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International audience; Purpose Protein synthesis and proteolysis are known to be controlled through mammalian target of rapamycin, AMP-activated kinase (AMPK) and general control non-derepressible 2 (GCN2) pathways, depending on the nutritional condition. This study aimed at investigating the contribution of liver AMPK and GCN2 on the adaptation to high variations in protein intake. Methods To evaluate the answer of protein pathways to high- or low-protein diet, male wild-type mice and genetically modified mice from C57BL/6 background with liver-specific AMPK- or GCN2-knockout were fed from day 25 diets differing in their protein level as energy: LP (5%), NP (14%) and HP (54%). Two hours after a 1 g test meal, protein synthesis rate was measured after a C-13 valine flooding dose. The gene expression of key enzymes involved in proteolysis and GNC2 signaling pathway were quantified. Results The HP diet but not the LP diet was associated with a decrease in fractional synthesis rate by 29% in the liver compared to NP diet. The expression of mRNA encoding ubiquitin and Cathepsin D was not sensitive to the protein content. The deletion of AMPK or GCN2 in the liver did not affect nor protein synthesis rates and neither proteolysis markers in the liver or in the muscle, whatever the protein intake. In the postprandial state, protein level alters protein synthesis in the liver but not in the muscle. Conclusions Taken together, these results suggest that liver AMPK and GCN2 are not involved in this adaptation to high- and low-protein diet observed in the postprandial period.

International audience; Amino acids are involved in energy homeostasis, just as are carbohydrates and lipids. Therefore, mechanisms controlling protein intake should operate independently and in combination with systems controlling overall energy intake to coordinate appropriate metabolic and behavioural responses. The objective of this study was to quantify the respective roles of dietary protein and carbohydrate levels on energy balance, plasma FGF21 and IGF-1 concentrations, and hypothalamic neurotransmitters (POMC, NPY, AgRP and CART). In a simplified geometric framework, 7-week-old male Wistar rats were fed 12 diets containing 3 to 30% protein for 3 weeks, in which carbohydrates accounted for 30 to 75% of the carbohydrate and fat part of the diet. As a result of this study, most of the studied parameters (body composition, energy expenditure, plasma FGF21 and IGF-1 concentrations and Pomc/Agrp ratio) responded mainly to the protein content and to a lesser extent to the carbohydrate content in the diet.

BACKGROUND Under dietary self-selection (DSS), rats ingest 25-30% of energy as protein. This high level appears to be explained by metabolic benefits related to reduced carbohydrate dependence and associated pathologies. However, the mechanisms underlying these choices remain largely misunderstood. OBJECTIVES The aim was to test the hypothesis that in a DSS model, rats select a protein-to-energy (PE) ratio to maintain the protein-to-carbohydrate (PC) ratio constant and that fibroblast growth factor 21 (FGF21) is involved in this response. METHODS Adult male Wistar rats were used in 3 experiments. The first was to determine whether the PE ratio was influenced by changes in carbohydrate content. The second was to test whether the PE ratio was defended with a modified DSS model. The third was to determine whether the selected PE ratio was of metabolic interest compared with a standard 15% protein diet. Food intake, body weight, and energy expenditure were measured. After 3 wk, plasma was sampled and rats were killed to determine body composition and gene expression. Statistical analyses were mainly done by ANOVA tests and correlation tests. RESULTS The selected PE ratio increased from 20% to 35% when the carbohydrate content of the protein-free diet increased from 30% to 75% (R2 = 0.56; P

International audience; The protein requirement is generally defined as the amount necessary to maintain the body's protein pool. However, under free choice conditions, animal models often ingest more protein than required for nitrogen balance (10 %–15 %). This behavior possibly reflects the search for a high protein-to-carbohydrate ratio (0.6–0.8), inducing metabolic benefits. This indicates that in addition to protein homeostasis, dietary proteins are also involved in energy homeostasis. The mechanisms controlling protein and energy intake are partly independent and in specific conditions, there may be a conflict between the two. Protein density in the human diet has decreased ∼ 2 % since the 1970s and, according to the protein leverage hypothesis, this decrease may be involved in the increase in energy intake and prevalence of obesity observed today.; Le besoin en protéines est en général défini comme la quantité nécessaire pour le maintien du pool de protéines corporelles, or dans des conditions de libre choix, les modèles animaux ingèrent souvent plus de protéines que nécessaire pour l’équilibre azoté (10 %–15 %). Ce comportement semble refléter la recherche d’un rapport protéines/glucides élevé (0,6–0,8), induisant des bénéfices métaboliques. Cela indique qu’en plus de l’homéostasie des protéines, les protéines alimentaires sont impliquées dans l’homéostasie énergétique. Les mécanismes contrôlant l’apport protéique et énergétique sont en partie indépendants et dans des conditions spécifiques, il peut y avoir un conflit entre les deux. La densité des protéines dans l’alimentation humaine a diminué d’environ 2 % depuis les années 1970 et, selon l’hypothèse de l’effet de levier des protéines, cette diminution pourrait contribuer à l’augmentation de l’apport énergétique et de la prévalence de l’obésité observées aujourd’hui.

Protein requirement has been determined at 10%-15% energy. Under dietary self-selection, rats ingest 25%-30% energy as protein and regulate FGF21 (a hormone signaling protein deficiency) to levels lower than those measured with a 15% protein (15P) diet. Our hypothesis is that if a 15P diet was indeed sufficient to ensure protein homeostasis, it is probably a too low protein level to ensure optimal energy homeostasis. Adult male Wistar rats were used in this study. The first objective was to determine the changes in food intake, body composition, and plasma FGF21, IGF-1, and PYY concentrations in rats fed 8P, 15P, 30P, 40P, or 50P diets. The second was to determine whether the FGF21 levels measured in the rats were related to spontaneous protein intake. Rats were fed a 15P diet and then allowed to choose between a protein diet and a protein-free diet. Food intake and body weight were measured throughout the experiments. Body composition was determined at different experimental stages. Plasma samples were collected to measure FGF21, IGF-1, and PYY concentrations. A 15P diet appears to result in higher growth than that observed with the 30P, 40P, and 50P diets. However, the 15P diet probably does not provide optimal progression of body composition owing to a tendency of 15P rats to fix more fat and energy in the body. The variable and higher concentrations of FGF21 in the 15P diet suggest a deficit in protein intake, but this does not appear to be a parameter reflecting the adequacy of protein intake relative to individual protein requirements.

The objective of this study is to evaluate the effects of a strictly essential amino acid (lysine or threonine; EAA) deficiency on energy metabolism in growing rats. Rats were fed for three weeks severely (15% and 25% of recommendation), moderately (40% and 60%), and adequate (75% and 100%) lysine or threonine-deficient diets. Food intake and body weight were measured daily and indirect calorimetry was performed the week three. At the end of the experimentation, body composition, gene expression, and biochemical analysis were performed. Lysine and threonine deficiency induced a lower body weight gain and an increase in relative food intake. Lysine or threonine deficiency induced liver FGF21 synthesis and plasma release. However, no changes in energy expenditure were observed for lysine deficiency, unlike threonine deficiency, which leads to a decrease in total and resting energy expenditure. Interestingly, threonine severe deficiency, but not lysine deficiency, increase orexigenic and decreases anorexigenic hypothalamic neuropeptides expression, which could explain the higher food intake. Our results show that the deficiency in one EAA, induces a decrease in body weight gain, despite an increased relative food intake, without any increase in energy expenditure despite an induction of FGF21.

International audience; Objectives The availability of indispensable amino acids (IAA) modulates protein turnover. More particularly AAI deficiency reduces protein synthesis while the consequence on proteolysis remains unclear. The present study aims to evaluate the specific response of both protein synthesis and proteolysis to a diet restricted on one strictly indispensable IAA, either lysine or threonine Methods Sixty-four growing rats were divided into 8 groups (n = 8/group). They were fed for 3 weeks isocaloric diets composed with different levels of lysine or threonine (L or T), 15, 25, 40, 60, 75, 100 or 170% of the theoretical lysine/threonine requirements. At the end of the experiment, rats were injected with valine13C and tissues and biological fluids were collected for gene expression measurement and blood amino acids (AA). Protein synthesis rate (Fractional and Absolute rate synthesis, ie FSR, ASR) were determined in liver and muscle. Statistical analysis was done by 1- or 2-factor ANOVA, when data were repeated. Results Severe (L/T15, L/T25) and moderate (T40) lysine or threonine deficiency resulted in a decrease in body weight gain due to a decrease in lean body mass. Severe restriction (L15, T15, T25) decreased the muscle FSR whereas no effect was observed in the liver. When the rate of protein synthesis was expressed per tissue, the ASR was decreased by severe restriction of lysine and threonine in liver and muscle and by moderate threonine deficiency (T40, T60, T75) in muscle. In liver, no effect of lysine and threonine on proteolysis was observed. In muscle, only severe lysine (L15) deficiency increased proteolysis. Dietary lysine deficiency induced a decrease in lysine concentration in the portal vein and in the vena cava whereas for threonine deficiency, all AAIs except threonine were decreased in the portal vein and vena cava. Conclusions These results indicate that the decreased protein synthesis is the primary mechanism involved in decreased lean body mass in response to the severe deficiency in a single AAI. Deficiency of a single AAI reproduce the effect of the low protein diet on protein synthesis. Lysine and threonine deficiency differently affect for a part protein turnover probably in relation with the tissue where they are metabolized. Funding Sources This study was funded by the doctoral school ABIES and AlimH-INRAE department.

Omnivores are able to correctly select adequate amounts of macronutrients from natural foods as well as purified macronutrients. In the rat model, the selected protein levels are often well above the requirements estimated from the nitrogen balance. These high intake levels were initially interpreted as reflecting poor control of protein intake, but the selected levels were later found to be precisely controlled for changes in dietary protein quality and adjusted for cold, exercise, pregnancy, lactation, age, etc. and therefore met physiological requirements. Several authors have also suggested that instead of a given level of protein intake, rodents regulate a ratio of protein to dietary carbohydrates in order to achieve metabolic benefits such as reduced insulin levels, improved blood glucose control, and, in the long term, reduced weight and fat gain. The objective of this review was to analyze the most significant results of studies carried out on rats and mice since the beginning of the 20th century, to consider what these results can bring us to interpret the current causes of the obesity pandemic and to anticipate the possible consequences of policies aimed at reducing the contribution of animal proteins in the human diet.

International audience; The protein requirement is generally defined as the amount necessary to maintain the body's protein pool. However, under free choice conditions, animal models often ingest more protein than required for nitrogen balance (10 %–15 %). This behavior possibly reflects the search for a high protein-to-carbohydrate ratio (0.6–0.8), inducing metabolic benefits. This indicates that in addition to protein homeostasis, dietary proteins are also involved in energy homeostasis. The mechanisms controlling protein and energy intake are partly independent and in specific conditions, there may be a conflict between the two. Protein density in the human diet has decreased ∼ 2 % since the 1970s and, according to the protein leverage hypothesis, this decrease may be involved in the increase in energy intake and prevalence of obesity observed today.; Le besoin en protéines est en général défini comme la quantité nécessaire pour le maintien du pool de protéines corporelles, or dans des conditions de libre choix, les modèles animaux ingèrent souvent plus de protéines que nécessaire pour l’équilibre azoté (10 %–15 %). Ce comportement semble refléter la recherche d’un rapport protéines/glucides élevé (0,6–0,8), induisant des bénéfices métaboliques. Cela indique qu’en plus de l’homéostasie des protéines, les protéines alimentaires sont impliquées dans l’homéostasie énergétique. Les mécanismes contrôlant l’apport protéique et énergétique sont en partie indépendants et dans des conditions spécifiques, il peut y avoir un conflit entre les deux. La densité des protéines dans l’alimentation humaine a diminué d’environ 2 % depuis les années 1970 et, selon l’hypothèse de l’effet de levier des protéines, cette diminution pourrait contribuer à l’augmentation de l’apport énergétique et de la prévalence de l’obésité observées aujourd’hui.

Fibroblast growth factor 21 (FGF21) is a hepatokine member of a subfamily of “fibroblast growth factors” that responds to multiple...

To study, in young growing rats, the consequences of different levels of dietary protein deficiency on food intake, body weight, body composition, and energy balance and to assess the role of FGF21 in the adaptation to a low protein diet. Thirty-six weanling rats were fed diets containing 3%, 5%, 8%, 12%, 15% and 20% protein for three weeks. Body weight, food intake, energy expenditure and metabolic parameters were followed throughout this period. The very low-protein diets (3% and 5%) induced a large decrease in body weight gain and an increase in energy intake relative to body mass. No gain in fat mass was observed because energy expenditure increased in proportion to energy intake. As expected, Fgf21 expression in the liver and plasma FGF21 increased with low-protein diets, but Fgf21 expression in the hypothalamus decreased. Under low protein diets (3% and 5%), the increase in liver Fgf21 and the decrease of Fgf21 in the hypothalamus induced an increase in energy expenditure and the decrease in the satiety signal responsible for hyperphagia. Our results highlight that when dietary protein decreases below 8%, the liver detects the low protein diet and responds by activating synthesis and secretion of FGF21 in order to activate an endocrine signal that induces metabolic adaptation. The hypothalamus, in comparison, responds to protein deficiency when dietary protein decreases below 5%.

The worldwide obesity and type 2 diabetes epidemics have led to an increase in nonalcoholic fatty liver disease (NAFLD). NAFLD covers a spectrum of hepatic pathologies ranging from simple steatosis to nonalcoholic steatohepatitis, characterized by fibrosis and hepatic inflammation. Nonalcoholic steatohepatitis predisposes to the onset of hepatocellular carcinoma (HCC). Here, we characterized the effect of a pharmacological activator of the intracellular energy sensor adenosine monophosphate–activated protein kinase (AMPK) on NAFLD progression in a mouse model. The compound stimulated fat oxidation by activating AMPK in both liver and skeletal muscle, as revealed by indirect calorimetry. This translated into an ameliorated hepatic steatosis and reduced fibrosis progression in mice fed a diet high in fat, cholesterol, and fructose for 20 weeks. Feeding mice this diet for 80 weeks caused the onset of HCC. The administration of the AMPK activator for 12 weeks significantly reduced tumor incidence and size. Conclusion: Pharmacological activation of AMPK reduces NAFLD progression to HCC in preclinical models., A pharmacological activator of the cellular energy sensor AMP‐activated protein kinase was tested in mice with diet‐induced, biopsy‐proven NAFLD. The compound reduced fibrosis progression and the onset of hepatocellular carcinoma.

OBJECTIVES: In mice, low protein (LP) diets increase food intake (FI) thereby energy intake, to compensate for protein deficiency, but mice do not gain fat because total energy expenditure (TEE) is also increased. Fibroblast Growth Factor 21 (FGF21), which is expressed in the liver and in the brain, appears as a key player in these effects. The present study hypothesized that both LP diet but also only lysine or threonine deficiency can modulate FGF21 in the liver and in the hypothalamus that in turn is involved in the control of FI and energy expenditure. METHODS: Growing rats were fed for 3 weeks: i) LP diets containing 3-5-8-12-15 or 20% of milk protein, or ii) the same diets but supplemented with free amino acids at the level of the 20% protein diet except for lysine or threonine leading to lysine or threonine deficient diets. Body weight and FI were measured daily and energy expenditure were measured by indirect calorimetry. At the end of the experiment, rats were euthanized, tissues and biological fluids were removed and frozen, and body composition as well as gene expression and plasma FGF21 were analyzed. RESULTS: Diets with 3 and 5% protein, and diets highly deficient in lysine or threonine (85% and 75%) result in significant growth retardation. LP 3% and 5% induced an increase in relative FI. Surprisingly, an increase in TEE was observed under LP 5% protein, due to an increase in motor activity. Hepatic FGF21 expression is increased at 3 and 5% and strongly decreased at 12, 15 and 20% protein. In contrast, in the hypothalamus, FGF21 expression was significantly lower in LP 3% compared to a 20% protein diet, and for the other LP diets, the values are intermediate. Plasma FGF21 was higher in 3, 5, 8 and 12% protein than in 15 and 20% protein diets. Lysine or threonine deficiency were able to reproduce the effect of LP diet at 3 and 5% whereas at 8%, only the deficiency of threonine was able to reproduce the LP effect. CONCLUSIONS: These results showed that hepatic and hypothalamic expression FGF21 are inversely affected by protein deficiency. Such situations of deficiency induced an up-regulation of hepatic expression of FGF21 that increased TEE and a down-regulation of hypothalamic expression of FGF21 that could led to hyperphagia. Interestingly, the levels of FGF21 observed for the 3, 5, 8% protein diets could be due at least to lysine and threonine deficiency. FUNDING SOURCES: ABIES, AlimH department of INRAe.

OBJECTIVES: Fibroblast Growth Factor 21 (FGF21), a response to metabolic stress, is influenced by the dietary protein content. Previous studies have shown that a protein level below 10% of energy increases FGF21 hepatic secretion in mice, and increases food intake and energy expenditure. However, it has also been shown in vitro that glucose stimulates FGF21 secretion in liver. The objective of this study was to determine the respective roles of dietary protein and carbohydrate contents on FGF21 secretion and associated metabolic responses. METHODS: 70 male Wistar rats were subjected at one to 12 diets with various milk protein contents (3, 5, 8, 15 and 30% P of energy) and a mixture of carbohydrates and fats in which carbohydrates amounted 30, 45, 60 or 75% of energy. Body weight and energy intake were measured twice a week, and energy expenditure was measured one time after 2 weeks by indirect calorimetry. After 3 weeks, plasma was collected and an ELISA test was used to determine plasma concentration of FGF21. Tissues were dissected and weighed to determine body composition. Pieces of liver were frozen for measuring expression of Fgf21 mRNA by RT-PCR. Statistical analyzes were done by analysis of variance. RESULTS: The decrease in %P in diets increased liver Fgf21 mRNA. Using the 30% P fed group as a reference, Fgf21 mRNA were increased not significantly 3x in 15% P, but significantly 19x in 8% P, 44x in 5% P and 60x in 3% P (P

Introduction et but de l’etude Differentes etudes ont rapporte qu’une restriction proteique impacte le metabolisme energetique de rats en croissance. Une deficience severe en proteine (3 % ou 5 % du besoin) induit notamment une diminution du gain de poids corporel, une augmentation de la prise alimentaire relative, de la depense energetique, ainsi que des taux de fibroblast growth factor 21 (FGF21) plasmatiques. Cependant, l’implication precise de tous ou certains acides amines (AA) dans ces effets reste inconnue. Cette etude a pour objectif d’evaluer les effets d’une deficience en un AA strictement indispensable (lysine ou threonine ; AAI) sur le metabolisme energetique de rats en croissance. Materiel et methodes Des rats en croissance ont ete nourris avec des regimes deficients en lysine ou threonine pendant trois semaines : severement deficients (P3 L/T15 et P5 L/T25), moderement deficients (P8 L/T40 et P12 L/T60), controles (P15 L/T75 et P20 L/T100), excessif (P20 L/T170) ou un controle de l’effet des AA libres dans le regime (P3 L/T100). Le poids corporel et la prise alimentaire ont ete mesures quotidiennement, la depense energetique a ete evaluee par calorimetrie indirecte durant la troisieme semaine. A la fin de l’experimentation, les tissus et fluides biologiques ont ete preleves et la composition corporelle a ete analysee. L’analyse statistique a ete faite par modele mixte en mesure repetee et ANOVA a 1 facteur. Resultats et analyse statistique Les regimes severement deficients en lysine induisent une reduction du gain de poids corporel, des 3 et 6 jours (pour P3 L15 et P5 L25 respectivement). Pour la threonine, les regimes severement deficients, ainsi que le regime P8 T40, provoquent une diminution du gain de poids corporel des 1 et 6 jours. Parallelement, on observe une augmentation de la prise alimentaire relative (PAR) pour les regimes severement deficients en lysine et threonine, ainsi que pour les regimes P8 L40/P8 T40. Les regimes deficients en lysine n’influent pas sur la depense energetique de repos (DER) et le cout d’activite, et ne sont pas significativement differents des regimes controles pour la depense energetique totale (DET) et d’activite (DEA). Pour la deficience en threonine, on observe une diminution de la DET pour le regime P3 T15, confortee par une diminution de la DER, sans difference significative de la DEA par rapport aux regimes controles. De plus, le cout d’activite tend a etre diminue pour le regime P3 T15. Les deficiences severes en lysine ou threonine induisent une augmentation de FGF21 plasmatique. Conclusion La deficience en un AAI, lysine ou threonine, induit une diminution du gain de poids corporel chez le rat en croissance, malgre une PAR augmentee, avec un effet plus marque dans le cas de la threonine. Comparativement aux regimes controles, la deficience en lysine n’impacte pas la depense energetique, contrairement a la deficience en threonine qui entraine une diminution de la DET et de la DER.

International audience; Protein sufficiency is tightly controlled through different sensing and signaling processes that modulate and adapt protein and energy metabolism and feeding behavior to reach and maintain a well-balanced protein status. High-protein diets, often discussed in the context of body weight management, usually activate anorexigenic pathways, leading to higher satiety, decreased food and energy intake, and decreased body weight and adiposity. Diets marginally low in protein (3-8% energy) or marginally deficient in some indispensable amino acid more often activate orexigenic pathways, with higher appetite and a specific appetite for protein, a response that leads to an increase in protein intake to partially compensate for the deficit in protein and amino acid. Diets severely deficient in protein (2-3% energy as protein) usually depress food intake and induce lower weight and lower fat mass and lean tissues that characterize a status of protein deficiency. The control of protein sufficiency involves various peripheral and central signals, including modulation of both metabolic pathways at the periphery as well as central pathways of the control of food and protein intake, including a reward-driven specific sensitivity to the protein content of foods.

Introduction:The quality of dietary protein sources became a particularly sensitive issue in the current debates on a rebalancing between animal and vegetable food sources.The ability of a protein to meet the nutritional requirements of essential amino acid (EAA) is the basis for assessing the quality of protein.The objective of this study was to characterize the impact of lysine- and threonine-deficient gluten-based diets on the metabolism of growing rats and to identify molecular markers of these diets.Materials and Methods:Growing rats were fed for 3 weeks with a threonine-supplemented and 70% lysine-deficient gluten diet; a lysine-supplemented and 47% threonine-deficient gluten diet; a gluten diet supplemented in lysine and threonine to meet all the AA requirements, and a control diet with milk protein to meet all the AA requirements.Body weight and food intake were measured daily. At the end of the experiment, tissues and biological fluids were removed. The body composition was analyzed, gene expression measurements involved in protein and lipid metabolism were performed and the urinary metabolome was analyzed by LC-MS. Statistical analysis was done by variance analysis and metabolome analysis by discriminant analysis of independent components.Results:This EAA deficiency does not modify the food intake. Lysine deficiency induces a decrease in body weight gain, and lean body mass, associated with an increased in proteolysis and a decreased in proteosynthesis, a decreased in bone mineral density, and no effect on lipid metabolism.Threonine deficiency induces a decrease in body weight gain, and liver and skin weight, without changes in protein metabolism, bone mineral density, and lipid metabolism. After approval of the deficiency model, the metabolomic analysis performed on urine samples revealed the presence of specific discriminating molecules of the diets and types of proteins.Discussion:EAA deficiency has an impact on the growth, and bone and protein metabolism of growing rats. These deficiency states have resulted in different metabolome profiles that could lead to the identification of specific molecular markers of protein sources and related to EAA deficiencies.

International audience; Objectives Severely low-protein (LP) diets induce a decrease in body weight and an increase in relative food including intake (FI) in rat. In the liver, changes in anabolic and catabolic protein pathways could transitorily participate to compensate for amino acid (AA) deficiency. The present study investigated these liver and muscle protein metabolic pathways on LP diet fed growing rats. Methods Growing rats were fed for three weeks different diets containing 3–5–8–12–15 or 20% energy from milk protein. Body weight and FI were measured daily. At the end of the experiment, rats were injected with 13C valine and tissues and biological fluids were collected for gene expression measurement, blood AA UPLC analysis and protein synthesis rate determination in liver and muscle. Statistical analysis was done by 1- or 2-factor ANOVA, when data were repeated. Results P3, P5 and P8% diets resulted in significant growth retardation and significant decrease in lean mass. Severe protein deficiency induced a decrease in the rate of protein synthesis in the liver and muscle. In addition, the results showed activation of the GCN2 pathway, via ATF4-CHOP-TRB3 both in the liver and in the muscle, which suggests the inhibition of the initiation of translation at the level of the binding of the RNAt-Met. Liver proteolytic pathways were up-regulated including the ubiquitin-proteasome, the caspase system and the autophagy. In muscle, both the ubiquitin-proteasome pathway, and autophagy were increased as well as the calpain system. The GCN2 pathway, via ATF4-CHOP-TRB3 was activated in both liver and muscle, confirming the activation of protein degradation by the ubiquitin-proteasome pathways, and autophagy. In portal vein, indispensable AA were lower in severe protein deficient diet whereas in vena cava no difference was observed. Conclusions Severe protein restriction lowered protein synthesis and activated protein catabolism in both liver and muscle whereas no effect was observed for moderate protein restriction. These results confirm that the liver and muscle play a major role in supplying the body with indispensable AA in response to severe protein restriction. Funding Sources This study was funded by the doctoral school ABIES and AlimH-INRAE department.

International audience; Objectives Severely low-protein diets (LP) induce behavioral and metabolic changes including a decrease in body weight, an increase in relative food intake (FI) and alterations in hepatic metabolism. During such protein restriction, changes in hepatic anabolic and catabolic protein pathways could transitory participate to compensate for amino acid (AA) deficiency. In the present study, liver expression of gene involved in proteosynthesis and proteolysis pathways, were related to FI, blood AA levels and body composition in rats fed LP diet. Methods Growing rats were fed for three weeks different diets containing 3-5-8-12-15 or 20% energy of milk protein. Body weight and FI were measured daily. At the end of the experiment, tissues and biological fluids were removed for gene expression measurement and blood AA UPLC analysis. Statistical analysis was done by 1- or 2-factor ANOVA, when data were repeated. Results Despite an increase in relative food intake under P3 and P5% diets, P3, P5 and P8% diets resulted in significant growth retardation compared to other groups. Lean mass was significantly decreased in rats under P3, P5 and P8% compared to P12, P15 and P20% diets, while there was no difference in fat mass between all groups. P3, P5 and P8% diets induced a decrease in essential amino acid concentrations in portal vein, whereas there was no significant difference between groups in veina cava. Severely protein restricted P3% and P5% diets induced an increase in hepatic gene expression involved in proteolysis as calpain 2 and ubiquitin, and an activation of ATF4-CHOP-TRB3 pathway. Conclusions These results suggested that under severe protein restriction, hepatic protein catabolism became a source of plasma amino acid that could partially compensate for the AA not provided by the diet. These observations confirm that liver plays a major role in the adaptation of the body to dietary protein restriction and highlight that severe dietary protein restriction induced liver protein catabolism by inducing an activation of ATF4-CHOP-TRB3 pathway in order to provide amino acids to body tissues. Funding Sources ABIES, AlimH-INRAE.

International audience; Low-protein diets most often induce increased energy intake in an attempt to increase protein intake to meet protein needs with a risk of accumulation as fat of the excess energy intake. In female adult BALB/c mice, a decrease in dietary casein from 20% to 6% and 3% increased energy intake and slightly increased adiposity, and this response was exacerbated with soy proteins with low methionine content. The effect on fat mass was however limited because total energy expenditure increased to the same extent as energy intake. Lean body mass was preserved in all 6% fed mice and reduced only in 3% casein-fed animals. Insulin response to an oral glucose tolerance test was reduced in soy-fed mice and in low-protein-fed mice. Low-protein diets did not affect uncoupling protein 1 and increased fibroblast growth factor 21 (FGF21) in brown adipose tissue and increased FGF21, fatty acid synthase, and cluster of differentiation 36 in the liver. In the hypothalamus, neuropeptide Y was increased and proopiomelanocortin was decreased only in 3% casein-fed mice. In plasma, when protein was decreased, insulin-like growth factor-1 decreased and FGF21 increased and plasma FGF21 was best described by using a combination of dietary protein level, protein-to-carbohydrate ratio, and protein-to-methionine ratio in the diet. In conclusion, reducing dietary protein and protein quality increases energy intake but also energy expenditure resulting in an only slight increase in adiposity. In this process, FGF21 is probably an important signal that responds to a complex combination of protein restriction, protein quality, and carbohydrate content of the diet.

International audience; BackgroundWe have reported large differences in adiposity (fat mass/body weight) gain between rats fed a low-fat, high-starch diet, leading to their classification into carbohydrate “sensitive” and “resistant” rats. In sensitive animals, fat accumulates in visceral adipose tissues, leading to the suggestion that this form of obesity could be responsible for rapid development of metabolic syndrome.ObjectiveWe investigated whether increased amylase secretion by the pancreas and accelerated starch degradation in the intestine could be responsible for this phenotype.MethodThirty-two male Wistar rats (7-wk-old) were fed a purified low-fat (10%), high-carbohydrate diet for 6 wk, in which most of the carbohydrate (64% by energy) was provided as corn starch. Meal tolerance tests of the Starch diet were performed to measure glucose and insulin responses to meal ingestion. Indirect calorimetry combined with use of 13C-labelled dietary starch was used to assess meal-induced changes in whole body and starch-derived glucose oxidation. Real-time polymerase chain reaction was used to assess mRNA expression in pancreas, liver, white and brown adipose tissues, and intestine. Amylase activity was measured in the duodenum, jejunum, and ileum contents. ANOVA and regression analyses were used for statistical comparisons.Results“Resistant” and “sensitive” rats were separated according to adiposity gain during the study (1.73% ± 0.20% compared with 4.35% ± 0.36%). Breath recovery of 13CO2 from 13C-labelled dietary starch was higher in “sensitive” rats, indicating a larger increase in whole body glucose oxidation and, conversely, a larger decrease in lipid oxidation. Amylase mRNA expression in pancreas, and amylase activity in jejunum, were also higher in sensitive rats.ConclusionDifferences in digestion of starch can promote visceral fat accumulation in rats when fed a low-fat, high-starch diet. This mechanism may have important implications in human obesity.

Epub ahead of print; OBJECTIVE: GCN2 is a kinase which detects amino acid deficiency and is involved in the control of protein synthesis and energy metabolism. However, the role of hepatic GCN2 in the metabolic adaptations in response to the modulation of dietary protein has been seldom studied. METHODS: Wild-type (WT) and liver GCN2-deficient (KO) mice were fed either a normo-protein diet, a low-protein diet or a high-protein diet for 3 weeks. During this period, body weight, food intake and metabolic parameters were followed. RESULTS: In mice fed normo- and high protein diets, GCN2 pathway in the liver is not activated in WT mice leading to a similar metabolic profile with the one of KO mice. On the contrary, a low protein diet activates GCN2 in WT mice inducing FGF21 secretion. In turn, FGF21 maintains a high level of lipid oxidation leading to a different postprandial oxidation profile compared with KO mice. CONCLUSIONS: Hepatic GCN2 controls FGF21 secretion under a low protein diet, and modulates a whole-body postprandial oxidation profile.

The energy sensor AMP-activated protein kinase (AMPK) is a key player in the control of energy metabolism. AMPK regulates hepatic lipid metabolism through the phosphorylation of its well-recognized downstream target acetyl CoA carboxylase (ACC). Although AMPK activation is proposed to lower hepatic triglyceride (TG) content via the inhibition of ACC to cause inhibition of de novo lipogenesis and stimulation of fatty acid oxidation (FAO), its contribution to the inhibition of FAO in vivo has been recently questioned. We generated a mouse model of AMPK activation specifically in the liver achieved by expression of a constitutively active AMPK using adenoviral delivery. Indirect calorimetry studies revealed that liver-specific AMPK activation is sufficient to induce a reduction in the respiratory exchange ratio and an increase in FAO rates in vivo. This led to a more rapid metabolic switch from carbohydrate to lipid oxidation during the transition from fed to fasting. Finally, mice with chronic AMPK activation in the liver display high fat oxidation capacity evidenced by increased [C14]-palmitate oxidation and ketone body production leading to reduced hepatic TG content and body adiposity. Our findings suggest a role for hepatic AMPK in the remodeling of lipid metabolism between the liver and adipose tissue.

Discipline Experimental/mecanismes cellulaires et moleculaires Introduction et but de l’etude La qualite des sources alimentaires de proteines est devenue une question particulierement sensible dans le cadre des debats actuels sur un reequilibrage entre les sources alimentaires animales et vegetales. Les proteines sont un composant alimentaire indispensable a la survie, fournissant de l’azote et des acides amines (AA), et en particulier les neuf acides amines indispensables (AAI). En outre, parmi les AAI, la lysine et la threonine sont dits strictement indispensables car ne participant pas aux processus de transamination. La capacite d’une proteine a couvrir les besoins nutritionnels en AAI est a la base de l’evaluation de la qualite des sources alimentaires de proteines. L’objectif de cette etude etait de caracteriser l’impact de regimes a base de gluten deficients en lysine et en threonine sur le metabolisme de rats en croissance et d’identifier des biomarqueurs de ces regimes. L’hypothese est qu’un deficit en un AAI entraine une activation des systemes enzymatiques de catabolisme des autres AA, produisant des metabolites specifiques de ces degradations detectables dans les fluides biologiques et susceptibles de representer des biomarqueurs de tels regimes. Materiel et methodes Des rats en croissance ont ete nourris pendant 3 semaines avec un regime a base de gluten supplemente en threonine et deficient a 70% en lysine, un regime a base de gluten supplemente en lysine et deficient a 47% en threonine, un regime a base de gluten supplemente en lysine et en threonine couvrant 100% des besoins en tous les AA, et un regime temoin a base de proteine de lait couvrant 100% des besoins en tous les AA (APAFiS #5497). Le poids et la prise alimentaire ont ete mesures quotidiennement. A la fin de l’experience, les tissus et fluides biologiques ont ete preleves. La composition corporelle a ete analysee, des mesures d’expression de genes impliques dans le metabolisme proteique et lipidique ont ete realisees et le metabolome urinaire a ete analyse par LC-MS. L’analyse statistique a ete faite par analyse de variance et l’analyse des metabolomes par analyse discriminante des composants independants (IC-DA). Resultats et analyse statistique La deficience en ces AAI ne modifie pas la prise alimentaire. La deficience en lysine induit une diminution du gain de poids corporel, et de la masse maigre, en lien avec une augmentation de la proteolyse et une diminution de la proteosynthese, une diminution de la densite minerale osseuse, et aucun effet sur le metabolisme lipidique. La deficience en threonine, induit une diminution du gain de poids corporel, et du poids du foie et de la peau, sans modification du metabolisme proteique, de la densite minerale osseuse et metabolisme lipidique. Apres validation du modele de deficience, l’analyse metabolomique realisee sur des echantillons d’urine a mis en evidence la presence de molecules discriminantes specifiques des regimes et des types de proteines. Conclusion La deficience en un AAI a un impact sur la croissance et le metabolisme osseux et proteique de rats en croissance. Ces etats de deficience se sont traduits par des profils de metabolomes differents qui pourraient conduire a l’identification de biomarqueurs specifiques des sources de proteines et des deficiences associees en AAI.

International audience; The ingestion of low or high lipid diets enriched with fructo-oligosaccharide (FOS) affects energy homeostasis. Ingesting protein diets also induces a depression of energy intake and decreases body weight. The goal of this study was to investigate the ability of FOS, combined or not with a high level of protein (P), to affect energy intake and body composition when included in diets containing different levels of lipids (L). We performed two studies of similar design over a period of 5 weeks. During the first experiment (exp1), after a 3-week period of adaptation to a normal protein-low fat diet, the rats received one of the following four diets for 5 weeks (6 rats per group): (i) normal protein (14% P/E (Energy) low fat (10% L/E) diet, (ii) normal protein, low fat diet supplemented with 10% FOS, (iii) high protein (55%P/E) low fat diet, and (iv) high protein, low fat diet supplemented with 10% FOS. In a second experiment (exp2) after the 3-week period of adaptation to a normal protein-high fat diet, the rats received one of the following 4 diets for 5 weeks (6 rats per group): (i) normal protein, high fat diet (35% of fat), (ii) normal protein, high fat diet supplemented with 10% FOS, (iii) high protein high fat diet and (iv) high protein high fat diet supplemented with 10% FOS. In low-fat fed rats, FOS did not affect lean body mass (LBM) and fat mass but the protein level reduced fat mass and tended to reduce adiposity. In high-fat fed rats, FOS did not affect LBM but reduced fat mass and adiposity. No additive or antagonistic effects between FOS and the protein level were observed. FOS reduced energy intake in low-fat fed rats, did not affect energy intake in normal-protein high-fat fed rats but surprisingly, and significantly, increased energy intake in high-protein high-fat fed rats. The results thus showed that FOS added to a high-fat diet reduced body fat and body adiposity.

International audience; Background: Hepatic AMP-activated kinase (AMPK) activity is sensitive to the dietary carbohydrate-to-protein ratio. However, the role of AMPK in metabolic adaptations to variations in dietary macronutrients remains poorly understood.Objective: The objective of this study was to determine the role of hepatic AMPK in the adaptation of energy metabolism in response to modulation of the dietary carbohydrate-to-protein ratio.Methods: Male 7-wk-old wild-type (WT) and liver AMPK–deficient (knockout) mice were fed either a normal-protein and normal-carbohydrate diet (NP-NC; 14% protein, 76% carbohydrate on an energy basis), a low-protein and high-carbohydrate diet (LP-HC; 5% protein, 85% carbohydrate), or a high-protein and low-carbohydrate diet (HP-LC; 55% protein, 35% carbohydrate) for 3 wk. During this period, after an overnight fast, metabolic parameters were measured and indirect calorimetry was performed in mice during the first hours after refeeding a 1-g calibrated meal of their own diet in order to investigate lipid and carbohydrate metabolism.Results: Knockout mice fed an LP-HC or HP-LC meal exhibited 24% and 8% lower amplitudes in meal-induced carbohydrate and lipid oxidation changes. By contrast, knockout mice fed an NP-NC meal displayed normal carbohydrate and lipid oxidation profiles. These mice exhibited a transient increase in hepatic triglycerides and a decrease in hepatic glycogen. These changes were associated with a 650% higher secretion of fibroblast growth factor 21 (FGF21) 2 h after refeeding.Conclusions: The consequences of hepatic AMPK deletion depend on the dietary carbohydrate-to-protein ratio. In mice fed the NP-NC diet, deletion of AMPK in the liver led to an adaptation of liver metabolism resulting in increased secretion of FGF21. These changes possibly compensated for the absence of hepatic AMPK, as these mice exhibited normal postprandial changes in carbohydrate and lipid oxidation. By contrast, in mice fed the LP-HC and HP-LC diets, the lack of adjustment in liver metabolism in knockout mice resulted in a metabolic inflexibility, leading to a reduced amplitude of meal-induced changes in carbohydrate and lipid oxidation.

International audience; Growth hormone (GH) and insulinlike growth factor (IGF) promote aging and age-related pathologies. Inhibiting this pathway by targeting IGF receptor (IGF-1R) is a promising strategy to extend life span, alleviate age-related diseases, and reduce tumor growth. Although anti-IGF-1R agents are being developed, long-term effects of IGF-1R blockade remain unknown. In this study, we used ubiquitous inducible IGF-1R knockout (UBIKOR) to suppress signaling in all adult tissues and screened health extensively. Surprisingly, UBIKOR mice showed no overt defects and presented with rather inconspicuous health, including normal cognition. Endocrine GH and IGF-1 were strongly upregulated without causing acromegaly. UBIKOR mice were strikingly lean with coordinate changes in body composition and organ size. They were insulin resistant but preserved physiological energy expenditure and displayed enhanced fasting metabolic flexibility. Thus, long-term IGF-1R blockade generated beneficial effects on aging-relevant metabolism, but exposed to high GH. This needs to be considered when targeting IGF-1R to protect from neurodegeneration, retard aging, or fight cancer.

OBJECTIVES: Low-protein diets are reported to induce hyperphagia to fulfil protein needs but at the expense of energy balance with a risk to gain in adiposity. However, different studies did not show body fat gain because an increased energy expenditure partly compensated the increase in energy intake and prevents the gain in adiposity. The present study evaluated in mice the consequence of protein restricted diets combined with protein quality (milk protein versus soy protein with slight methionine deficiency) on energy balance and adiposity and the role of FGF21 in the response to the protein restricted diets. METHODS: The study investigated in female BalbC mice the behavioural, metabolic and phenotypic responses to 8 weeks feeding a very low (3% energy - P3), moderately low (6% - P6) or adequate (20% - P20) dietary protein diet and evaluated if methionine scarcity, using soy protein (S) vs casein (C), affected these responses. Food intake, body weight, adiposity (assessed by DEXA), were measured throughout the study and body composition determined at the end of the study. Plasma, liver, muscle, adipose tissue and hypothalamus samples were collected for nutrient, hormones and gene expression measurements. RESULTS: Decreasing dietary casein from 20% to 3% increased energy intake, slightly increased adiposity, and this was exacerbated with methionine-deficient soy protein (figure 1). Lean body mass was reduced in 3% casein fed mice but preserved in all 6% fed mice. The effect on fat mass was limited because energy expenditure was also increased (figure 2). In plasma, low protein diets decreased IGF-1 and increased FGF21 that was related to protein level, protein to carbohydrate ratio and methionine content in the diet (figure 3). Insulin response to an oral glucose test was reduced in soy and low-protein fed mice. Low-protein diets did not affect Ucp1 but increased Fgf21 in brown adipose tissue and Fgf21, Fas, and Cd36 in the liver. In the hypothalamus, Npy was increased and Pomc was decreased only in 3% casein fed mice. CONCLUSIONS: Reducing dietary protein and protein quality increases both energy intake and energy expenditure resulting only in slight increase in adiposity. In this process FGF21 is probably a signal that responds to a combination of protein restriction and carbohydrate content of the diet. FUNDING SOURCES: Institut National de la Recherche Agronomique, AgroParisTech. SUPPORTING TABLES, IMAGES AND/OR GRAPHS

OBJECTIVES: Low protein diet and essential amino acid deficient-diet have an impact on body weight and growth and different studies also showed an impact of lysine intake on bone metabolism. Lysine has been shown to promote the absorption of intestinal calcium and to participate in the collagen synthesis through its involvement in the reticulation process of the tropocollagen beams. The assembly of tropocollagen bundle into mature collagen fibers is essential for bone formation and remodeling (civitelli et al, 1992; Fini et al, 2001). The objective of this study was to characterize the impact of low protein diet and lysine-deficient diet on bone metabolism of growing rats. METHODS: Study 1: 6 group of growing rats were fed for 3 weeks different diet with different content of milk protein at levels of 3%, 5%, 8%, 12%, 15% or 20% (% total energy). Study 2: 7 group of growing rats were fed diets with different lysine content (as % of lysine requirement), for 3 weeks: 15%, 25%, 40%, 60%, 75%, 100% or 170% (% Lysine requirement). Body weight was measured daily. At the end of the experiment, the body composition was analyzed and tissues were removed for measurements of the expression of genes involved in protein and bone metabolism. Statistical analysis was done by variance analysis. RESULTS: Rats fed low protein diet (3% and 5% of milk protein), compared to control have a lower growth, with a lower body weight and naso-anal length. This weak growth was associated with a lower lean body mass, and also had an impact on bone metabolism. There was a decrease in the bone mineral density, bone mineral content and femur size, associated with a decrease of markers of bone turnover and formation. The same results on bone metabolism were observed on rats fed the 85% lysine deficient diet. CONCLUSIONS: Low protein diet and lysine-deficient diet reduce growth and bone metabolism. The impact of low protein diet could be related to the lysine deficiency, which have an impact on the calcium intestinal absorption and on collagen synthesis. FUNDING SOURCES: INRA, AgroParisTech. Supporting Tables, Images and/or Graphs

Orexin-A and -B are hypothalamic neuropeptides of 33 and 28-amino acids, which regulate many homeostatic systems including sleep/wakefulness states, energy balance, energy homeostasis, reward seeking and drug addiction. Orexin-A treatment was also shown to reduce tumor development in xenografted nude mice and is thus a potential treatment for carcinogenesis. The aim of this work was to explore in healthy mice the consequences on energy expenditure components of an orexin-A treatment at a dose previously shown to be efficient to reduce tumor development. Physiological approaches were used to evaluate the effect of orexin-A on food intake pattern, energy metabolism body weight and body adiposity. Modulation of the expression of brain neuropeptides and receptors including NPY, POMC, AgRP, cocaine-and amphetamine related transcript (CART), corticotropin-releasing hormone (CRH) and prepro-orexin (HCRT), and Y-2 and Y-5 neuropeptide Y, MC4 (melanocortin), OX1 and OX2 orexin receptors (Y2R, Y5R, MC4R, OX1R and OX2R, respectively) was also explored. Our results show that orexin-A treatment does not significantly affect the components of energy expenditure, and glucose metabolism but reduces intraperitoneal fat deposit, adiposity and the expression of several brain neuropeptide receptors suggesting that peripheral orexin-A was able to reach the central nervous system. These findings establish that orexin-A treatment which is known for its activity as an inducer of tumor cell death, do have minor parallel consequence on energy homeostasis control.

International audience; The components of energy expenditure, total metabolic rate (TMR), resting metabolic rate (RMR), thermogenic response to feeding (TEF), activity, and cost of activity were measured in fed and fasted mice housed at 22 and 30 degrees C. Mice housed at 22 degrees C had more than two times larger TMR and RMR. Mice at 22 degrees C were less active when fasted but more active when fed. Cost of activity was nearly doubled in the fasted and in the fed state. Analysis of the short-term relation between TMR, RMR, and bouts of activity showed that, at 22 degrees C, the bouts of activity induced a decrease in the intensity of RMR that reflected the reduced need for thermal regulation induced by the heat released from muscular contraction. This phenomenon induced a considerable underestimation of TEF and prevented its reliable measurement when mice were housed at 22 degrees C. Correlation between TMR and activity measured across time in individual mice was very strong at both 22 and 30 degrees C, but the correlation measured across mice was much weaker at 30 degrees C and no longer significant at 22 degrees C. We suspect that this phenomenon was due to the fact that RMR is a much more reliable predictor of TMR than activity. RMR is more variable at 22 degrees C than at 30 degrees C because of heat transfers between thermal regulation and heat released by other discontinuous processes, such as activity and TEE Therefore, more noise is introduced into the correlations performed across multiple mice between TMR and activity at 22 degrees C. On the other hand, it should be kept in mind that the doubling of TMR and RMR at 22 degrees C is fueled by an increased non-shivering thermogenesis that can obviously modify how the mouse responds to pharmacological and nutritional challenges. Taken together, these results suggest that in pre-clinical studies, mice should be housed in conditions where thermal regulation is limited as is generally the case in humans. However, the increased sensitivity of mice to small changes in ambient temperature can also be used as a versatile tool to investigate the role of thermal regulation on the energy balance equation in humans.

WOS:000385213000006; International audience; Fibroblast growth factor 21 (FGF21) is a polypeptide secreted by the liver and involved in several metabolic processes such as thermogenesis and lipid oxidation. The nutritional mechanisms controlling FGF21 production are poorly understood. This study aimed to investigate how dietary carbohydrates and proteins impact FGF21 production and how in turn, FGF21 is involved in the metabolic adaptation to changes in the carbohydrate and protein contents of the diet. For that purpose, we fed 25 male C57BL/6 mice diets composed of different protein and carbohydrate contents (normal-protein and carbohydrate diet (N=9, NPNC), low-protein high-carbohydrate diet (N=8, LPHC), high-protein low-carbohydrate diet (N=8, HPLC) for 3 weeks. We measured liver Fell gene expression, synthesis and secretion as well as different parameters related to energy and glucose metabolism. We also investigated the direct role of amino acids and glucose in the control of Fgf21 gene expression in hepatocyte primary cultures (n=6). In vivo, FGF21 responds acutely to LPHC intake whereas under an HPLC diet, plasma FGF21 circulating levels are low in the fasted and refed states. In hepatocytes, Fgf21 expression was controlled by glucose but not amino acids. Both diets increased the thermic effect of feeding (TEF) and ketogenesis was increased in fasted HPLC mice. The results presented suggest that dietary glucose, rather than amino acids, directly controls FGF21 secretion, and that FGF21 may be involved in the increased TEF response to LPHC. The effects of the HPLC diet on ketogenesis and TEF are probably controlled by other metabolic pathways. (C) 2016 Elsevier Inc. All rights reserved.

WOS:000377021700018; International audience; We tested the hypothesis that, for rats fed a high-fat diet (HFD), a prioritization of maintaining protein intake may increase energy consumption and hence result in obesity, particularly for individuals prone to obesity ("fat sensitive," FS, vs. "fat resistant," FR). Male Wistar rats (n = 80) first received 3 wk of HFD (protein 15%, fat 42%, carbohydrate 42%), under which they were characterized as being FS (n = 18) or FR (n = 20) based on body weight gain. They then continued on the same HFD but in which protein (100%) was available separately from the carbohydrate: fat (50: 50%) mixture. Under this second regimen, all rats maintained their previous protein intake, whereas intake of fat and carbohydrate was reduced by 50%. This increased protein intake to 26% and decreased fat intake to 37%. Adiposity gain was prevented in both FR and FS rats, and gain in fat-free mass was increased only in FS rats. At the end of the study, the rats were killed 2 h after ingestion of a protein meal, and their tissues and organs were collected for analysis of body composition and measurement of mRNA levels in the liver, adipose tissue, arcuate nucleus, and nucleus accumbens. FS rats had a higher expression of genes encoding enzymes involved in lipogenesis in the liver and white adipose tissue. These results show that FS rats strongly reduced food intake and adiposity gain through macronutrient selection, despite maintenance of a relatively high-fat intake and overexpression of genes favoring lipogenesis.

WOS:000375891000010; International audience; To prevent the development of adiposity-associated metabolic diseases, early biomarkers are needed. Such markers could bring insight to understand the complexity of susceptibility to obesity. Urine and plasma metabolomics fingerprints have been successfully associated with metabolic dysfunctions. Fat resistance (FR) was found to be associated with higher urinary levels of acylglycines and leucine. However, no differences were observed before the diet switch. In this context, we aimed at characterizing metabolic signatures predictive of resistance or sensitivity to fat in the C57Bl6/J mouse model. Urinary metabolic profiles of FR (n = 15) and fat sensitivity (FS) mice (n = 14) were performed on liquid chromatography mass spectrometry. Urinary and plasma metabolic profiles were first collected at baseline (during low-fat diet), then after 10 weeks of high-fat (HF) feeding. Mice were sorted a posteriori into FS and FR based on their final adiposity. After HF feeding for 10 weeks, FS mice tended to have lower plasma levels of p-hydroxybutyrate than FR ones. Urinary metabolic profiles showed that baseline levels of octanoylglycine, leucine and valine were significantly lower in FS mice. Moreover, expressions in the adipose tissue of Baat and Glyat mRNA were lower in FS than in FR mice. In muscle, mRNA encoding CaD and UbE2b tended to be lower in FS mice than in FR mice (P=.056 and P=.071, respectively). The data show that lower levels of urinary octanoylglycine, leucine and valine are potential predictive biomarkers of FS and could be related to a lower stimulation in adipose acyl-coenzyme A conjugation to glycine and to muscle protein breakdown. (C) 2016 Elsevier Inc. All rights reserved.

Summary In this study we investigated the relationship between activity and energy expenditure (EE) in mice. By determining the relationship between activity and EE over a 24 hr period in an individual mouse, activity was calculated to account for 26.6% ± 1.1% of total EE at 30°C. However, when comparing across multiple mice, only 9.53% ± 1.1% of EE from activity appeared to be independent of other components involved in the thermogenic response, suggesting other metabolic processes may mask the contribution of activity to EE. In line with this concept, below thermoneutrality mice still expended a substantial amount of energy on activity; however, at 24°C, 20°C, or 5°C, no independent effect of EE from activity on total daily EE could be detected. Overall these results suggest that when studying mice at temperatures below thermoneutrality, activity is unlikely to explain differences in EE between groups of animals., Highlights ► Under standard lab conditions, more-active mice do not expend more energy

Introduction et but de l’etude Nous avons revele l’existence de rats sensibles a l’obesite sous regime standard riche en glucides et pauvres en graisses (Nadkarni et al., 2013). Ces rats sont appeles « carbohydrate sensitive » (CS) pour etre demarques des rats sensibles a l’obesite sous regime gras (rats dits « obesity prone » [OP]). Nous avons observe que les rats CS presentaient une augmentation des taux d’oxydation des glucides en reponse a un repas test (Chaumontet et al., 2015). L’objectif de notre etude a donc ete de rechercher les mecanismes responsables de cette reponse. Materiel et methodes Les rats ont ete soumis pendant 7 semaines a un regime riche en glucides complexes (amidon). Le poids et la prise alimentaire ont ete mesures 3 fois par semaine. La composition corporelle a ete mesuree par IRM au debut de l’introduction du regime (T0), puis apres 3 (T3) et 6 (T6) semaines de regime. Le gain de masse grasse entre T0 et T3 nous a permis de selectionner les rats sensibles (CS) et resistants (CR). Afin de mesurer l’oxydation postprandiale glucidique, les rats ont recu un repas test de 56 kJ de meme composition que leur regime, mais contenant soit de l’amidon (AMIDON) soit de la maltodextrine (MALTO) marques au 13C. Les rats ont ete places en cage calorimetrique et leurs echanges respiratoires mesures. L’enrichissement en 13CO2 dans les gaz expires a ete mesure par multiflow-IRMS. A la fin de l’experience, les animaux ont ete euthanasies, la composition corporelle mesuree par dissection des principaux tissus et le sang et les tissus cibles collectes pour les mesures biochimiques et d’expression de genes. Resultats et analyse statistique Comme nous l’avions prealablement observe, apres trois semaines de regime, les rats CS sont significativement plus gras que les rats CR, mais le poids et la prise alimentaire ne different pas significativement. Suite a l’ingestion d’un repas MALTO, les taux postprandiaux d’oxydation des glucides et l’apparition de 13CO2 dans les gaz expires augmentent rapidement mais de facon identique chez les rats CR et CS, ce qui indique que l’absorption et l’utilisation metabolique du glucose ne different pas entre les rats CS et CR. Par contre, suite a l’ingestion d’un repas AMIDON, les taux postprandiaux d’oxydation des glucides sont plus eleves chez les rats CS que chez les CR. Cette augmentation coincide avec une apparition plus rapide du 13CO2 dans les gaz expires qui indique que la digestion et l’utilisation metabolique des glucides alimentaires complexes sont plus rapides chez les rats CS. En outre, les ARNm codant l’amylase dans le pancreas sont augmentes chez les rats CS, de meme que l’activite de l’amylase dans le jejunum et dans le sang portal. Conclusion L’augmentation de l’expression de l’amylase pancreatique chez les rats CS induit une acceleration de la digestion et de l’utilisation metabolique du glucose derive des glucides complexes et est probablement responsable du developpement progressif de obesite de type visceral (Nadkarni et al., 2013) observee chez les rats « carbohydrate-sensitive ».

WOS:000369207000023; International audience; Consumption of sugar-sweetened beverages is associated with overweight and obesity. In this study, we hypothesized that obesity-prone (OP) mice fed a high-fat high-sucrose diet (HFHS) are more sensitive to consumption of sucrose-sweetened water (SSW) than obesity-resistant (OR) mice. After 3 weeks of ad libitum access to the HFHS diet (7.5 h/day), 180 male mice were classified as either OP (upper quartile of body weight gain, 5.2 +/- 0.1 g, n = 45) or OR (lower quartile, 3.2 +/- 0.1 g, n = 45). OP and OR mice were subsequently divided into 3 subgroups that had access to HFHS (7.5 h/day) for 16 weeks, supplemented with: i) water (OP/water and OR/water); ii) water and SSW (12.6% w/v), available for 2 h/day randomly when access to HFHS was available and for 5 randomly-chosen days/week (OP/SSW and OR/SSW); or iii) water and SSW for 8 weeks, then only water for 8 weeks (OP/SSW-water and OR/SSW-water). OR/SSW mice decreased their food intake compared to OR/water mice, while OP/SSW mice exhibited an increase in food and total energy intake compared to OP/water mice. OP/SSW mice also gained more body weight and fat mass than OP/water mice, showed an increase in liver triglycerides and developed insulin resistance. These effects were fully reversed in OP/SSW-water mice. In the gut, OR/SSW mice, but not OP/SSW mice, had an increase GLP-1 and CCK response to a liquid meal compared to mice drinking only water. OP/SSW mice had a decreased expression of melanocortin receptor 4 in the hypothalamus and increased expression of delta opioid receptor in the nucleus accumbens compared to OP/water mice when fasted that could explain the hyperphagia in these mice. When access to the sucrose solution was removed for 8 weeks, OP mice had increased dopaminergic and opioidergic response to a sucrose solution. Thus, intermittent access to a sucrose solution in mice fed a HFHS diet induces changes in the gut and brain signaling, leading to increased energy intake and adverse metabolic consequences only in mice prone to HFHSinduced obesity. (C) 2015 Elsevier Inc. All rights reserved.

International audience; Adaptation to a high-protein diet progressively increases the postprandial accumulation of carbon skeletons from dietary amino acids in rats..—We aimed to determine whether oxidative pathways adapt to the overproduction of carbon skeletons resulting from the progressive activation of amino acid (AA) deamination and ureagenesis under a high-protein (HP) diet. Ninety-four male Wistar rats, of which 54 were implanted with a permanent jugular catheter, were fed a normal protein diet for 1 wk and were then switched to an HP diet for 1, 3, 6, or 14 days. On the experimental day, they were given their meal containing a mixture of 20 U-[ 15 N]-[ 13 C] AA, whose metabolic fate was followed for 4 h. Gastric emptying tended to be slower during the first 3 days of adaptation. 15 N excretion in urine increased progressively during the first 6 days, reaching 29% of ingested protein. 13 CO2 excretion was maximal, as early as the first day, and represented only 16% of the ingested proteins. Consequently, the amount of carbon skeletons remaining in the metabolic pools 4 h after the meal ingestion progressively increased to 42% of the deaminated dietary AA after 6 days of HP diet. In contrast, 13 C enrichment of plasma glucose tended to increase from 1 to 14 days of the HP diet. We conclude that there is no oxidative adaptation in the early postprandial period to an excess of carbon skeletons resulting from AA deamination in HP diets. This leads to an increase in the postprandial accumulation of carbon skeletons throughout the adaptation to an HP diet, which can contribute to the sustainable satiating effect of this diet. dietary amino acids; oxidation; deamination; glycogen; stable isotopes THE EFFECT OF HIGH-PROTEIN (HP) diets on energy metabolism has been often questioned (15, 29, 43). In rats fed ad libitum, an HP diet has been shown to lower adiposity, an effect that is not only due to a reduction in the spontaneous energy intake, as revealed by pair-fed studies (4, 30), but also due to metabolic adaptations. When dietary proteins are given in excess, the metabolic pathways adapt in a few days to cope with the excess nitrogen originating from amino acids through an activation of various liver enzymes involved in their catabolism. This has been well documented for the urea cycle (25, 33). In contrast, less is known about the remaining carbon skeletons. It seems that the carbon skeletons produced from the increased deami-nation are poorly catabolized during the postprandial window (14). Indeed, using 15 N and 13 C amino acids administered as oral tracers in an HP meal given to rats, we found that 4 h after ingestion almost half of the carbon skeletons produced from dietary amino acids were still not oxidized (14). Minimal data exist on the adaptation of energy pathways to a large increase in dietary protein intake. In a previous study, we examined the time course adaptation of energy metabolism components to a high-protein diet in rats (38). Together with the adaptation of key enzymes involved in energy pathways (39), we reported a progressive adaptation of glucose metabolism over 2 wk, characterized by an increase in carbohydrate (CHO) oxidation above CHO intake. In contrast, we found no significant changes in energy expenditure. A better understanding of the channeling of carbon skeletons from dietary amino acids toward metabolic pathways is of importance to better understand how amino acids contribute to energy metabolism in situations of protein excess. To compare the time course adaptation of dietary nitrogen and carbon skeleton metabolism to an HP diet, we used a double stable isotope method (15 N and 13 C) to trace dietary proteins and determine the metabolic fate of the nitrogen and carbon compounds .

Dietary protein content has long been investigated for its influence on food behavior. High protein diets promote satiety and reduce calorie intake, while the effects of low protein diets are more contradictory and less well established. Protein sensing may take place in the oral cavity or more certainly in the postoral gastrointestinal tract, where relevant receptors have been found. Protein signaling to the brain may involve the vagal nerve, as well as gastric hormones such as cholecystokinin and peptide YY. Other pathways that may be involved in protein sensing include postabsorptive signaling and the direct influence of amino acid levels in the brain. The consumption of a high protein diet enhances the activity of brain satiety centers, mainly the area postrema, the nucleus of the solitary tract, the arcuate nucleus and other hypothalamic nuclei, and may modify the activity of brain reward centers. A better understanding of the role of both homeostatic and hedonic systems is needed to fully describe the influence of protein consumption on food intake.

Although antipsychotics are established drugs in schizophrenia treatment, they are admittedly known to induce side effects favoring the onset of obesity and worsening its complications. Despite potential involvement of histamine receptor antagonism, or of other neurotransmitter systems, the mechanism by which antipsychotic drugs increase body weight is not elucidated. The aim of the present study was to investigate whether chronic antipsychotic treatments can directly alter the regulation of two main functions of white adipose tissue: lipolysis and glucose utilization. The influence of a classical antipsychotic (haloperidol) was compared to that of two atypical antipsychotics, one known to favor weight gain (olanzapine), the other not (ziprasidone). Cell size, lipolytic capacity and glucose transport activity were determined in white adipocytes of rats subjected to 5-week oral treatment with these antipsychotics. Gene expression of adipocyte proteins involved in glucose transport or fat storage and mobilization, such as glucose transporters (GLUT1 and GLUT4), leptin, matrix metallo-proteinase-9 (MMP9), hormone-sensitive lipase (HSL) and fatty acid synthase (FAS) was also evaluated. Adipocytes from chronic olanzapine-treated rats exhibited decreased lipolytic activity, lowered HSL expression and increased FAS expression. These changes were concomitant to enlarged fat deposition and adipocyte size. Alterations were observed in adipocytes from olanzapine-treated rats whereas the other antipsychotics did not induce any notable disorder. Our results therefore show evidence of an effect of chronic antipsychotic treatment on rat adipocyte metabolism. Thus, impairment of fat cell lipolysis should be considered as a side effect of certain antipsychotics, leading, along with the already documented hyperphagia, to the excessive weight gain observed in patients under prolonged treatment..

WOS:000349588400007; International audience; Intake of sodas has been shown to increase energy intake and to contribute to obesity in humans and in animal models, although the magnitude and importance of these effects are still debated. Moreover, intake of sugar sweetened beverages is often associated with high-fat food consumption in humans. We studied two different accesses to a sucrose-sweetened water (SSW, 12.3%, a concentration similar to that usually found in sugar sweetened beverages) in C57BL/6 mice fed a normal-fat (NF) or a high-fat (HF) diet in a scheduled access (7.5 h). NF-fed and HF-fed mice received during 5 weeks access to water, to SSW continuously for 7.5 h (SSW), or to water plus SSW for 2 h (randomly-chosen time slot for only 5 random days/week) (SSW-2h). Mouse preference for SSW was greater in HF-fed mice than NF-fed mice. Continuous SSW access induced weight gain whatever the diet and led to greater caloric intake than mice drinking water in NF-fed mice and in the first three weeks in HF-fed mice. In HF-fed mice, 2 h-intermittent access to SSW induced a greater body weight gain than mice drinking water, and led to hyperphagia on the HF diet when SSW was accessible compared to days without SSW 2 h-access (leading to greater overall caloric intake), possibly through inactivation of the anorexigenic neuropeptide POMC in the hypothalamus. This was not observed in NF-fed mice, but 2 h-intermittent access to SSW stimulated the expression of dopamine, opioid and endocannabinoid receptors in the nucleus accumbens compared to water-access. In conclusion, in mice, a sucrose solution provided 2 h-intermittently and a high-fat diet have combined effects on peripheral and central homeostatic systems involved in food intake regulation, a finding which has significant implications for human obesity. (C) 2014 Elsevier Inc. All rights reserved.

Angiotensin (Ang) II is produced locally in various tissues, but its role in the regulation of tissue metabolism is still unclear. Recent studies have revealed the role of type 2 Ang II receptor (AT2R) in the control of energy homeostasis and lipid metabolism. The contribution of the AT2R to adaptation to starvation was tested using AT2R-deficient (AT2R (y)(/-)) mice. Fasted AT2R (y)(/-) mice exhibited a lower loss of adipose tissue weight associated to a decreased free fatty acid (FFA) release from stored lipids than the controls. In vitro studies show that Ang II causes an AT1R-mediated antilipolytic effect in isolated adipocytes. AT1R expression is up-regulated by fasting in both genotypes, but the increase is more pronounced in AT2R (y/-) mice. In addition, the increased muscle beta-oxidation displayed in AT2R (y/-) mice on a fed state, persists after fasting compared with wild-type mice. In liver from fed mice, AT2R deficiency did not modify the expression of genes involved in fatty acid oxidation. However, in response to fasting, the large increase of the expression of this subset of genes exhibited by wild-type mice, was impaired in AT2R (y/-) mice. Taken together, decreased lipolytic capacity and increased muscle fatty acid oxidation participate in the decreased plasma FFA observed in fasted AT2R (y/-) mice and could account for the lower FFA metabolism in the liver. These data reveal an important physiological role of AT2R in metabolic adaptations to fasting.

Weight gain is a common and severe side effect of antipsychotic drugs. A usual tool to study the side effects of psychotropic drugs is animal models. However, attempts to create an animal model of antipsychotic-induced weight gain were not successful so far. Female rodents are sensitive to the effects of antipsychotics, but not males. This does not match the human clinical situation. Antipsychotics have different pharmacokinetic properties in rats and humans, and rats and humans have different spontaneous diets.In the present study, we tested the hypothesis that the insensitivity of male rats to the weight-promoting effects of antipsychotics could be related to the mode of administration of antipsychotics and to the animals' diet. Antipsychotics were mixed with the food, and rats were fed a diet resembling the human diet. Rats were treated with 0.01, 0.1, 0.5, and 2 mg/kg of olanzapine or with a control solution for 6 weeks. Their weight and food intake were recorded, and their body composition were analyzed. The effects on weight and food intake of olanzapine (1 mg/kg), haloperidol (1 mg/kg), and ziprasidone (10 mg/kg) were also compared in a 3-week treatment experiment.The results showed that 0.5 and 2 mg of olanzapine, but not lower doses, increase body weight and subcutaneous fat deposition. After the 3-week treatment, olanzapine-treated rats, but not haloperidol- or ziprasidone-treated rats, had significantly increased their weight.This study shows that a rat model of obesity induced by antipsychotics can be created under specific conditions of drug administration, diet, and dose.

The putative effects of a tryptic bovine alphaS1-casein hydrolysate on stress-induced sleep disorders were investigated and their possible link with typical blood stress parameters such as plasma corticosterone concentrations and glycaemia was assessed. Rats were subjected to chronic stress in the form of environmental disturbances, while receiving an oral administration of the alphaS1-casein hydrolysate (CH). Chronic stress significantly reduced sleep duration in control rats during the first 2 days of the stress period, but stress-induced sleep disturbance was prevented in CH-treated rats. Indeed, CH administration allowed the maintenance of slow wave sleep (SWS) duration and even a slight increase in paradoxical sleep (PS) duration in treated rats. Results on plasma corticosterone concentrations and on glycemia values were inconclusive with respect to the implication of the HPA axis in this study. However, the protective effect of the alphaS1-casein hydrolysate on sleep during exposure to our chronic mild stress conditions may be mediated by modulation of the central adrenergic response.

Recent data have suggested that the activity of various brain nuclei is modulated during sleep. In this context, we developed the idea that sleep may participate in adapting brain responsiveness to feeding, so as to tune the control of peripheral energy metabolism. In order to characterize the nature of a possible link between sleep and peripheral energy metabolism, we have investigated the relationship between sleep parameters [wakefulness (W), slow wave sleep (SWS), paradoxical sleep (PS)] and the intensity of peripheral lipid and protein deposition processes. To achieve this, by manipulating the amount and quality of food available to rats, we induced states of energy or protein depletion/repletion which would specifically affect lean or fat body mass, which was quantified by an analysis of body composition. In parallel, using a permanently implanted cortical electrode, we measured electroencephalogram signals (EEG) to quantify the time spent in W, SWS and PS. Analysis of EEG changes in relation to the changes induced in body composition, showed that (1) the amount of sleep (PS and SWS) followed the evolution of energy supply levels, and (2) the time spent in PS relative to SWS varied to a considerable degree (14-23.5%) and followed the same trend as the ratio of lean body mass to fat mass. These results suggest the possible existence of quantitative and qualitative interactions between sleep quality and the anabolic and catabolic processes of peripheral fat and protein deposition.