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Brown adipose tissue (BAT) thermogenesis results from the uncoupling of mitochondrial inner membrane proton gradient mediated by uncoupling protein 1 (UCP-1), which is activated by lipolysis-derived fatty acids. Norepinephrine (NE) secreted by sympathetic innervation not only activates BAT lipolysis and UCP-1 but, uniquely in brown adipocytes, promotes "futile" metabolic cycles and enhances BAT thermogenic capacity by increasing UCP-1 content, mitochondrial biogenesis, and brown adipocyte hyperplasia. NE exerts these actions by triggering signaling in the canonical G protein-coupled β-adrenergic receptors, cAMP, and protein kinase A (PKA) pathway, which in brown adipocytes is under a complex and intricate cross talk with important growth-promoting signaling pathways such as those of mechanistic target of rapamycin (mTOR) complexes 1 (mTORC1) and 2 (mTORC2). This article reviews evidence suggesting that mTOR complexes are modulated by and participate in the thermogenic, metabolic, and growth-promoting effects elicited by NE in BAT and discusses current gaps and future directions in this field of research.

Chloroquine diphosphate (CQ), a weak base used to inhibit autophagic flux and treat malaria and rheumatoid diseases, has been shown, through unknown mechanisms, to improve glucose and lipid homeostasis in patients and rodents. We investigate herein the molecular mechanisms underlying these CQ beneficial metabolic actions in diet-induced obese mice. For this, C57BL6/J mice fed with either a chow or a high-fat diet (HFD) and uncoupling protein 1 (UCP-1) KO and adipocyte Atg7-deficient mice fed with a HFD were treated or not with CQ (60 mg/kg of body weight/day) during 8 weeks and evaluated for body weight, adiposity, glucose homeostasis and brown and white adipose tissues (BAT and WAT) UCP-1 content. CQ reduced body weight gain and adipose tissue and liver masses in mice fed with a HFD, without altering food intake, oxygen consumption, respiratory exchange ratio, spontaneous motor activity and feces caloric content. CQ attenuated the insulin intolerance, hyperglycemia, hyperinsulinemia, hypertriglyceridemia and hypercholesterolemia induced by HFD intake, such effects that were associated with increases in serum and liver fibroblast growth factor 21 (FGF-21) and BAT and WAT UCP-1 content. Interestingly, CQ beneficial metabolic actions of reducing body weight and adiposity and improving glucose homeostasis were preserved in HFD-fed UCP-1 KO and adipocyte Atg7 deficient mice. CQ reduces body weight gain and adiposity and improves glucose homeostasis in diet-induced obese mice through mechanisms that might involve FGF-21, but not UCP1-mediated nonshivering thermogenesis or inhibition of adipocyte autophagy., Competing Interests: Declaration of competing interest The authors declare no competing and conflicting of interests., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Hepatocellular carcinoma (HCC) markedly enhances liver secretion of fibroblast growth factor 21 (FGF-21), a hepatokine that increases brown and subcutaneous inguinal white adipose tissues (BAT and iWAT, respectively) uncoupling protein 1 (UCP-1) content, thermogenesis and energy expenditure. Herein, we tested the hypothesis that an enhanced BAT and iWAT UCP-1-mediated thermogenesis induced by high levels of FGF-21 is involved in HCC-associated catabolic state and fat mass reduction. For this, we evaluated body weight and composition, liver mass and morphology, serum and tissue levels of FGF-21, BAT and iWAT UCP-1 content, and thermogenic capacity in mice with Pten deletion in hepatocytes that display a well-defined progression from steatosis to steatohepatitis (NASH) and HCC upon aging. Hepatocyte Pten deficiency promoted a progressive increase in liver lipid deposition, mass, and inflammation, culminating with NASH at 24 weeks and hepatomegaly and HCC at 48 weeks of age. NASH and HCC were associated with elevated liver and serum FGF-21 content and iWAT UCP-1 expression (browning), but reduced serum insulin, leptin, and adiponectin levels and BAT UCP-1 content and expression of sympathetically regulated gene glycerol kinase (GyK), lipoprotein lipase (LPL), and fatty acid transporter protein 1 (FATP-1), which altogether resulted in an impaired whole-body thermogenic capacity in response to CL-316,243. In conclusion, FGF-21 pro-thermogenic actions in BAT are context-dependent, not occurring in NASH and HCC, and UCP-1-mediated thermogenesis is not a major energy-expending process involved in the catabolic state associated with HCC induced by Pten deletion in hepatocytes., (© 2023. The Author(s) under exclusive licence to University of Navarra.)

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive loss of motor neurons, systemic hypermetabolism, and inflammation. In this context, oxylipins have been investigated as signaling molecules linked to neurodegeneration, although their specific role in ALS remains unclear. Importantly, most methods focused on oxylipin analysis are based on low-resolution mass spectrometry, which usually confers high sensitivity, but not great accuracy for molecular characterization, as provided by high-resolution MS (HRMS). Here, we established an ultra-high performance liquid chromatography HRMS (LC-HRMS) method for simultaneous analysis of 126 oxylipins in plasma. Intra- and inter-day method validation showed high sensitivity (0.3-25 pg), accuracy and precision for more than 90% of quality controls. This method was applied in plasma of ALS rats overexpressing the mutant human Cu/Zn-superoxide dismutase gene (SOD1-G93A) at asymptomatic (ALS 70 days old) and symptomatic stages (ALS 120 days old), and their respective age-matched wild type controls. From the 56 oxylipins identified in plasma, 17 species were significantly altered. Remarkably, most of oxylipins linked to inflammation and oxidative stress derived from arachidonic acid (AA), like prostaglandins and mono-hydroxides, were increased in ALS 120 d rats. In addition, ketones derived from AA and linoleic acid (LA) were increased in both WT 120 d and ALS 120 d groups, supporting that age also modulates oxylipin metabolism in plasma. Interestingly, the LA-derived diols involved in fatty acid uptake and β-oxidation, 9(10)-DiHOME and 12(13)-DiHOME, were decreased in ALS 120 d rats and showed significant synergic effects between age and disease factors. In summary, we validated a high-throughput LC-HRMS method for oxylipin analysis and provided a comprehensive overview of plasma oxylipins involved in ALS disease progression. Noteworthy, the oxylipins altered in plasma have potential to be investigated as biomarkers for inflammation and hypermetabolism in ALS., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

Uncoupling protein 1 (UCP1) plays a central role in thermogenic tissues by uncoupling cellular respiration to dissipate energy. Beige adipocytes, an inducible form of thermogenic cells in subcutaneous adipose tissue (SAT), have become a major focus in obesity research. We have previously shown that eicosapentaenoic acid (EPA) ameliorated high-fat diet (HFD)-induced obesity by activating brown fat in C57BL/6J (B6) mice at thermoneutrality (30 °C), independently of UCP1. Here, we investigated whether ambient temperature (22 °C) impacts EPA effects on SAT browning in wild-type (WT) and UCP1 knockout (KO) male mice and dissected underlying mechanisms using a cell model. We observed resistance to diet-induced obesity in UCP1 KO mice fed HFD at ambient temperature, with significantly higher expression of UCP1-independent thermogenic markers, compared to WT mice. These markers included the fibroblast growth factor 21 (FGF21) and sarco/endoplasmic reticulum Ca 2+ -ATPase 2b (SERCA2b), suggesting the indispensable role of temperature in beige fat reprogramming. Surprisingly, although EPA induced thermogenic effects in SAT-derived adipocytes harvested from both KO and WT mice, EPA only increased thermogenic gene and protein expression in the SAT of UCP1 KO mice housed at ambient temperature. Collectively, our findings indicate that the thermogenic effects of EPA, which are independent of UCP1, occur in a temperature-dependent manner.

Cold acclimation and pharmacological peroxisome proliferator-activated receptor γ (PPARγ) activation have each earlier been shown to recruit brown adipose tissue (BAT) and beige adipocytes thermogenic machinery, enhancing uncoupling protein 1 (UCP1)-mediated thermogenic capacity. We here investigated whether cold acclimation and PPARγ agonism combined have additive effects in inducing brown and beige adipocytes UCP1 content and whether this translates into a higher thermogenic capacity and energy expenditure. C57BL/6J mice treated or not with pioglitazone (30 mg/kg/day) were maintained at 21°C or exposed to cold (7°C) for 15 days and evaluated for thermogenic capacity, energy expenditure and interscapular BAT (iBAT) and inguinal white adipose tissue (iWAT) mass, morphology, UCP1 content and gene expression, glucose uptake and oxygen consumption. Cold acclimation and PPARγ agonism combined synergistically increased iBAT and iWAT total UCP1 content and mRNA levels of the thermogenesis-related proteins PGC1a, CIDEA, FABP4, GYK, PPARa, LPL, GLUTs (GLUT1 in iBAT and GLUT4 in iWAT), and ATG when compared to cold and pioglitazone individually. This translated into a stronger increase in body temperature in response to the β 3 -adrenergic agonist CL316,243 and iBAT and iWAT respiration induced by succinate and pyruvate in comparison to that seen in either cold-acclimated or pioglitazone-treated mice. However, basal energy expenditure, BAT glucose uptake and glucose tolerance were not increased above that seen in cold-acclimated untreated mice. In conclusion, cold acclimation and PPARγ agonism combined induced a robust increase in brown and beige adipocytes UCP1 content and thermogenic capacity, much higher than each treatment individually. However, our findings enforce the concept that increases in total UCP1 do not innately lead to higher energy expenditure. NEW & NOTEWORTHY Cold acclimation and PPARγ agonism combined markedly increase brown and white adipose tissue total UCP1 content and mRNA levels of thermogenesis-related proteins. Higher UCP1 protein levels did not result in higher energy expenditure. The high thermogenic capacity induced by PPARγ agonism in cold-exposed animals markedly increases animals' body temperature in response to the β3-adrenergic agonist CL316,243.

Total absence of adipose tissue (lipoatrophy) is associated with the development of severe metabolic disorders including hepatomegaly and fatty liver. Here, we sought to investigate the impact of severe lipoatrophy induced by deletion of peroxisome proliferator-activated receptor gamma (PPARγ) exclusively in adipocytes on lipid metabolism in mice. Untargeted lipidomics of plasma, gastrocnemius and liver uncovered a systemic depletion of the essential linoleic (LA) and α-linolenic (ALA) fatty acids from several lipid classes (storage lipids, glycerophospholipids, free fatty acids) in lipoatrophic mice. Our data revealed that such essential fatty acid depletion was linked to increased: 1) capacity for liver mitochondrial fatty acid β-oxidation (FAO), 2) citrate synthase activity and coenzyme Q content in the liver, 3) whole-body oxygen consumption and reduced respiratory exchange rate in the dark period, and 4) de novo lipogenesis and carbon flux in the TCA cycle. The key role of de novo lipogenesis in hepatic steatosis was evidenced by an accumulation of stearic, oleic, sapienic and mead acids in liver. Our results thus indicate that the simultaneous activation of the antagonic processes FAO and de novo lipogenesis in liver may create a futile metabolic cycle leading to a preferential depletion of LA and ALA. Noteworthy, this previously unrecognized cycle may also explain the increased energy expenditure displayed by lipoatrophic mice, adding a new piece to the metabolic regulation puzzle in lipoatrophies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Aims: Blood vessels are surrounded by perivascular adipose tissue (PVAT), which plays an important role in vascular tonus regulation due to its anticontractile effect; however, this effect is impaired in obesity. We previously demonstrated that miRNA-22 is involved in obesity-related metabolic disorders. However, the impact of miRNA-22 on vascular reactivity and PVAT function is unknown., Aim: To investigate the role of miRNA-22 on vascular reactivity and its impact on obesity-induced PVAT dysfunction., Main Methods: Wild-type and miRNA-22 knockout (KO) mice were fed a control or a high-fat (HF) diet. To characterize the vascular response, concentration-responses curves to noradrenaline were performed in PVAT- or PVAT+ thoracic aortic rings in absence and presence of L-NAME. Expression of adipogenic and thermogenic markers and NOS isoforms were evaluated by western blotting or qPCR., Key Findings: HF diet and miRNA-22 deletion reduced noradrenaline-induced contraction in PVAT- aortic rings. Additionally, miRNA-22 deletion increased noradrenaline-induced contraction in PVAT+ aortic rings without affecting its sensitivity; however, this effect was not observed in miRNA-22 KO mice fed a HF diet. Interestingly, miRNA-22 deletion reduced the contraction of aortic rings to noradrenaline via a NOS-dependent mechanism. Moreover, HF diet abolished the NOS-mediated anticontractile effect of PVAT, which was attenuated by miRNA-22 deletion. Mechanistically, we found that PVAT from miRNA-22 KO mice fed a HF diet presented increased protein expression of nNOS., Significance: These results suggest that miRNA-22 is important for aorta reactivity under physiological circumstances and its deletion attenuates the loss of the NOS-mediated anticontractile effect of PVAT in obesity., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Mutations in the mitochondrial genome (mtDNA) are ubiquitous in humans and can lead to a broad spectrum of disorders. However, due to the presence of multiple mtDNA molecules in the cell, co-existence of mutant and wild-type mtDNAs (termed heteroplasmy) can mask disease phenotype unless a threshold of mutant molecules is reached. Importantly, the mutant mtDNA level can change across lifespan as mtDNA segregates in an allele- and cell-specific fashion, potentially leading to disease. Segregation of mtDNA is mainly evident in hepatic cells, resulting in an age-dependent increase of mtDNA variants, including non-synonymous potentially deleterious mutations. Here we modeled mtDNA segregation using a well-established heteroplasmic mouse line with mtDNA of NZB/BINJ and C57BL/6N origin on a C57BL/6N nuclear background. This mouse line showed a pronounced age-dependent NZB mtDNA accumulation in the liver, thus leading to enhanced respiration capacity per mtDNA molecule. Remarkably, liver-specific atg7 (autophagy related 7) knockout abolished NZB mtDNA accumulat ion, resulting in close-to-neutral mtDNA segregation through development into adulthood. prkn (parkin RBR E3 ubiquitin protein ligase) knockout also partially prevented NZB mtDNA accumulation in the liver, but to a lesser extent. Hence, we propose that age-related liver mtDNA segregation is a consequence of macroautophagic clearance of the less-fit mtDNA. Considering that NZB/BINJ and C57BL/6N mtDNAs have a level of divergence comparable to that between human Eurasian and African mtDNAs, these findings have potential implications for humans, including the safe use of mitochondrial replacement therapy. Abbreviations: Apob : apolipoprotein B; Atg1 : autophagy-related 1; Atg7 : autophagy related 7; Atp5a1 : ATP synthase, H+ transporting, mitochondrial F1 complex, alpha subunit 1; BL6: C57BL/6N mouse strain; BNIP3 : BCL2/adenovirus E1B interacting protein 3; FCCP: carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; MAP1LC3A: microtubule-associated protein 1 light chain 3 alpha; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; mt-Atp8 : mitochondrially encoded ATP synthase 8; MT-CO1: mitochondrially encoded cytochrome c oxidase I; MT-CO2: mitochondrially encoded cytochrome c oxidase II; mt-Co3 : mitochondrially encoded cytochrome c oxidase III; mt-Cytb : mitochondrially encoded cytochrome b; mtDNA: mitochondrial DNA; MUL1: mitochondrial ubiquitin ligase activator of NFKB 1; nDNA: nuclear DNA; Ndufa9 : NADH:ubiquinone oxireductase subunit A9; NDUFB8: NADH:ubiquinone oxireductase subunit B8; Nnt : nicotinamide nucleotide transhydrogenase; NZB: NZB/BINJ mouse strain; OXPHOS: oxidative phosphorylation; PINK1: PTEN induced putative kinase 1; Polg2 : polymerase (DNA directed), gamma 2, accessory subunit; Ppara : peroxisome proliferator activated receptor alpha; Ppia : peptidylprolyl isomerase A; Prkn : parkin RBR E3 ubiquitin protein ligase; P10: post-natal day 10; P21: post-natal day 21; P100: post-natal day 100; qPCR: quantitative polymerase chain reaction; Rpl19 : ribosomal protein L19; Rps18 : ribosomal protein S18; SD: standard deviation; SEM: standard error of the mean; SDHB: succinate dehydrogenase complex, subunit B, iron sulfur (Ip); SQSTM1: sequestosome 1; Ssbp1 : single-stranded DNA binding protein 1; TFAM: transcription factor A, mitochondrial; Tfb1m : transcription factor B2, mitochondrial; TOMM20: translocase of outer mitochondrial membrane 20; UQCRC2: ubiquinol cytochrome c reductase core protein 2; WT: wild-type.Tfb2m : transcription factor B2, mitochondrial; TOMM20: translocase of outer mitochondrial membrane 20; UQCRC2: ubiquinol cytochrome c reductase core protein 2; WT: wild-type.

Intracellular peptides (InPeps) generated by proteasomes were previously suggested as putative natural regulators of protein-protein interactions (PPI). Here, the main aim was to investigate the intracellular effects of intracellular peptide VFDVELL (VFD7) and related peptides on PPI. The internalization of the peptides was achieved using a C-terminus covalently bound cell-penetrating peptide (cpp; YGRKKRRQRRR). The possible inhibition of PPI was investigated using a NanoBiT ® luciferase structural complementation reporter system, with a pair of plasmids vectors each encoding, simultaneously, either FK506-binding protein (FKBP) or FKBP-binding domain (FRB) of mechanistic target of rapamycin complex 1 (mTORC1). The interaction of FKBP-FRB within cells occurs under rapamycin induction. Results shown that rapamycin-induced interaction between FKBP-FRB within human embryonic kidney 293 (HEK293) cells was inhibited by VFD7-cpp (10-500 nM) and FDVELLYGRKKRRQRRR (VFD6-cpp; 1-500 nM); additional VFD7-cpp derivatives were either less or not effective in inhibiting FKBP-FRB interaction induced by rapamycin. Molecular dynamics simulations suggested that selected peptides, such as VFD7-cpp, VFD6-cpp, VFAVELLYGRKKKRRQRRR (VFA7-cpp), and VFEVELLYGRKKKRRQRRR (VFA7-cpp), bind to FKBP and to FRB protein surfaces. However, only VFD7-cpp and VFD6-cpp induced changes on FKBP structure, which could help with understanding their mechanism of PPI inhibition. InPeps extracted from HEK293 cells were found mainly associated with macromolecular components (i.e., proteins and/or nucleic acids), contributing to understanding InPeps' intracellular proteolytic stability and mechanism of action-inhibiting PPI within cells. In a model of cell death induced by hypoxia-reoxygenation, VFD6-cpp (1 µM) increased the viability of mouse embryonic fibroblasts cells (MEF) expressing mTORC1-regulated autophagy-related gene 5 (Atg5), but not in autophagy-deficient MEF cells lacking the expression of Atg5. These data suggest that VFD6-cpp could have therapeutic applications reducing undesired side effects of rapamycin long-term treatments. In summary, the present report provides further evidence that InPeps have biological significance and could be valuable tools for the rational design of therapeutic molecules targeting intracellular PPI.

Intracellular peptides were shown to derive from proteasomal degradation of proteins from mammalian and yeast cells, being suggested to play distinctive roles both inside and outside these cells. Here, the role of intracellular peptides previously identified from skeletal muscle and adipose tissues of C57BL6/N wild type (WT) and neurolysin knockout mice were investigated. In differentiated C2C12 mouse skeletal muscle cells, some of these intracellular peptides like insulin activated the expression of several genes related to muscle contraction and gluconeogenesis. One of these peptides, LASVSTVLTSKYR (Ric4; 600 µg/kg), administrated either intraperitoneally or orally in WT mice, decreased glycemia. Neither insulin (10 nM) nor Ric4 (100 µM) induced glucose uptake in adipose tissue explants obtained from conditional knockout mice depleted of insulin receptor. Ric4 (100 µM) similarly to insulin (100 nM) induced Glut4 translocation to the plasma membrane of C2C12 differentiated cells, and increased GLUT4 mRNA levels in epididymal adipose tissue of WT mice. Ric4 (100 µM) increased both Erk and Akt phosphorylation in C2C12, as well as in epididymal adipose tissue from WT mice; Erk, but not Akt phosphorylation was activated by Ric4 in tibial skeletal muscle from WT mice. Ric4 is rapidly degraded in vitro by WT liver and kidney crude extracts, such a response that is largely reduced by structural modifications such as N-terminal acetylation, C-terminal amidation, and substitution of Leu8 for DLeu8 (Ac-LASVSTV[DLeu]TSKYR-NH2; Ric4-16). Ric4-16, among several Ric4 derivatives, efficiently induced glucose uptake in differentiated C2C12 cells. Among six Ric4-derivatives evaluated in vivo, Ac-LASVSTVLTSKYR-NH2 (Ric4-2; 600 µg/kg) and Ac-LASVSTV[DLeu]TSKYR (Ric4-15; 600 µg/kg) administrated orally efficiently reduced glycemia in a glucose tolerance test in WT mice. The potential clinical application of Ric4 and Ric4-derivatives deserves further attention.

Deletion of mechanistic target of rapamycin complex 2 (mTORC2) essential component rapamycin insensitive companion of mTOR (Rictor) by a Cre recombinase under control of the broad, nonadipocyte-specific aP2/FABP4 promoter impairs thermoregulation and brown adipose tissue (BAT) glucose uptake on acute cold exposure. We investigated herein whether adipocyte-specific mTORC2 deficiency affects BAT and inguinal white adipose tissue (iWAT) signaling, metabolism, and thermogenesis in cold-acclimated mice. For this, 8-wk-old male mice bearing Rictor deletion and therefore mTORC2 deficiency in adipocytes (adiponectin-Cre) and littermates controls were either kept at thermoneutrality (30 ± 1°C) or cold-acclimated (10 ± 1°C) for 14 days and evaluated for BAT and iWAT signaling, metabolism, and thermogenesis. Cold acclimation inhibited mTORC2 in BAT and iWAT, but its residual activity is still required for the cold-induced increases in BAT adipocyte number, total UCP-1 content and mRNA levels of proliferation markers Ki67 and cyclin 1 D, and de novo lipogenesis enzymes ATP-citrate lyase and acetyl-CoA carboxylase. In iWAT, mTORC2 residual activity is partially required for the cold-induced increases in multilocular adipocytes, mitochondrial mass, and uncoupling protein 1 (UCP-1) content. Conversely, BAT mTORC1 activity and BAT and iWAT glucose uptake were upregulated by cold independently of mTORC2. Noteworthy, the impairment in BAT and iWAT total UCP-1 content and thermogenic capacity induced by adipocyte mTORC2 deficiency had no major impact on whole body energy expenditure in cold-acclimated mice due to a compensatory activation of muscle shivering. In conclusion, adipocyte mTORC2 deficiency impairs, through different mechanisms, BAT and iWAT total UCP-1 content and thermogenic capacity in cold-acclimated mice, without affecting glucose uptake and whole body energy expenditure. NEW & NOTEWORTHY BAT and iWAT mTORC2 is inhibited by cold acclimation, but its residual activity is required for cold-induced increases in total UCP-1 content and thermogenic capacity, but not glucose uptake and mTORC1 activity. The impaired BAT and iWAT total UCP-1 content and thermogenic capacity induced by adipocyte mTORC2 deficiency are compensated by activation of muscle shivering in cold-acclimated mice.

Visceral obesity may be a driving factor in nonalcoholic fatty liver disease (NAFLD) development. Previous studies have shown that the omega-3 polyunsaturated fatty acid, eicosapentaenoic acid (EPA), ameliorates obesity in high-fat (HF) fed male, C57Bl/6 mice at thermoneutral conditions, independent of uncoupling protein 1 (UCP1). Our goals herein were to investigate sex-dependent mechanisms of EPA in the livers of wild type (WT) and UCP1 knockout (KO) male and female mice fed a HF diet (45% kcal fat; WT-HF, KO-HF) with or without supplementation of 36 g/kg EPA (WT-EPA, KO-EPA). KO significantly increased body weight in males, with no significant reductions with EPA in the WT or KO groups. In females, there were no significant differences in body weight among KO groups and no effects of EPA. In males, liver TGs were significantly higher in the KO-HF group and reduced with EPA, which was not observed in females. Accordingly, gene and protein markers of mitochondrial oxidation, peroxisomal biogenesis and oxidation, as well as metabolic futile cycles were sex-dependently impacted by KO and EPA supplementation. These findings suggest a genotypic difference in response to dietary EPA supplementation on the livers of male and female mice with diet-induced obesity and housed at thermoneutrality.

The nutrient sensors peroxisome proliferator-activated receptor γ (PPARγ) and mechanistic target of rapamycin complex 1 (mTORC1) closely interact in the regulation of adipocyte lipid storage. The precise mechanisms underlying this interaction and whether this extends to other metabolic processes and the endocrine function of adipocytes are still unknown. We investigated herein the involvement of mTORC1 as a mediator of the actions of the PPARγ ligand rosiglitazone in subcutaneous inguinal white adipose tissue (iWAT) mass, endocrine function, lipidome, transcriptome and branched-chain amino acid (BCAA) metabolism. Mice bearing regulatory associated protein of mTOR (Raptor) deletion and therefore mTORC1 deficiency exclusively in adipocytes and littermate controls were fed a high-fat diet supplemented or not with the PPARγ agonist rosiglitazone (30 mg/kg/day) for 8 weeks and evaluated for iWAT mass, lipidome, transcriptome (Rnaseq), respiration and BCAA metabolism. Adipocyte mTORC1 deficiency not only impaired iWAT adiponectin transcription, synthesis and secretion, PEPCK mRNA levels, triacylglycerol synthesis and BCAA oxidation and mRNA levels of related proteins but also completely blocked the upregulation in these processes induced by pharmacological PPARγ activation with rosiglitazone. Mechanistically, adipocyte mTORC1 deficiency impairs PPARγ transcriptional activity by reducing PPARγ protein content, as well as by downregulating C/EBPα, a co-partner and facilitator of PPARγ. In conclusion, mTORC1 and PPARγ are essential partners involved in the regulation of subcutaneous adipose tissue adiponectin production and secretion and BCAA oxidative metabolism., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Obesity is a critical public health problem worldwide that has been associated to non-communicable diseases (NCD), such as type 2 diabetes (T2DM), non-alcoholic fatty lipid diseases (NAFLD) and inflammatory diseases. Polyphenols from several food sources have been studied as one option against these health problems. Sabara jaboticaba (Plinia jaboticaba (Vell.) Berg) is a Brazilian berry rich in ellagic acid derivatives and anthocyanins. Here we investigated the effects of a phenolic-rich extract from Sabara jaboticaba (PEJ) in a diet-induced obesity animal model. PEJ at two doses, 50 mg gallic acid equivalent (GAE)/kg body weight (BW) and 100 mg GAE/kg BW, were administered by daily gavage to obese C57BL/6J mice for 14 weeks. PEJ prevented the excessive body weight and adiposity, adipocyte hypertrophy, inflammation, hyperglycemia, glucose intolerance, insulin resistance, hypercholesterolemia, and hepatic lipid accumulation, as well as increased energy expenditure. In conclusion, polyphenols from Sabara jaboticaba presented several powerful therapeutic properties relevant for fighting obesity and associated health problems., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Production of the proinflammatory cytokine tumor necrosis factor (TNF) must be precisely regulated for effective host immunity without the induction of collateral tissue damage. Here, we showed that TNF production was driven by a spleen-liver axis in a rat model of systemic inflammation induced by bacterial lipopolysaccharide (LPS). Analysis of cytokine expression and secretion in combination with splenectomy and hepatectomy revealed that the spleen generated not only TNF but also factors that enhanced TNF production by the liver, the latter of which accounted for nearly half of the TNF secreted into the circulation. Using mass spectrometry-based lipidomics, we identified leukotriene B 4 (LTB 4 ) as a candidate blood-borne messenger in this spleen-liver axis. LTB 4 was essential for spleen-liver communication in vivo, as well as for humoral signaling between splenic macrophages and Kupffer cells in vitro. LPS stimulated the splenic macrophages to secrete LTB 4 , which primed Kupffer cells to secrete more TNF in response to LPS in a manner dependent on LTB 4 receptors. These findings provide a framework to understand how systemic inflammation can be regulated at the level of interorgan communication., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Background: Obesity, characterized by excessive expansion of white adipose tissue (WAT), is associated with numerous metabolic complications. Conversely, brown adipose tissue (BAT) and beige fat are thermogenic tissues that protect mice against obesity and related metabolic disorders. We recently reported that deletion of miR-22 enhances energy expenditure and attenuates WAT expansion in response to a high-fat diet (HFD). However, the molecular mechanisms involved in these effects mediated by miR-22 loss are unclear., Methods and Results: Here, we show that miR-22 expression is induced during white, beige, and brown adipocyte differentiation in vitro. Deletion of miR-22 reduced white adipocyte differentiation in vitro. Loss of miR-22 prevented HFD-induced expression of adipogenic/lipogenic markers and adipocyte hypertrophy in murine WAT. In addition, deletion of miR-22 protected mice against HFD-induced mitochondrial dysfunction in WAT and BAT. Loss of miR-22 induced WAT browning. Gain- and loss-of-function studies revealed that miR-22 did not affect brown adipogenesis in vitro. Interestingly, miR-22 KO mice fed a HFD displayed increased expression of genes involved in thermogenesis and adrenergic signaling in BAT when compared to WT mice fed the same diet., Conclusions: Collectively, our findings suggest that loss of miR-22 attenuates fat accumulation in response to a HFD by reducing white adipocyte differentiation and increasing BAT activity, reinforcing miR-22 as a potential therapeutic target for obesity-related disorders., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Polyphenols from cambuci (CBC) (Campomanesia phaea (O. Berg.)), a Brazilian native fruit, were investigated on therapeutic actions mitigating insulin resistance and hepatic steatosis in high-fat-sucrose diet (HFS) induced obese mice. For this, C57BL/6J mice fed with a obesogenic and diabetogenic HFS diet were administered with either water or two CBC doses (36 or 74 mg gallic acid equivalent (GAE)/kg body weight) by gavage from week 6 to week 14 (end-point) of HFS feeding. CBC reduced body weight gain, inflammation, hepatic steatosis, hyperglycemia, glucose intolerance, and insulin resistance in liver and skeletal muscle of obese mice, and such effects were associated with activation of Akt and AMPK in these tissues. In conclusion, polyphenols from CBC show important therapeutic actions ameliorating obesity-associated complications., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Dietary sugar is an important determinant of the development and progression of nonalcoholic fatty liver disease (NAFLD). However, the molecular mechanisms underlying the deleterious effects of sugar intake on NAFLD under energy-balanced conditions are still poorly understood. Here, we provide a comprehensive analysis of the liver lipidome and mechanistic insights into the pathogenesis of NAFLD induced by the chronic consumption of high-sugar diet (HSD). Newly weaned male Wistar rats were fed either a standard chow diet or an isocaloric HSD for 18 weeks. Livers were harvested for histological, oxidative stress, gene expression, and lipidomic analyses. Intake of HSD increased oxidative stress and induced severe liver injury, microvesicular steatosis, and ballooning degeneration of hepatocytes. Using untargeted lipidomics, we identified and quantified 362 lipid species in the liver. Rats fed with HSD displayed increased hepatic levels of triacylglycerol enriched in saturated and monounsaturated fatty acids, lipids related to mitochondrial function/structure (phosphatidylglycerol, cardiolipin, and ubiquinone), and acylcarnitine (an intermediate lipid of fatty acid beta-oxidation). HSD-fed animals also presented increased levels of some species of membrane lipids and a decreased content of phospholipids containing omega-6 fatty acids. These changes in the lipidome were associated with the downregulation of genes involved in fatty acid oxidation in the liver. In conclusion, our data suggest that the chronic intake of a HSD, even under isocaloric conditions, induces lipid overload, and inefficient/impaired fatty acid oxidation in the liver. Such events lead to marked disturbance in hepatic lipid metabolism and the development of NAFLD., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Scope: Evidence gathered in the last decades suggests that lipotoxicity and inflammation are the main factors connecting adipose tissue dysfunction to the development of metabolic diseases such as insulin resistance, nonalcoholic fatty liver disease (NAFLD), cardiovascular disease, and certain types of cancer, among others. The mechanistic target of rapamycin (mTOR) is a serine threonine kinase that functions as the catalytic entity of two multiprotein complexes, mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). These complexes are important components of signaling pathways activated by nutrients, growth factors, and inflammatory mediators and are therefore directly involved in the regulation of adipocyte and macrophage metabolism and function., Methods and Results: In this article, studies that evaluate the involvement of mTORC1 and 2 in the regulation of macrophage and adipocyte function and their implication in the development of metabolic-disease-associated adipose tissue dysfunction are reviewed., Conclusion: In adipocytes, optimal levels of mTORC1 activity are required for its pro-lipogenic actions, while in macrophages, mTORC1 regulates features of both M1 and M2 polarization. mTORC2, on the other hand, promotes glucose uptake and de novo lipogenesis in adipocytes and counteracts macrophage inflammatory response., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

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 ± 0.1 g) 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.

Palmitoleic acid (POA, 16:1n-7) is a lipokine that has potential nutraceutical use to treat non-alcoholic fatty liver disease. We tested the effects of POA supplementation (daily oral gavage, 300 mg/Kg, 15 days) on murine liver inflammation induced by a high fat diet (HFD, 59% fat, 12 weeks). In HFD-fed mice, POA supplementation reduced serum insulin and improved insulin tolerance compared with oleic acid (OA, 300 mg/Kg). The livers of POA-treated mice exhibited less steatosis and inflammation than those of OA-treated mice with lower inflammatory cytokine levels and reduced toll-like receptor 4 protein content. The anti-inflammatory effects of POA in the liver were accompanied by a reduction in liver macrophages (LM, CD11c + ; F4/80 + ; CD86 + ), an effect that could be triggered by peroxisome proliferator activated receptor (PPAR)-γ, a lipogenic transcription factor upregulated in livers of POA-treated mice. We also used HFD-fed mice with selective deletion of PPAR-γ in myeloid cells (PPAR-γ KO LyzCre+ ) to test whether the beneficial anti-inflammatory effects of POA are dependent on macrophages PPAR-γ. POA-mediated improvement of insulin tolerance was tightly dependent on myeloid PPAR-γ, while POA anti-inflammatory actions including the reduction in liver inflammatory cytokines were preserved in mice bearing myeloid cells deficient in PPAR-γ. This overlapped with increased CD206 + (M2a) cells and downregulation of CD86 + and CD11c + liver macrophages. Moreover, POA supplementation increased hepatic AMPK activity and decreased expression of the fatty acid binding scavenger receptor, CD36. We conclude that POA controls liver inflammation triggered by fat accumulation through induction of M2a macrophages independently of myeloid cell PPAR-γ., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

DICER is a key enzyme in microRNA (miRNA) biogenesis. Here we show that aerobic exercise training up-regulates DICER in adipose tissue of mice and humans. This can be mimicked by infusion of serum from exercised mice into sedentary mice and depends on AMPK-mediated signaling in both muscle and adipocytes. Adipocyte DICER is required for whole-body metabolic adaptations to aerobic exercise training, in part, by allowing controlled substrate utilization in adipose tissue, which, in turn, supports skeletal muscle function. Exercise training increases overall miRNA expression in adipose tissue, and up-regulation of miR-203-3p limits glycolysis in adipose under conditions of metabolic stress. We propose that exercise training-induced DICER-miR-203-3p up-regulation in adipocytes is a key adaptive response that coordinates signals from working muscle to promote whole-body metabolic adaptations., Competing Interests: The authors declare no competing interest.

mTOR, a serine/threonine protein kinase that is involved in a series of critical cellular processes, can be found in two functionally distinct complexes, mTORC1 and mTORC2. In contrast to mTORC1, little is known about the mechanisms that regulate mTORC2. Here we show that mTORC2 activity is reduced in mice with a hypomorphic mutation of the Ric-8B gene. Ric-8B is a highly conserved protein that acts as a non-canonical guanine nucleotide exchange factor (GEF) for heterotrimeric Gαs/olf type subunits. We found that Ric-8B hypomorph embryos are smaller than their wild type littermates, fail to close the neural tube in the cephalic region and die during mid-embryogenesis. Comparative transcriptome analysis revealed that signaling pathways involving GPCRs and G proteins are dysregulated in the Ric-8B mutant embryos. Interestingly, this analysis also revealed an unexpected impairment of the mTOR signaling pathway. Phosphorylation of Akt at Ser473 is downregulated in the Ric-8B mutant embryos, indicating a decreased activity of mTORC2. Knockdown of the endogenous Ric-8B gene in cultured cell lines leads to reduced phosphorylation levels of Akt (Ser473), further supporting the involvement of Ric-8B in mTORC2 activity. Our results reveal a crucial role for Ric-8B in development and provide novel insights into the signals that regulate mTORC2., Competing Interests: The authors have declared that no competing interests exist.

Moderate aerobic training may be therapeutic for chronic low-grade inflammatory diseases due to the associated anti-inflammatory response that is mediated by immune cells. The peroxisome proliferator-activated receptor gamma (PPARγ) regulates the M1 (pro-inflammatory) and M2 (anti-inflammatory) polarization, as well as the immunometabolic response of macrophages. Against this background, the present study seeks to clarify whether the conditional deletion of PPARγ in macrophages would have any effect on the anti-inflammatory role of moderate aerobic training. To test this hypothesis, two mice strains were used: PPARγ LyzCre+/+ (KO) and littermates control animals (WT). Each genotype was divided into 1) sedentary high-fat diet (HF) and 2) high-fat diet and moderate aerobic training (HFT) (n = 5-8 per group). The experimental protocol lasted for 12 weeks, comprising 4 weeks of HF diet only and 8 weeks of HF diet and aerobic training (5 times/week, 50-60 minutes/day at 60% of maximum speed). Metabolic analyses were carried out on the serum glucose homeostase, adipose tissue morphology and cytokine content, and macrophage cytokine production.Immunophenotyping and gene expression were also performed. KO male mice were more prone to hypertrophy in the subcutaneous adipose tissue, though only the IL-1β (p = 0.0049) was higher compared to the values observed in WT animals. Peritoneal macrophages from KO animals exhibited a marked inflammatory environment with an increase in TNF-α (p = 0.0008), IL- 1β (p = 0.0017), and IL-6 (p < 0.0001) after lipopolysaccharide stimulation. The moderate aerobic training protected both genotypes from weight gain and reduced the caloric intake in the KO animals. Despite the attenuation of the M2 marker CD206 (p < 0.001) in the absence of PPAR-γ, the aerobic training modulated cytokine production in LPS stimulated peritoneal macrophages from both genotypes, reducing proinflammatory cytokines such as TNF-α (p = 0.0002) and IL-6 (p < 0.0001). Overall, our findings demonstrate the essential role of PPARγ in macrophage immunophenotypes. However, the deletion of PPARγ did not inhibit the exercise-mediated anti-inflammatory effect, underscoring the important role of exercise in modulating inflammation., (Copyright © 2020 International Society of Exercise and Immunology. All rights reserved.)

Objectives: Hepatokines are proteins secreted by the liver that impact the functions of the liver and various tissues through autocrine, paracrine, and endocrine signaling. Recently, Tsukushi (TSK) was identified as a new hepatokine that is induced by obesity and cold exposure. It was proposed that TSK controls sympathetic innervation and thermogenesis in brown adipose tissue (BAT) and that loss of TSK protects against diet-induced obesity and improves glucose homeostasis. Here we report the impact of deleting and/or overexpressing TSK on BAT thermogenic capacity, body weight regulation, and glucose homeostasis., Methods: We measured the expression of thermogenic genes and markers of BAT innervation and activation in TSK-null and TSK-overexpressing mice. Body weight, body temperature, and parameters of glucose homeostasis were also assessed in the context of TSK loss and overexpression., Results: The loss of TSK did not affect the thermogenic activation of BAT. We found that TSK-null mice were not protected against the development of obesity and did not show improvement in glucose tolerance. The overexpression of TSK also failed to modulate thermogenesis, body weight gain, and glucose homeostasis in mice., Conclusions: TSK is not a significant regulator of BAT thermogenesis and is unlikely to represent an effective target to prevent obesity and improve glucose homeostasis., (Copyright © 2019 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Objective: Dietary restriction (DR) improves health and prolongs lifespan in part by upregulating type III endoribonuclease DICER in adipose tissue. In this study, we aimed to specifically test which missing dietary component was responsible for DICER upregulation., Methods: We performed a nutrient screen in mouse preadipocytes and validated the results in vivo using different kinds of dietary interventions in wild type or genetically modified mice and worms, also testing the requirement of DICER on the effects of the diets., Results: We found that sulfur amino acid restriction (i.e., methionine or cysteine) is sufficient to increase Dicer mRNA expression in preadipocytes. Consistently, while DR increases DICER expression in adipose tissue of mice, this effect is blunted by supplementation of the diet with methionine, cysteine, or casein, but not with a lipid or carbohydrate source. Accordingly, dietary methionine or protein restriction mirrors the effects of DR. These changes are associated with alterations in serum adiponectin. We also found that DICER controls and is controlled by adiponectin. In mice, DICER plays a role in methionine restriction-induced upregulation of Ucp1 in adipose tissue. In C. elegans, DR and a model of methionine restriction also promote DICER expression in the intestine (an analog of the adipose tissue) and prolong lifespan in a DICER-dependent manner., Conclusions: We propose an evolutionary conserved mechanism in which dietary sulfur amino acid restriction upregulates DICER levels in adipose tissue leading to beneficial health effects., (Copyright © 2019 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Cachexia is a wasting syndrome characterized by the continuous loss of skeletal muscle mass due to imbalance between protein synthesis and degradation, which is related with poor prognosis and compromised quality of life. Dysfunctional mitochondria are associated with lower muscle strength and muscle atrophy in cancer patients, yet poorly described in human cachexia. We herein investigated mitochondrial morphology, autophagy and apoptosis in the skeletal muscle of patients with gastrointestinal cancer-associated cachexia (CC), as compared with a weight-stable cancer group (WSC). CC showed prominent weight loss and increased circulating levels of serum C-reactive protein, lower body mass index and decreased circulating hemoglobin, when compared to WSC. Electron microscopy analysis revealed an increase in intermyofibrillar mitochondrial area in CC, as compared to WSC. Relative gene expression of Fission 1, a protein related to mitochondrial fission, was increased in CC, as compared to WSC. LC3 II, autophagy-related (ATG) 5 and 7 essential proteins for autophagosome formation, presented higher content in the cachectic group. Protein levels of phosphorylated p53 (Ser46), activated caspase 8 (Asp384) and 9 (Asp315) were also increased in the skeletal muscle of CC. Overall, our results demonstrate that human cancer-associated cachexia leads to exacerbated muscle-stress response that may culminate in muscle loss, which is in part due to disruption of mitochondrial morphology, dysfunctional autophagy and increased apoptosis. To the best of our knowledge, this is the first report showing quantitative morphological alterations in skeletal muscle mitochondria in cachectic patients.

Nonalcoholic fatty liver disease (NAFLD) prevails in obesity and is linked to several health complications including dyslipidemia and atherosclerosis. How exactly NAFLD induces atherogenic dyslipidemia to promote cardiovascular diseases is still elusive. Here, we identify Tsukushi (TSK) as a hepatokine induced in response to NAFLD. We show that both endoplasmic reticulum stress and inflammation promote the expression and release of TSK in mice. In humans, hepatic TSK expression is also associated with steatosis, and its circulating levels are markedly increased in patients suffering from acetaminophen-induced acute liver failure (ALF), a condition linked to severe hepatic inflammation. In these patients, elevated blood TSK levels were associated with decreased transplant-free survival at hospital discharge, suggesting that TSK could have a prognostic significance. Gain- and loss-of-function studies in mice revealed that TSK impacts systemic cholesterol homeostasis. TSK reduces circulating HDL cholesterol, lowers cholesterol efflux capacity, and decreases cholesterol-to-bile acid conversion in the liver. Our data identify the hepatokine TSK as a blood biomarker of liver stress that could link NAFLD to the development of atherogenic dyslipidemia and atherosclerosis.

Scope: Brown adipose tissue (BAT) dissipates energy through uncoupling protein 1 (UCP1) and has been proposed as an anti-obesity target. It was reported previously that a high-fat (HF) diet enriched in eicosapentaenoic acid (EPA) significantly increased UCP1 and other thermogenic markers in BAT. It is hypothesized that these effects are mediated through UCP1-dependent regulation., Methods and Results: Wild-type (WT) and UCP1 knockout (KO) B6 male mice were housed at thermoneutrality and fed a HF diet, without or with eicosapentaenoic acid (EPA)-enriched fish oil. HF-fed KO mice were heavier and had higher BAT lipid content than other groups. Protective effects of EPA in WT, previously observed at 22 °C (reduced adiposity, improved glucose tolerance, and increased UCP1), disappeared at thermoneutrality. Mitochondrial proteins, cytochrome c oxidase subunit 1 (COX I), COX I, II, and IV were reduced in the KO mice compared to WT. Unexpectedly, EPA attenuated weight and fat mass gain and improved glucose tolerance in the KO mice. Finally, EPA increased BAT peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α) protein and gene expression, and whole-body oxygen consumption in KO mice, consistent with increased mitochondria DNA (mtDNA)/nuclear DNA (nucDNA) ratio., Conclusions: EPA rescued the weight gain and glucose intolerance in UCP1 KO mice at thermoneutrality, independent of UCP1; these effects may be mediated in part via increased oxygen consumption and BAT PGC1α., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Scope: The mechanisms and involvement of uncoupling protein 1 (UCP1) in the protection from obesity and insulin resistance induced by intake of a high-fat diet rich in omega-3 (n-3) fatty acids are investigated., Methods and Results: C57BL/6J mice are fed either a low-fat (control group) or one of two isocaloric high-fat diets containing either lard (HFD) or fish oil (HFN3) as fat source and evaluated for body weight, adiposity, energy expenditure, glucose homeostasis, and inguinal white and interscapular brown adipose tissue (iWAT and iBAT, respectively) gene expression, lipidome, and mitochondrial bioenergetics. HFN3 intake protected from obesity, glucose and insulin intolerances, and hyperinsulinemia. This is associated with increased energy expenditure, iWAT UCP1 expression, and incorporation of n-3 eicosapentaenoic and docosahexaenoic fatty acids in iWAT and iBAT triacylglycerol. Importantly, HFN3 is equally effective in reducing body weight gain, adiposity, and glucose intolerance and increasing energy expenditure in wild-type and UCP1-deficient mice without recruiting other thermogenic processes in iWAT and iBAT, such as mitochondrial uncoupling and SERCA-mediated calcium and creatine-driven substrate cyclings., Conclusion: Intake of a high-fat diet rich in omega-3 fatty acids protects both wild-type and UCP1-deficient mice from obesity and insulin resistance by increasing energy expenditure through unknown mechanisms., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Exercise is a powerful tool for prevention and treatment of many conditions related to the cardiovascular system and also chronic low-grade inflammation. Peroxisome proliferator-activated receptors γ (PPARγ) exerts an import role on the regulation of metabolic profile and subsequent inflammatory response, especially in macrophages., Purpose: To investigate the effects of 8-week moderate-exercise training on metabolic and inflammatory parameters in mice with PPARγ deficiency in myeloid cells., Methods: Twelve-week old mice bearing PPARγ deletion exclusively in myeloid cells (PPARγlox/lox Lys Cre -/+ , knockout [KO]) and littermate controls (PPARγlox/lox Lys Cre -/- , wild type [WT]) were submitted to 8-week exercise training (treadmill running at moderate intensity, 5 days/week). Animals were evaluated for food intake, glucose homeostasis, serum metabolites, adipose tissue and peritoneal macrophage inflammation, and basal and stimulated cytokine secretion., Results: Exercise protocol did not improve glucose metabolism or adiponectin concentrations in serum of KO mice. Moreover, the absence of PPARγ in macrophages exacerbated the proinflammatory profile in sedentary mice. Peritoneal cultured cells had higher tumor necrosis factor-α (TNF-α) secretion in nonstimulated and lipopolysaccharide (LPS)-stimulated conditions and higher Toll-4 receptor (TLR4) gene expression under LPS stimulus. Trained mice showed reduced TNF-α content in adipose tissue independently of the genotype. M2 polarization ability was impaired in KO peritoneal macrophages after exercise training, while adipose tissue-associated macrophages did not present any effect by PPARγ ablation., Conclusion: Overall, PPARγ seems necessary to maintain macrophages appropriate response to inflammatory stimulus and macrophage polarization, affecting also whole body lipid metabolism and adiponectin profile. Exercise training showed as an efficient mechanism to restore the immune response impaired by PPARγ deletion in macrophages., (© 2018 Wiley Periodicals, Inc.)

Background: Isocitrate deyhydrogenase (IDH) mutant glioma comprises the majority of grades II-III gliomas and nearly all secondary glioblastomas. These progressive gliomas arise from mutations in IDH1 or IDH2 that pathologically produce D-2-hydroxyglutarate (2HG), which interferes with cell reactions using alpha ketoglutarate, leading to a hypermethylated genome and epigenetic dysregulation of gene expression initiating tumorigenesis., Methods: Human IDH1 wild type (wt) and IDH1 R132H cell lines and patient-derived xenografts (PDXs) were used to evaluate the FDA-approved DNA demethylating agent 5-azacytidine (5-aza). Cell growth, protein and gene expression, chromatin immunoprecipitation, and nucleosome position assays were performed in 5-aza treated cells. To evaluate antitumor activity in vivo, 5-aza was administered alone and in combination with temozolomide (TMZ) in a PDX glioma model harboring IDH1 R132H mutation., Results: 5-Aza treatment has been found to reduce cell growth and increase expression of glial fibrillary acid protein (GFAP). Chromatin immunoprecipitation and nucleosome position assay showed that the mechanism of increased GFAP expression induction is associated with histone modification and nucleosome repositioning of the GFAP promoter, respectively. In vivo, 5-aza treatment extended survival in IDH1 R132H mutant but not in an IDH1 wt glioma model. Additionally, 5-aza enhances the therapeutic effect of the DNA damaging agent TMZ in both subcutaneous and orthotopic PDX models of IDH1 R132H mutant glioma., Conclusion: 5-Aza provided a survival benefit as a single agent but worked best in combination with TMZ in 2 different IDH1 R132H mutant glioma models., (© The Author(s) 2018. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Malaria in pregnancy is a public health concern in malaria-endemic areas. Accumulation of maternal immune cells in the placenta and increased levels of inflammatory cytokines caused by sequestration of Plasmodium falciparum -infected erythrocytes have been associated to poor neonatal outcomes, including low birth weight because of fetal growth restriction. Little is known about the molecular changes occurring in a P. falciparum -infected placenta that has developed placental malaria during pregnancy but had the parasites cleared by pharmacological treatment (past infection). We conducted an integrated proteome, phosphoproteome and glycoproteome analysis in past P. falciparum -infected placentas aiming to find molecular changes associated with placental malaria. A total of 2946 proteins, 1733 N -linked glycosites and 4100 phosphosites were identified and quantified in this study, disclosing overrepresented processes related to oxidative stress, protein folding and regulation of apoptosis in past-infected placentas Moreover, AKT and ERK signaling pathways activation, together with clinical data, were further correlated to an increased apoptosis in past-infected placentas. This study showed apoptosis-related mechanisms associated with placental malaria that can be further explored as therapeutic target against adverse pregnancy outcomes., (© 2019 Kawahara et al.)

Objective: We investigated whether PPARγ modulates adipose tissue BCAA metabolism, and whether this mediates the attenuation of obesity-associated insulin resistance induced by pharmacological PPARγ activation., Methods: Mice with adipocyte deletion of one or two PPARγ copies fed a chow diet and rats fed either chow, or high fat (HF) or HF supplemented with BCAA (HF/BCAA) diets treated with rosiglitazone (30 or 15 mg/kg/day, 14 days) were evaluated for glucose and BCAA homeostasis., Results: Adipocyte deletion of one PPARγ copy increased mice serum BCAA and reduced inguinal white (iWAT) and brown (BAT) adipose tissue BCAA incorporation into triacylglycerol, as well as mRNA levels of branched-chain aminotransferase (BCAT)2 and branched-chain α-ketoacid dehydrogenase (BCKDH) complex subunits. Adipocyte deletion of two PPARγ copies induced lipodystrophy, severe glucose intolerance and markedly increased serum BCAA. Rosiglitazone abolished the increase in serum BCAA induced by adipocyte PPARγ deletion. In rats, HF increased serum BCAA, such levels being further increased by BCAA supplementation. Rosiglitazone, independently of diet, lowered serum BCAA and upregulated iWAT and BAT BCAT and BCKDH activities. This was associated with a reduction in mTORC1-dependent inhibitory serine phosphorylation of IRS1 in skeletal muscle and whole-body insulin resistance evaluated by HOMA-IR., Conclusions: PPARγ, through the regulation of both BAT and iWAT BCAA catabolism in lipoeutrophic mice and muscle insulin responsiveness and proteolysis in lipodystrophic mice, is a major determinant of circulating BCAA levels. PPARγ agonism, therefore, may improve whole-body and muscle insulin sensitivity by reducing blood BCAA, alleviating mTORC1-mediated inhibitory IRS1 phosphorylation., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Brown adipose tissue (BAT) thermogenesis is a key controller of energy metabolism. In response to cold or other adrenergic stimuli, brown adipocytes increase their substrate uptake and oxidative activity while uncoupling ATP synthesis from the mitochondrial respiratory chain activity. Brown adipocytes are found in classic depots such as in the interscapular BAT (iBAT). They can also develop in white adipose tissue (WAT), such as in the inguinal WAT (iWAT), where their presence has been associated with metabolic improvements. We previously reported that the induction of oxidative metabolism in iWAT is low compared with that of iBAT, even after sustained adrenergic stimulation. One explanation to this apparent lack of thermogenic ability of iWAT is the presence of an active iBAT, which may prevent the full activation of iWAT. In this study, we evaluated whether iBAT denervation-induced browning of white fat enhanced the thermogenic activity of iWAT following cold acclimation, under beta-3 adrenergic stimulation (CL 316,243). Following a bilateral denervation of iBAT, we assessed energy balance, evaluated the oxidative activity of iBAT and iWAT using 11 C-acetate, and quantified the dynamic glucose uptake of those tissues using 2-deoxy-2-[ 18 F]- fluoro-d-glucose. Our results indicate that despite portraying marked browning and mildly enhanced glucose uptake, iWAT of cold-adapted mice does not exhibit significant oxidative activity following beta-3 adrenergic stimulation in the absence of a functional iBAT. The present results suggest that iWAT is not readily recruitable as a thermogenic organ even when functional iBAT is lacking.

Scope: To test whether myeloid cells Tsc1 deletion and therefore constitutive activation of the nutrient sensor mTORC1 protects from high-fat diet (HFD)-induced obesity, glucose intolerance, and adipose tissue inflammation., Methods and Results: Mice with Tsc1 deletion in myeloid cells (MTsc1KO) and littermate controls (MTsc1WT) were fed with HFD for 8 weeks and evaluated for body weight, glucose homeostasis, and adipose tissue inflammation. MTsc1KO mice were protected from HFD-induced obesity and glucose intolerance. MTsc1KO, however, displayed, independently of the diet, abnormal behavior, episodes of intense movement, and muscle spasms followed by temporary paralysis. To investigate whether obesity protection was due to myeloid cells Tsc1 deletion, bone marrow was transplanted from MTsc1WT and MTsc1KO into irradiated C57BL6/J mice. Mice transplanted with MTsc1KO bone marrow displayed reduced body weight gain, adiposity, and inflammation, and enhanced energy expenditure, glucose tolerance and adipose tissue M2 macrophage content upon HFD feeding, in the absence of abnormal behavior. In vitro, Tsc1 deletion increased in a mTORC1-dependent manner macrophage polarization to M2 profile and mRNA levels of fatty acid binding protein 4 and PPARγ., Conclusion: Constitutive mTORC1 activation in myeloid cells protects mice from HFD-induced obesity, adipose tissue inflammation, and glucose intolerance by promoting macrophage polarization to M2 pro-resolution profile and increasing energy expenditure., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Visceral obesity is frequently associated with the development of type 2 diabetes (T2D), a highly prevalent chronic disease that features insulin resistance and pancreatic β -cell dysfunction as important hallmarks. Recent evidence indicates that the chronic, low-grade inflammation commonly associated with visceral obesity plays a major role connecting the excessive visceral fat deposition with the development of insulin resistance and pancreatic β -cell dysfunction. Herein, we review the mechanisms by which nutrients modulate obesity-associated inflammation.

Polyphenol-rich cagaita (Eugenia dysenterica DC.) extracts (PCE) have previously shown to prevent body weight and adiposity induced by high-fat/high-sucrose (HFS) diet. Whether PCE also exerts protective effects in already developed obesity is unknown. In order to test this hypothesis, male C57BL/6J obese mice (previously feed with a HFS diet for six weeks) were treated with PCE at two doses, 7mg gallic acid equivalent (GAE)/kg body weight (PCE I group), and 14mg GAE/kg body weight (PCE II group) or water (HFS and Chow groups) by oral gavage for eight weeks. PCE did not affect body weight and adiposity of obese mice. However, PCE did protect against dyslipidemia, fasting hyperglycemia, and glucose intolerance, and attenuated both hepatic gluconeogenesis and inflammation as observed by the expression of tumor necrosis factor-α and transcriptional factor NF-κB. These results indicate that PCE improves glucose homeostasis of obese mice by attenuating hepatic gluconeogenesis and inflammation., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Intracellular peptides generated by limited proteolysis are likely to function inside and outside cells and could represent new possibilities for drug development. Here, we used several conformational-sensitive antibodies targeting G-protein coupled receptors to screen for novel pharmacological active peptides. We find that one of these peptides, DITADDEPLT activates cannabinoid type 1 receptors. Single amino acid modifications identified a novel peptide, DIIADDEPLT (Pep19), with slightly better inverse agonist activity at cannabinoid type 1 receptors. Pep19 induced uncoupling protein 1 expression in both white adipose tissue and 3T3-L1 differentiated adipocytes; in the latter, Pep19 activates pERK1/2 and AKT signaling pathways. Uncoupling protein 1 expression induced by Pep19 in 3T3-L1 differentiated adipocytes is blocked by AM251, a cannabinoid type 1 receptors antagonist. Oral administration of Pep19 into diet-induced obese Wistar rats significantly reduces adiposity index, whole body weight, glucose, triacylglycerol, cholesterol and blood pressure, without altering heart rate; changes in the number and size of adipocytes were also observed. Pep19 has no central nervous system effects as suggested by the lack of brain c-Fos expression, cell toxicity, induction of the cannabinoid tetrad, depressive- and anxiety-like behaviors. Therefore, Pep19 has several advantages over previously identified peripherally active cannabinoid compounds, and could have clinical applications.

Obesity is characterized by an excessive increase in the adipose tissue mass, and is associated with higher incidence of several chronic metabolic diseases, such as type 2 diabetes. Therefore, its increasing prevalence is a public health concern, and it is important to better understand its etiology to develop new therapeutic strategies. Evidence accumulated over the years indicates that obesity is associated with a marked activation in adipose tissue of the mechanistic target of rapamycin complex 1 (mTORC1), a signaling pathway that controls lipid metabolism, and adipocyte formation and maintenance. Curiously, mTORC1 is also involved in the control of nonshivering thermogenesis and recruitment as well as browning of white adipose tissue. In this review, we explored mTORC1 functions in adipocytes and presented evidence, suggesting that mTORC1 may either increase or reduce adiposity, depending on the conditions and activation levels.

Mechanistic target of rapamycin complex (mTORC)1 activity is increased in adipose tissue of obese insulin-resistant mice, but its role in the regulation of tissue inflammation is unknown. Herein, we investigated the effects of adipocyte mTORC1 deficiency on adipose tissue inflammation and glucose homeostasis. For this, mice with adipocyte raptor deletion and controls fed a chow or a high-fat diet were evaluated for body mass, adiposity, glucose homeostasis, and adipose tissue inflammation. Despite reducing adiposity, adipocyte mTORC1 deficiency promoted hepatic steatosis, insulin resistance, and adipose tissue inflammation (increased infiltration of macrophages, neutrophils, and B lymphocytes; crown-like structure density; TNF-α, interleukin (IL)-6, and monocyte chemoattractant protein 1 expression; IL-1β protein content; lipid peroxidation; and de novo ceramide synthesis). The anti-oxidant, N -acetylcysteine, partially attenuated, whereas treatment with de novo ceramide synthesis inhibitor, myriocin, completely blocked adipose tissue inflammation and nucleotide oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3)-inflammasome activation, but not hepatic steatosis and insulin resistance induced by adipocyte raptor deletion. Rosiglitazone treatment, however, completely abrogated insulin resistance induced by adipocyte raptor deletion. In conclusion, adipocyte mTORC1 deficiency induces adipose tissue inflammation and NLRP3-inflammasome activation by promoting oxidative stress and de novo ceramide synthesis. Such adipose tissue inflammation, however, is not an underlying cause of the insulin resistance displayed by these mice., (Copyright © 2017 by the American Society for Biochemistry and Molecular Biology, Inc.)

Beige (or brite, "brown in white") adipocytes are uncoupling protein 1 (UCP1)-positive cells residing in white adipose depots that, depending on the conditions, behave either as classic white adipocytes, storing energy as lipids, or as brown adipocytes, dissipating energy from oxidative metabolism as heat through non-shivering thermogenesis. Because of their thermogenic potential and, therefore, possible usage to treat metabolic diseases such as obesity and type 2 diabetes, beige cells have attracted the attention of many scientists worldwide aiming to develop strategies to safely recruit and activate their thermogenic activity. Indeed, in recent years, a large variety of conditions, molecules (including nutrients) and signaling pathways were reported to promote the recruitment of beige adipocytes. Despite of those advances, the true contribution of beige adipocyte thermogenesis to whole-body energy expenditure is still not completely defined. Herein, we discuss some important aspects that should be considered when studying beige adipocyte biology and the contribution to energy balance and whole-body metabolism.

The underlying mechanism by which MyD88 regulates the development of obesity, metainflammation, and insulin resistance (IR) remains unknown. Global deletion of MyD88 in high-fat diet (HFD)-fed mice resulted in increased weight gain, impaired glucose homeostasis, elevated Dectin-1 expression in adipose tissue (AT), and proinflammatory CD11c+ AT macrophages (ATMs). Dectin-1 KO mice were protected from diet-induced obesity (DIO) and IR and had reduced CD11c+ AT macrophages. Dectin-1 antagonist improved glucose homeostasis and decreased CD11c+ AT macrophages in chow- and HFD-fed MyD88 KO mice. Dectin-1 agonist worsened glucose homeostasis in MyD88 KO mice. Dectin-1 expression is increased in AT from obese individuals. Together, our data indicate that Dectin-1 regulates AT inflammation by promoting CD11c+ AT macrophages in the absence of MyD88 and identify a role for Dectin-1 in chronic inflammatory states, such as obesity. This suggests that Dectin-1 may have therapeutic implications as a biomarker for metabolic dysregulation in humans., (Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.)

Genetic- and diet-induced obesity and insulin resistance are associated with an increase in mechanistic target of rapamycin complex (mTORC) 1 activity in adipose tissue. We investigated herein the effects of pharmacological mTORC1 inhibition in the development of adipose tissue inflammation induced by high-fat diet (HFD) feeding, as well as in the polarization, metabolism and function of bone marrow-derived macrophages (BMDM). For this, C57BL/6J mice fed with a standard chow diet or a HFD (60% of calories from fat) and treated with either vehicle (0.1% Me 2 SO, 0.2% methylcellulose) or rapamycin (2mg/kg/ day, gavage) during 30days were evaluated for body weight, adiposity, glucose tolerance and adipose tissue inflammation. Although rapamycin did not affect the increase in body weight and adiposity, it exacerbated the glucose intolerance and adipose tissue inflammation induced by HFD feeding, as evidenced by the increased adipose tissue percentage of M1 macrophages, naive and activated cytotoxic T lymphocytes, and mRNA levels of proinflammatory molecules, such as TNF-α, IL-6 and MCP-1. In BMDM in vitro, pharmacological mTORC1 inhibition induced phosphorylation of NFκB p65 and spontaneous polarization of macrophages to a proinflammatory M1 profile, while it impaired M2 polarization induced by IL-4+IL-13, glycolysis and phagocytosis. Altogether, these findings indicate that mTORC1 activity is an important determinant of adipose tissue inflammatory profile and macrophage plasticity, metabolism and function., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Brown adipose tissue (BAT), an organ specialized in the conversion of chemical energy from nutrients into heat through a process denominated as nonshivering thermogenesis, plays an important role in defence of body weight and homoeothermy in mammals. BAT nonshivering thermogenesis relies on the activity of the uncoupling protein 1 (UCP-1), a mitochondrial protein that, on demand, deviates proton gradient from ATP synthesis to heat generation. Energetically, this process is supported by BAT-elevated mitochondrial density and outstanding capacity to oxidize fatty acids and glucose. These unique features place BAT as an important determinant of whole-body energy, lipid and glucose homoeostases. In the present issue of Clinical Science, Poekes et al. have gathered supporting evidence indicating that, along with hyperphagia, impaired BAT diet-induced thermogenesis is an important factor driving the exacerbated diet-induced obesity, glucose intolerance and hepatic steatosis featured by foz/foz, a mouse strain that carries mutations in Alström syndrome protein 1 (ALMS1) gene mimicking human Alström syndrome. They also show that restoration of BAT nonshivering thermogenesis by intermittent cold exposure attenuated foz/foz mice obesity, glucose intolerance and liver steatosis. Altogether, these findings highlight the important contribution of BAT nonshivering thermogenesis to whole-body energy expenditure, lipid and glucose homoeostases and further support its potential utilization as a therapeutic strategy to treat metabolic diseases., (© 2017 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.)