Dietary Sources, Sex, and rs5888 (SCARB1) as Modulators of Vitamin A's Effect on Cardiometabolic Health.

Although preclinical studies have attributed vitamin A (VA) cardiometabolic benefits, these effects are still controversial and not always supported in large human studies. Here, the outcomes associated with VA and its relationship with habitual dietary sources, sex, and genetic background have been studied. To do so, the data from an observational study (n = 455) (64% females, mean age of 36 years) showing that suboptimal VA intake (mainly from retinol rather than carotene) is associated with cardiometabolic risk (CMR) were considered. A higher odds ratio (OR) of suffering ≥ 2 simultaneous CMR factors was observed in men in the low consumption tercile of retinol (OR = 2.04; p = 0.019). In women, however, this relationship was not evident. Then, incubation of peripheral blood mononuclear cells (PBMCs) with VA-related compounds (ex vivo functional assay from 81 men and women) induced specific changes in the activity of genes involved in lipid homeostasis and inflammatory status, which were dependent on the type of compound tested and the sex of the person. In addition, the presence of the genetic variant rs5888 in SCARB1 was identified as having a high influence on VA-related metabolic response. The new evidence derived from this study could be relevant for personalized nutritional advice concerning VA and CMR.

The Metabolically Obese, Normal-Weight Phenotype in Young Rats Is Associated with Cognitive Impairment and Partially Preventable with Leptin Intake during Lactation.

The intake of high-fat diets (HFDs) and obesity are linked to cognitive impairment. Here, we aimed to investigate whether an early metabolically obese, normal-weight (MONW) phenotype, induced with an HFD in young rats, also leads to cognitive dysfunction and to evaluate the potential cognitive benefits of neonatal intake of leptin. To achieve this, Wistar rats orally received physiological doses of leptin or its vehicle during lactation, followed by 11 weeks of pair-feeding with an HFD or control diet post-weaning. Working memory was assessed using a T-maze, and gene expression in the hippocampus and peripheral blood mononuclear cells (PBMCs) was assessed with real-time RT-qPCR to identify cognition biomarkers. Young MONW-like rats showed hippocampal gene expression changes and decreased working memory. Animals receiving leptin during lactation presented similar gene expression changes but preserved working memory despite HFD intake, partly due to improved insulin sensitivity. Notably, PBMC Syn1 expression appears as an accessible biomarker of cognitive health, reflecting both the detrimental effect of HFD intake at early ages despite the absence of obesity and the positive effects of neonatal leptin treatment on cognition. Thus, the MONW phenotype developed at a young age is linked to cognitive dysfunction, which is reflected at the transcriptomic level in PBMCs. Neonatal leptin intake can partly counteract this impaired cognition resulting from early HFD consumption.

Leptin as a key regulator of the adipose organ.

Leptin is a hormone primarily produced by the adipose tissue in proportion to the size of fat stores, with a primary function in the control of lipid reserves. Besides adipose tissue, leptin is also produced by other tissues, such as the stomach, placenta, and mammary gland. Altogether, leptin exerts a broad spectrum of short, medium, and long-term regulatory actions at the central and peripheral levels, including metabolic programming effects that condition the proper development and function of the adipose organ, which are relevant for its main role in energy homeostasis. Comprehending how leptin regulates adipose tissue may provide important clues to understand the pathophysiology of obesity and related diseases, such as type 2 diabetes, as well as its prevention and treatment. This review focuses on the physiological and long-lasting regulatory effects of leptin on adipose tissue, the mechanisms and pathways involved, its main outcomes on whole-body physiological homeostasis, and its consequences on chronic diseases.

Lower miR-26a levels in breastmilk affect gene expression in adipose tissue of offspring.

Breastmilk miRNAs may act as epigenetic regulators of metabolism and energy homeostasis in offspring. Here, we aimed to investigate the regulatory effects of miR-26a on adipose tissue development. First, the 3T3-L1 cell model was used to identify putative target genes for miR-26a. Then, target genes were analysed in adipose tissue of offspring from dams that supplied lower levels of breastmilk miR-26a to determine whether miR-26a milk concentration might have a long-lasting impact on adipose tissue in the progeny. In the in vitro model, both over- and under-expression of miR-26a were induced by transfecting into 3T3-L1 with miR-26a mimic and inhibitor. Array analysis was performed after induction of miR-26a to ascertain the impact on mRNA target genes and influence of differentiation status. Focusing on genes related to adipose tissue development, transfection with miR-26a mimic reduced the expression of Pten, Hmga1, Stk11, Rb1, and Adam17 in both pre- and mature adipocytes. Data mostly confirmed the results found in the animal model. After weaning, descendants of cafeteria-fed dams breastfed with lower levels of miR-26a displayed greater expression of Hmag1, Rb1, and Adam17 in retroperitoneal white adipose tissue in comparison with controls. Hence, alterations in the amount of miR-26a supplied through milk during lactation is able to alter the expression of target genes in the descendants and may affect adipose tissue development. Thus, milk miR-26a may act as an epigenetic regulator influencing early metabolic program in the progeny, which emerges as a relevant component of an optimal milk composition for correct development.

Breast Milk MicroRNAs Related to Leptin and Adiponectin Function Can Be Modulated by Maternal Diet and Influence Offspring Phenotype in Rats.

There is evidence of the role of milk components in the metabolic programming of offspring. Here, we aimed to investigate the effects of a diet during lactation on breast milk leptin, adiponectin, and related miRNAs' expression, and their impact on dams and their offspring. Dams were fed a control diet (controls) or a diet enriched with oleic acid, betaine, and leucine (TX) throughout lactation. A TX diet promoted higher leptin at lactation day (LD) five and lower adiponectin on LD15 (vs. controls) in milk, resulting in increased leptin to adiponectin (L/A) ratio throughout lactation. Moreover, TX diet reduced milk levels of miR-27a, miR-103, miR-200a, and miR-222. Concerning TX offspring, higher body fat was early observed and maintained into adult life, accompanied by higher HOMA-IR than controls at three months of age. Offspring body fat content in adulthood correlated positively with milk L/A ratio at LD15 and negatively with miRNAs modulated by the TX diet. In conclusion, maternal diet during lactation can modulate leptin and adiponectin interplay with miRNAs in milk, setting up the metabolic programming of the offspring. Better knowledge about the influence of diet on this process is necessary to promote a healthy adult life in the progeny.

CUN-BAE Index as a Screening Tool to Identify Increased Metabolic Risk in Apparently Healthy Normal-Weight Adults and Those with Obesity.

BACKGROUND: Imbalanced dietary intake is related to increased adiposity, which is linked to increased metabolic risk even in the absence of obesity. BMI is traditionally used to classify body fatness and weight range, but it only considers body weight and height. The Clínica Universidad de Navarra-Body Adiposity Estimator (CUN-BAE) equation has appeared as an additional tool to estimate adiposity considering also other relevant parameters, i.e., sex and age. OBJECTIVES: We aimed to determine whether the CUN-BAE index could estimate adiposity-related metabolic risk in apparently healthy, normoglycemic adults. METHODS: In this case-control study, men and women (18-45 y old) were classified as normal-weight (NW) [n = 20; BMI (in kg/m2) <25] or overweight-obese (OW-OB) (n = 34; BMI ≥25). The primary outcome was body fat content and clinical circulating parameters to assess by correlation analysis CUN-BAE's usefulness as a predictor of metabolic risk. In addition, transcriptomic biomarkers of lipid metabolism were analyzed in peripheral blood mononuclear cells (PBMCs) as secondary outcome indicators of metabolic impairment. Data were analyzed by correlation analysis and comparison of means. RESULTS: CUN-BAE values correlated directly with body fatness obtained by DXA (r = 0.89, P < 0.01), with classical molecular biomarkers of metabolic risk, and with PBMC gene expression of carnitine palmitoyltransferase 1A (CPT1A), sterol regulatory element binding transcription factor 1c (SREBP-1c), and fatty acid synthase (FASN), early markers of metabolic impairment (P < 0.05). Moreover, CUN-BAE allowed identification of NW individuals with excessive body fatness, who were not yet presenting obesity-related molecular alterations. In these subjects, visceral fat correlated directly with circulating glucose, triglycerides, and total and LDL cholesterol, and with triglyceride-glucose and fatty liver indexes (P < 0.05). This is indicative of a metabolically obese NW phenotype. CONCLUSIONS: Data obtained in our cohort of young normoglycemic volunteers support the use of the CUN-BAE index as a tool to estimate accurately body fat mass, but also as a first easy/effective screening tool to identify lean people with increased fat mass and increased metabolic risk.This trial was registered at clinicaltrials.gov as NCT04402697.

Identification of blood cell transcriptome-based biomarkers in adulthood predictive of increased risk to develop metabolic disorders using early life intervention rat models.

Calorie restriction during gestation in rats has long-lasting adverse effects in the offspring. It induces metabolic syndrome-related alterations, which are partially reversed by leptin supplementation during lactation. We employed these conditions to identify transcript-based nutrient sensitive biomarkers in peripheral blood mononuclear cells (PBMCs) predictive of later adverse metabolic health. The best candidate was validated in humans. Transcriptome analysis of PBMCs from adult male Wistar rats of three experimental groups was performed: offspring of control dams (CON), and offspring of 20% calorie-restricted dams during gestation without (CR) and with leptin supplementation throughout lactation (CR-LEP). The expression of 401 genes was affected by gestational calorie restriction and reversed by leptin. The changes preceded metabolic syndrome-related phenotypic alterations. Of these genes, Npc1 mRNA levels were lower in CR vs CON, and normalized to CON in CR-LEP. In humans, NPC1 mRNA levels in peripheral blood cells (PBCs) were decreased in subjects with mildly impaired metabolic health compared to healthy subjects. Therefore, a set of potential transcript-based biomarkers indicative of a predisposition to metabolic syndrome-related alterations were identified, including NPC1, which was validated in humans. Low NPC1 transcript levels in PBCs are a candidate biomarker of increased risk for impaired metabolic health in humans.

Regulation of thermogenic capacity in brown and white adipocytes by the prebiotic high-esterified pectin and its postbiotic acetate.

OBJECTIVES: High-esterified pectin (HEP) is a prebiotic able to modulate gut microbiota, associated with health-promoting metabolic effects in glucose and lipid metabolism and adipostatic hormone sensitivity. Possible effects regulating adaptive thermogenesis and energy waste are poorly known. Therefore, we aimed to study how physiological supplementation with HEP is able to affect microbiota, energy metabolism and adaptive thermogenic capacity, and to contribute to the healthier phenotype promoted by HEP supplementation, as previously shown. We also attempted to decipher some of the mechanisms involved in the HEP effects, including in vitro experiments. SUBJECTS AND EXPERIMENTAL DESIGN: We used a model of metabolic malprogramming consisting of the progeny of rats with mild calorie restriction during pregnancy, both under control diet and an obesogenic (high-sucrose) diet, supplemented with HEP, combined with in vitro experiments in primary cultured brown and white adipocytes treated with the postbiotic acetate. RESULTS: Our main findings suggest that chronic HEP supplementation induces markers of brown and white adipose tissue thermogenic capacity, accompanied by a decrease in energy efficiency, and prevention of weight gain under an obesogenic diet. We also show that HEP promotes an increase in beneficial bacteria in the gut and peripheral levels of acetate. Moreover, in vitro acetate can improve adipokine production, and increase thermogenic capacity and browning in brown and white adipocytes, respectively, which could be part of the protection mechanism against excess weight gain observed in vivo. CONCLUSION: HEP and acetate stand out as prebiotic/postbiotic active compounds able to modulate both brown-adipocyte metabolism and browning and protect against obesity.

Alterations in plasma acylcarnitine and amino acid profiles may indicate poor nutrition during the suckling period due to maternal intake of an unbalanced diet and may predict later metabolic dysfunction.

Plasma profiles of acylcarnitines (ACs) and amino acids (AAs) may have interest as potential biomarkers. Here we analyzed plasma AC and AA profiles in 2 rat models with different metabolic programming outcomes: offspring of dams fed a cafeteria diet during lactation (O-CAF, with a thin-outside-fat-inside phenotype) and the offspring of dams with diet-induced obesity subjected to dietary normalization before gestation [offspring of postcafeteria dams (O-PCaf), nonaltered phenotype]. The purpose was to identify early variables that might indicate a propensity for a dysmetabolic state. O-CAF rats presented higher circulating levels of most of the lipid-derived ACs and higher hepatic expression of genes related to fatty acid oxidation ( Ppara and Cpt1a) than controls [offspring of control dams (O-C)]. They also exhibited an altered plasma AA profile. These differences were not observed in O-PCaf animals. A partial least squares-discriminant analysis score plot of the metabolomics data showed a clear separation between O-CAF and O-C animals. The long-chain ACs (C18, C18:1, C18:2, C16:1, and C16DC) and the AAs glycine, alanine, isoleucine, serine, and proline are the variables mainly influencing this separation. In summary, we have identified a cluster of ACs and AAs whose alterations may indicate poor nutrition during lactation due to maternal unbalanced diet intake and predict the later dysmetabolic phenotype observed in the offspring.-Pomar, C. A., Kuda, O., Kopecky, J., Rombaldova, M., Castro, H., Picó, C., Sánchez, J., Palou, A. Alterations in plasma acylcarnitine and amino acid profiles may indicate poor nutrition during the suckling period due to maternal intake of an unbalanced diet and may predict later metabolic dysfunction.

Metabolomic approach in milk from calorie-restricted rats during lactation: a potential link to the programming of a healthy phenotype in offspring.

PURPOSE: Mild/moderate maternal calorie restriction during lactation in rats has been associated with a lower predisposition to obesity and a healthier metabolic profile in adult offspring. Here, we aimed to assess the impact of maternal calorie restriction during lactation on milk composition to identify potential candidate components that could be involved in the programming effects in offspring. METHODS: An untargeted metabolomic approach in milk samples from 20%-calorie-restricted lactating (CRL) dams and their controls was performed. Levels of leptin, adiponectin, and irisin hormones in milk were also determined at lactating days 5, 10, and 15. RESULTS: Metabolomic analyses revealed a different metabolite pattern in milk between controls and CRL dams. 29 differential metabolites were tentatively identified (p < 0.05, FC > 1.5). Among them, myo-inositol, which showed greater levels in milk from CRL rats than controls, may be highlighted as one of the biologically plausible candidates that could be related to the beneficial effects of CRL in offspring. Results regarding myo-inositol were validated spectrophotometrically at days 10 and 15 of lactation, and levels in milk were correlated with maternal plasma levels. In addition, milk from CRL dams presented increased levels of adiponectin, decreased levels of irisin, and no changes in leptin levels vs controls throughout lactation. CONCLUSION: These data reveal important changes in milk composition due to calorie restriction during lactation that may be involved in the metabolic programming of the healthier phenotype of adult offspring. However, the possible contribution of the specific components is yet to be determined.

Blood cell transcript levels in 5-year-old children as potential markers of breastfeeding effects in those small for gestational age at birth.

BACKGROUND: Nutrition of the newborn during the early postnatal period seems to be of capital importance and there is clinical evidence showing the protective effect of breastfeeding compared with formula feeding on childhood obesity and its comorbidities. Infants born small for gestation age may be more sensitive to the type of feeding during lactation. Here, we aimed to analyze the impact of birth weight and the type of infant feeding on the expression levels in peripheral blood cells of selected candidate genes involved in energy homeostasis in 5-year-old children, to find out potential early biomarkers of metabolic programming effects during this period of metabolic plasticity. METHODS: Forty subjects were recruited at birth and divided in four groups according to birth weight (adequate or small for gestational age) and type of infant feeding (breastfeeding or formula feeding). They were followed from birth to the age of 5 years. RESULTS: At 5 years, no significant differences regarding anthropometric parameters were found between groups, and all children had normal biochemical values. Expression levels of UCP2 and MC4R in peripheral blood cells were lower and higher, respectively, in formula feeding children compared with breastfeeding ones (P = 0.002 and P = 0.064, two-way ANOVA). Differences were more marked and significant by Student's t test in small for gestation age children (P < 0.001 and P = 0.017, respectively). Transcript levels of FASN and FTO in peripheral blood cells were also different according to the type of infant feeding, but only in small for gestation age children. CONCLUSIONS: Altogether, these results suggest that small for gestation age infants are more sensitive to the type of feeding during lactation, and transcript levels of particular genes in peripheral blood cells, especially the MC4R/UCP2 mRNA ratio, may precisely reflect these effects in the absence of clear differences in phenotypic traits.

Novel Markers of the Metabolic Impact of Exogenous Retinoic Acid with A Focus on Acylcarnitines and Amino Acids.

Treatment with all-trans retinoic acid (ATRA), the carboxylic form of vitamin A, lowers body weight in rodents by promoting oxidative metabolism in multiple tissues including white and brown adipose tissues. We aimed to identify novel markers of the metabolic impact of ATRA through targeted blood metabolomics analyses, with a focus on acylcarnitines and amino acids. Blood was obtained from mice treated with a high ATRA dose (50 mg/kg body weight/day, subcutaneous injection) or placebo (controls) during the 4 days preceding collection. LC-MS/MS analyses with a focus on acylcarnitines and amino acids were conducted on plasma and PBMC. Main results showed that, relative to controls, ATRA-treated mice had in plasma: increased levels of carnitine, acetylcarnitine, and longer acylcarnitine species; decreased levels of citrulline, and increased global arginine bioavailability ratio for nitric oxide synthesis; increased levels of creatine, taurine and docosahexaenoic acid; and a decreased n-6/n-3 polyunsaturated fatty acids ratio. While some of these features likely reflect the stimulation of lipid mobilization and oxidation promoted by ATRA treatment systemically, other may also play a causal role underlying ATRA actions. The results connect ATRA to specific nutrition-modulated biochemical pathways, and suggest novel mechanisms of action of vitamin A-derived retinoic acid on metabolic health.

Retinoic Acid Increases Fatty Acid Oxidation and Irisin Expression in Skeletal Muscle Cells and Impacts Irisin In Vivo.

BACKGROUND/AIMS: All-trans retinoic acid (ATRA) has protective effects against obesity and metabolic syndrome. We here aimed to gain further insight into the interaction of ATRA with skeletal muscle metabolism and secretory activity as important players in metabolic health. METHODS: Cultured murine C2C12 myocytes were used to study direct effects of ATRA on cellular fatty acid oxidation (FAO) rate (using radioactively-labelled palmitate), glucose uptake (using radioactively-labelled 2-deoxy-D-glucose), triacylglycerol levels (by an enzymatic method), and the expression of genes related to FAO and glucose utilization (by RT-real time PCR). We also studied selected myokine production (using ELISA and immunohistochemistry) in ATRA-treated myocytes and intact mice. RESULTS: Exposure of C2C12 myocytes to ATRA led to increased fatty acid consumption and decreased cellular triacylglycerol levels without affecting glucose uptake, and induced the expression of the myokine irisin at the mRNA and secreted protein level in a dose-response manner. ATRA stimulatory effects on FAO-related genes and the Fndc5 gene (encoding irisin) were reproduced by agonists of peroxisome proliferator-activated receptor ß/δ and retinoid X receptors, but not of retinoic acid receptors, and were partially blocked by an AMP-dependent protein kinase inhibitor. Circulating irisin levels were increased by 5-fold in ATRA-treated mice, linked to increased Fndc5 transcription in liver and adipose tissues, rather than skeletal muscle. Immunohistochemistry analysis of FNDC5 suggested that ATRA treatment enhances the release of FNDC5/irisin from skeletal muscle and the liver and its accumulation in interscapular brown and inguinal white adipose depots. CONCLUSION: These results provide new mechanistic insights on how ATRA globally stimulates FAO and enhances irisin secretion, thereby contributing to leaning effects and improved metabolic status.

Peripheral Blood Cells, a Transcriptomic Tool in Nutrigenomic and Obesity Studies: Current State of the Art.

Gene expression profile of peripheral blood cells (PBC) is able to reflect useful aspects of the whole body metabolic status. Therefore, and favored by the huge development of "omic" technologies, blood cells and, particularly, the peripheral blood mononuclear cell (PBMC) fraction, are emerging as a potent source of transcriptomic biomarkers of health and disease. In this review we describe and discuss the available information concerning the use of the PBC and the PBMC fraction as a crucial tool for nutrigenomic studies. Results of these studies reveal, as these cells are good indicators of metabolic adaptations to diet and, moreover, as they allow us to monitor from early stages on, the metabolic alterations associated with dietary imbalances. In this way, blood cells present the capacity of reflecting higher risks of suffering from diet-related pathologies, such as obesity and its medical complications. What is more, different studies also show how PBMC are able to evidence the metabolic recovery associated with weight loss or dietary interventions. Besides, recent research points to the utility of ex vivo systems of blood cells to test the efficacy of food bioactives. All in all, PBC constitutes an easily obtainable source of predictive biomarkers of metabolic imbalance and disease related to diet and obesity, and also of metabolic recovery, which appears as highly relevant for developing nutritional preventive strategies in dietetics. Moreover, they could serve to perform relatively simple and economic in vitro tests to assess food bioactive compounds, promoting in this way functional food research and related industry developments.

Consumption of a Mango Fruit Powder Protects Mice from High-Fat Induced Insulin Resistance and Hepatic Fat Accumulation.

BACKGROUND/AIMS: The aim of this study was to gain more insight into the beneficial effects of mango fruit powder on the early metabolic adverse effects of a high-fat diet. METHODS: The progressive dose-response effects of mango fruit powder on body composition, circulating parameters, and the expression of genes related to fatty acid oxidation and insulin sensitivity in key tissues were studied in mice fed a moderate (45%) high-fat diet. RESULTS: Findings suggest that mango fruit powder exerts physiological protective effects in the initial steps of insulin resistance and hepatic lipid accumulation induced by a high-fat diet in mice. Moreover, AMPK and SIRT1 appear as key regulators of the observed improvement in fatty acid oxidation capacity, as well as of the improved insulin sensitivity and the increased glucose uptake and metabolism through the glycolytic pathway capacity in liver and skeletal muscle. CONCLUSION: In summary, this study provides evidence that the functional food ingredient (CarelessTM) from mango fruit prevents early metabolic alterations caused by a high-fat diet in the initial stages of the metabolic syndrome.

Methylation analysis in fatty-acid-related genes reveals their plasticity associated with conjugated linoleic acid and calcium supplementation in adult mice.

PURPOSE: DNA methylation is one of the most extensively studied mechanisms within epigenetics, and it is suggested that diet-induced changes in methylation status could be involved in energy metabolism regulation. Conjugated linoleic acid (CLA) and calcium supplementation counteract body weight gain, particularly under a high-fat (HF) diet, in adult mice. The aim was to determine whether the modulation of DNA methylation pattern in target genes and tissues could be an underlying mechanism of action. METHODS: Mice (C57BL/6J) were divided into five groups according to diet and treatment: normal fat as the control group (12 % kJ content as fat), HF group (43 % kJ content as fat), HF + CLA (6 mg CLA/day), HF + calcium (12 g/kg of calcium) and HF with both compounds. Gene expression and methylation degree of CpG sites in promoter sequences of genes involved in fatty acid metabolism, including adiponectin (Adipoq), stearoyl-CoA desaturase (Scd1) and fatty acid synthase (Fasn), were determined by bisulphite sequencing in liver and epididymal white adipose tissue. RESULTS: Results showed that the methylation profile of promoters was significantly altered by dietary supplementation in a gene- and tissue-specific manner, whereas only slight changes were observed in the HF group. Furthermore, changes in specific CpG sites were also associated with an overall healthier metabolic profile, in particular for calcium-receiving groups. CONCLUSIONS: Both CLA and calcium were able to modify the methylation pattern of genes involved in energy balance in adulthood, which opens a novel area for increasing efficiency in body weight management strategies.

Carotenoids in Adipose Tissue Biology and Obesity.

Cell, animal and human studies dealing with carotenoids and carotenoid derivatives as nutritional regulators of adipose tissue biology with implications for the etiology and management of obesity and obesity-related metabolic diseases are reviewed. Most studied carotenoids in this context are ß-carotene, cryptoxanthin, astaxanthin and fucoxanthin, together with ß-carotene-derived retinoids and some other apocarotenoids. Studies indicate an impact of these compounds on essential aspects of adipose tissue biology including the control of adipocyte differentiation (adipogenesis), adipocyte metabolism, oxidative stress and the production of adipose tissue-derived regulatory signals and inflammatory mediators. Specific carotenoids and carotenoid derivatives restrain adipogenesis and adipocyte hypertrophy while enhancing fat oxidation and energy dissipation in brown and white adipocytes, and counteract obesity in animal models. Intake, blood levels and adipocyte content of carotenoids are reduced in human obesity. Specifically designed human intervention studies in the field, though still sparse, indicate a beneficial effect of carotenoid supplementation in the accrual of abdominal adiposity. In summary, studies support a role of specific carotenoids and carotenoid derivatives in the prevention of excess adiposity, and suggest that carotenoid requirements may be dependent on body composition.

Cell-Autonomous Brown-Like Adipogenesis of Preadipocytes From Retinoblastoma Haploinsufficient Mice.

Mechanisms behind the emergence of brown adipocyte-like (brite or beige) adipocytes within white adipose tissue (WAT) are of interest. Retinoblastoma protein gene (Rb) haploinsufficiency associates in mice with improved metabolic regulation linked to a greater capacity for fatty acid oxidation and thermogenesis in WAT. We aimed to explain a feasible mechanism of WAT-to-BAT remodeling in this model. Differentiated primary adipocytes and Sca1-positive preadipocytes derived from adipose depots of Rb(+/-) mice and wild-type siblings were compared. Primary white Rb(+/-) adipocytes displayed under basal conditions increased glucose uptake and an enhanced expression of brown adipocyte-related genes (Pparg, Ppargc1a, Ppargc1b, Prdm16, Cpt1b) but not of purported beige/brite transcriptional markers (Cd137, Tmem26, Tbx1, Slc27a1, Hoxc9, Shox2). Lack of induction of beige markers phenocopied results in WAT of adult Rb(+/-) mice. Flow cytometry analysis evidenced an increased number of preadipocytes in WAT depots of Rb(+/-) mice. Sca1(+) preadipocytes from WAT of Rb(+/-) mice displayed increased gene expression of several transcription factors common to the brown and beige adipogenic programs (Prdm16, Pparg, Ppargc1a) and of receptors of bone morphogenetic proteins (BMPs); however, among the recently proposed beige markers, only Tbx1 was upregulated. Adult Rb(+/-) mice had increased circulating levels of BMP7. These results indicate that preadipose cells resident in WAT depots of Rb(+/-) mice retain an increased capacity for brown-like adipogenesis that appears to be different from beige adipogenesis, and suggest that the contribution of these precursors to the Rb(+/-) adipose phenotype is driven, at least in part, by interaction with BMP7 pathways. J. Cell. Physiol. 231: 1941-1952, 2016. © 2016 Wiley Periodicals, Inc.

Leptin Effect on Acetylation and Phosphorylation of Pgc1α in Muscle Cells Associated With Ampk and Akt Activation in High-Glucose Medium.

Leptin is crucial in energy metabolism, including muscle regulation. Peroxisome proliferator activated receptor gamma co-activator 1α (PGC1α) orchestrates energy metabolism and is tightly controlled by post-translational covalent modifications such as phosphorylation and acetylation. We aimed to further the knowledge of PGC1α control by leptin (at physiological levels) in muscle cells by time-sequentially analysing the activation of AMP activated protein kinase (AMPK), P38 mitogen-activated protein kinase (P38 MAPK) and Akt (Protein kinase B)--all known to phosphorylate PGC1α and to be involved in the regulation of its acetylation status--in C2C12 myotubes placed in a high-glucose serum-free medium. We also studied the protein levels of PGC1α, Sirtuin 1, adiponectin, COX IV, mitofusin 2 (Mfn2), and pyruvate dehydrogenase kinase 4 (PDK4). Our main findings suggest an important role of leptin regulating AMPK and Akt phosphorylation, Mfn2 induction and PGC1α acetylation status, with the novelty that the latter in transitorily increased in response to leptin, an effect dependent, at least in part, on AMPK regulation. These post-translational reversible changes in PGC1α in response to leptin, especially the increase in acetylation status, may be related to the physiological role of the hormone in modulating muscle cell response to the physiological/nutritional status.

Retinoblastoma Protein Knockdown Favors Oxidative Metabolism and Glucose and Fatty Acid Disposal in Muscle Cells.

Deficiency in the retinoblastoma protein (Rb) favors leanness and a healthy metabolic profile in mice largely attributed to activation of oxidative metabolism in white and brown adipose tissues. Less is known about Rb modulation of skeletal muscle metabolism. This was studied here by transiently knocking down Rb expression in differentiated C2C12 myotubes using small interfering RNAs. Compared with control cells transfected with non-targeting RNAs, myotubes silenced for Rb (by 80-90%) had increased expression of genes related to fatty acid uptake and oxidation such as Cd36 and Cpt1b (by 61% and 42%, respectively), increased Mitofusin 2 protein content (∼2.5-fold increase), increased mitochondrial to nuclear DNA ratio (by 48%), increased oxygen consumption (by 65%) and decreased intracellular lipid accumulation. Rb silenced myotubes also displayed up-regulated levels of glucose transporter type 4 expression (∼5-fold increase), increased basal glucose uptake, and enhanced insulin-induced Akt phosphorylation. Interestingly, exercise in mice led to increased Rb phosphorylation (inactivation) in skeletal muscle as evidenced by immunohistochemistry analysis. In conclusion, the silencing of Rb enhances mitochondrial oxidative metabolism and fatty acid and glucose disposal in skeletal myotubes, and changes in Rb status may contribute to muscle physiological adaptation to exercise.