which encodes the succinate receptor, as highly differentially expressed (20-fold) in brown versus white adipocytes. Succinate is known to increase BAT thermogenesis, while SUCNR1 activation inhibits lipolysis in white adipose tissue (WAT). We hypothesized that SUCNR1 activation increases BAT thermogenesis in humans and in mice. To test this, we investigated the effect of succinate and the selective SUCNR1 agonist cis-epoxysuccinic acid (C-ESA) in human and murine primary brown/ beige adipocytes. In addition, we determined the metabolic effects of disruption of Sucnr1 in mice either fed control or a high fat (HFD) for 12 weeks and measured energy expenditure when housed at standard room temperature (21°C) and during cold exposure (4°C). In human brown and white adipocytes, succinate increased basal and noradrenaline-stimulated respiration, while C-ESA increased basal respiration only in human brown adipocytes without altering UCP1 mRNA levels. SUCNR1 activation reduced glycerol release only in human white adipocytes. Male (but not female) Sucnr1-/- mice exhibited impaired glucose tolerance versus Sucnr1+/+ littermates after 2 weeks of HFD, and greater fat mass was also observed in the Sucnr1-/- mice following chronic HFD. However, energy expenditure, locomotor activity and tail vein temperature were unchanged in male Sucnr1-/- mice either at 21°C or 4°C. In addition, expression of Ucp1 and Pgc1α in BAT and inguinal WAT were similar between genotypes. Sucnr1 deletion had no effect on fat mass or glucose tolerance in mice fed a control diet. In vitro, differentiated brown adipocytes from Sucnr1-/- mice demonstrated comparable cellular respiration and Ucp1 mRNA levels both basally and during noradrenaline stimulation to Sucnr1+/+ adipocytes. Succinate and C-ESA did not stimulate respiration in murine brown or white adipocytes. These data reveal that SUCNR1 activation enhances brown adipocyte respiration in human but not murine brown adipocytes, highlighting a novel pathway regulating BAT function in addition to differences between species. These data also reveal novel sex-specific differences in the metabolic effects of SUCNR1, although the mechanisms through which female mice are protected from the adverse metabolic effects of Sucnr1 deletion are unclear. SUCNR1 activation may be a novel therapeutic target to treat obesity and associated metabolic disease, but requires further research in humans to determine the specific mechanisms through which SUCNR1 regulates BAT thermogenesis. Presentation: Friday, June 16, 2023

HDAC11 inhibition triggers bimodal thermogenic pathways to circumvent adipocyte catecholamine resistance.

Stimulation of adipocyte β-adrenergic receptors (β-ARs) induces expression of uncoupling protein 1 (UCP1), promoting nonshivering thermogenesis. Association of β-ARs with a lysine-myristoylated form of A kinase-anchoring protein 12 (AKAP12, also known as gravin-α) is required for downstream signaling that culminates in UCP1 induction. Conversely, demyristoylation of gravin-α by histone deacetylase 11 (HDAC11) suppresses this pathway. Whether inhibition of HDAC11 in adipocytes is sufficient to drive UCP1 expression independently of β-ARs is not known. Here, we demonstrate that adipocyte-specific deletion of HDAC11 in mice leads to robust induction of UCP1 in adipose tissue (AT), resulting in increased body temperature. These effects are mimicked by treating mice in vivo or human AT ex vivo with an HDAC11-selective inhibitor, FT895. FT895 triggers biphasic, gravin-α myristoylation-dependent induction of UCP1 protein expression, with a noncanonical acute response that is posttranscriptional and independent of protein kinase A (PKA), and a delayed response requiring PKA activity and new Ucp1 mRNA synthesis. Remarkably, HDAC11 inhibition promotes UCP1 expression even in models of adipocyte catecholamine resistance where β-AR signaling is blocked. These findings define cell-autonomous, multimodal roles for HDAC11 as a suppressor of thermogenesis, and highlight the potential of inhibiting HDAC11 to therapeutically alter AT phenotype independently of β-AR stimulation.

Metabolite Profile Characterization of Cyanobacterial Strains with Bioactivity on Lipid Metabolism Using In Vivo and In Vitro Approaches.

Cyanobacteria have demonstrated their therapeutic potential for many human diseases. In this work, cyanobacterial extracts were screened for lipid reducing activity in zebrafish larvae and in fatty-acid-overloaded human hepatocytes, as well as for glucose uptake in human hepatocytes and ucp1 mRNA induction in murine brown adipocytes. A total of 39 cyanobacteria strains were grown and their biomass fractionated, resulting in 117 chemical fractions. Reduction of neutral lipids in zebrafish larvae was observed for 12 fractions and in the human hepatocyte steatosis cell model for five fractions. The induction of ucp1 expression in murine brown adipocytes was observed in six fractions, resulting in a total of 23 bioactive non-toxic fractions. All extracts were analyzed by untargeted UPLC-Q-TOF-MS mass spectrometry followed by multivariate statistical analysis to prioritize bioactive strains. The metabolite profiling led to the identification of two markers with lipid reducing activity in zebrafish larvae. Putative compound identification using mass spectrometry databases identified them as phosphatidic acid and aromatic polyketides derivatives-two compound classes, which were previously associated with effects on metabolic disorders. In summary, we have identified cyanobacterial strains with promising lipid reducing activity, whose bioactive compounds needs to be identified in the future.

Electroacupuncture improves obesity and promotes white adipose tissue browning by regulating central glucagon-like peptide-1

To observe the effect of electroacupuncture （EA） on white adipose tissue （WAT） browning by regulating central glucagon-like peptide-1 （GLP-1）, so as to explore the possible central mechanisms of EA in improving obesity. Thirty male Wistar rats were randomly divided into normal group, model group, EA group, HM3D group, and EA+HM4D group, with 6 rats in each group. The obesity rat model was obtained by feeding with high-fat diet for 8 weeks. Adeno-associated virus combined with DREADDs was injected into bilateral nucleus of solitary tract （NTS）, with rAAV-GLP-1+rAAV-4D applied to the EA+HM4D group, rAAV-GLP-1+rAAV-3D applied to the HM3D group, and rAAV-GLP-1+rAAV-GFP applied to other 3 groups. After modeling, rats in the EA and EA+HM4D groups received EA treatment at bilateral "Zusanli"（ST36）, "Fenglong"（ST40）, "Guanyuan"（CV4） and "Zhongwan"（CV12）, with successive waves （2 Hz, 1 mA） for 10 minutes, 3 times a week, for a total of 8 weeks. Body mass of rats in each group were measured before and 2, 4, 6, and 8 weeks after intervention. Abdominal and perirenal WAT mass was weighed, serum triglyceride （TG） and total cholesterol （TC） contents were detected by using automatic analyzer, and nonestesterified fatty acid （NEFA） content was detected by using colorimetric assay kit. The morphology of abdominal WAT lipid droplets was observed by HE staining. The mRNA expressions of GLP-1 in NTS, AMPK in ventromedial nucleus of hypothalamus（VMH）, UCP1 and PGC-1α in subcutaneous fat were detected by real-time PCR. The protein expression levels of GLP-1, AMPK, phosphorylated-AMPK, UCP1 and PGC-1α were detected by Western blot. The activation level of GLP-1 neurons in NTS was observed by immunofluorescence. Compared with the normal group, abdominal WAT lipid droplets were enlarged, body weight, serum TG, TC, NEFA contents, abdominal and perirenal WAT mass, mRNA and protein expression levels of AMPK were significantly increased（P<0.01, P<0.05）, while GLP-1 neurons activation level, mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α, and AMPK protein phosphorylation were decreased （P<0.01） in the model group. After EA intervention, body weight at 6 and 8 weeks after intervention and other indexes mentioned above were all significantly reversed （P<0.01, P<0.05） in the EA group in comparison with those of the model group. Compared with the EA group, the HM3D group had reduced abdominal WAT lipid droplets size, decreased serum TG, TC, and NEFA contents, and protein expression level of AMPK（P<0.01, P<0.05）, with increased mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α, and phosphorylation level of AMPK protein（P<0.01, P<0.05）, while the EA+HM4D group had enlarged abdominal WAT lipid droplets, increased body weight 6 and 8 weeks after intervention, abdominal and renal WAT mass, and NEFA content （P<0.01, P<0.05）, with decreased serum TG content, activation level of GLP-1 neurons in the NTS, mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α （P<0.01, P<0.05）, as well as down-regulated phosphorylation of AMPK protein and mRNA （P<0.01, P<0.05）. EA can effectively promote the browning of WAT, which may be related to the activation of GLP-1 neurons in the NTS, as well as the promotion of the phosphorylation of AMPK in the VMH and up-regulation of UCP1.