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Background: Nutritional conditions during fetal life influence the risk of the development of metabolic syndrome and cardiovascular diseases in adult life (metabolic programming). Impaired glucose tolerance and dysregulated fatty acid metabolism are hallmarks of metabolic syndrome., Objective: We aimed to establish a mouse model of metabolic programming focusing on the sex-specific effects of a maternal low-protein diet during gestation on glucose and lipid metabolism in the adult offspring., Methods: Pregnant C57BL/6 mice received a control or a low-protein diet (18% vs 9% casein) throughout gestation. Male and female offspring received a low-fat or a high-fat diet from 6 to 22 weeks of age., Results: Maternal low-protein diet during gestation led to deteriorated insulin sensitivity on high-fat feeding in female offspring, as determined by biochemical and microarray analyses. Female offspring of control diet-fed dams were relatively resistant to high-fat diet-induced metabolic dysregulation. In contrast, maternal low-protein diet did not specifically affect the metabolic parameters addressed in male offspring. In males, the high-fat diet led to insulin insensitivity regardless of the diet of the dam., Conclusions: Our findings show that fetal malnutrition has a limited impact on male mouse offspring, yet it does influence the metabolic response to a high-fat diet in females. These findings may have implications for future early diagnostics in metabolic syndrome and for the development of sex-specific treatment regimens., (Copyright © 2012 Elsevier HS Journals, Inc. All rights reserved.)

The primary products of de novo lipogenesis (DNL) are saturated fatty acids, which confer adverse cellular effects. Human adipocytes differentiated with no exogenous fat accumulated triacylglycerol (TG) in lipid droplets and differentiated normally. TG composition showed the products of DNL (saturated fatty acids from 12:0 to 18:0) together with unsaturated fatty acids (particularly 16:1n-7 and 18:1n-9) produced by elongation/desaturation. There was parallel upregulation of expression of genes involved in DNL and in fatty acid elongation and desaturation, suggesting coordinated control of expression. Enzyme products (desaturation ratios, elongation ratios, and total pathway flux) were also correlated with mRNA levels. We used (13)C-labeled substrates to study the pathway of DNL. Glucose (5 mM or 17.5 mM in the medium) provided less than half the carbon used for DNL (42% and 47%, respectively). Glutamine (2 mM) provided 9-10%, depending upon glucose concentration. In contrast, glucose provided most (72%) of the carbon of TG-glycerol. Pathway analysis using mass isotopomer distribution analysis (MIDA) revealed that the pathway for conversion of glucose to palmitate is complex. DNL in human fat cells is tightly coupled with further modification of fatty acids to produce a range of saturated and unsaturated fatty acids consistent with normal maturation.

A growing body of evidence indicates that peroxisome proliferator-activated receptor alpha (PPARalpha) not merely serves as a transcriptional regulator of fatty acid catabolism but also exerts a much broader role in hepatic lipid metabolism. We determined adaptations in hepatic lipid metabolism and related aspects of carbohydrate metabolism upon treatment of C57Bl/6 mice with the PPARalpha agonist fenofibrate. Stable isotope procedures were applied to assess hepatic fatty acid synthesis, fatty acid elongation, and carbohydrate metabolism. Fenofibrate treatment strongly induced hepatic de novo lipogenesis and chain elongation (+/-300, 150, and 600% for C16:0, C18:0, and C18:1 synthesis, respectively) in parallel with an increased expression of lipogenic genes. The lipogenic induction in fenofibrate-treated mice was found to depend on sterol regulatory element-binding protein 1c (SREBP-1c) but not carbohydrate response element-binding protein (ChREBP). Fenofibrate treatment resulted in a reduced contribution of glycolysis to acetyl-CoA production, whereas the cycling of glucose 6-phosphate through the pentose phosphate pathway presumably was enhanced. Altogether, our data indicate that beta-oxidation and lipogenesis are induced simultaneously upon fenofibrate treatment. These observations may reflect a physiological mechanism by which PPARalpha and SREBP-1c collectively ensure proper handling of fatty acids to protect the liver against cytotoxic damage.

Background: High-fat diets promote hepatic lipid accumulation. Paradoxically, these diets also induce lipogenic gene expression in rodent liver. Whether high expression of these genes actually results in an increased flux through the de novo lipogenic pathway in vivo has not been demonstrated., Methodology/principal Findings: To interrogate this apparent paradox, we have quantified de novo lipogenesis in C57Bl/6J mice fed either chow, a high-fat or a n-3 polyunsaturated fatty acid (PUFA)-enriched high-fat diet. A novel approach based on mass isotopomer distribution analysis (MIDA) following 1-(13)C acetate infusion was applied to simultaneously determine de novo lipogenesis, fatty acid elongation as well as cholesterol synthesis. Furthermore, we measured very low density lipoprotein-triglyceride (VLDL-TG) production rates. High-fat feeding promoted hepatic lipid accumulation and induced the expression of lipogenic and cholesterogenic genes compared to chow-fed mice: induction of gene expression was found to translate into increased oleate synthesis. Interestingly, this higher lipogenic flux (+74 microg/g/h for oleic acid) in mice fed the high-fat diet was mainly due to an increased hepatic elongation of unlabeled palmitate (+66 microg/g/h) rather than to elongation of de novo synthesized palmitate. In addition, fractional cholesterol synthesis was increased, i.e. 5.8+/-0.4% vs. 8.1+/-0.6% for control and high fat-fed animals, respectively. Hepatic VLDL-TG production was not affected by high-fat feeding. Partial replacement of saturated fat by fish oil completely reversed the lipogenic effects of high-fat feeding: hepatic lipogenic and cholesterogenic gene expression levels as well as fatty acid and cholesterol synthesis rates were normalized., Conclusions/significance: High-fat feeding induces hepatic fatty acid synthesis in mice, by chain elongation and subsequent desaturation rather than de novo synthesis, while VLDL-TG output remains unaffected. Suppression of lipogenic fluxes by fish oil prevents from high fat diet-induced hepatic steatosis in mice.

Unlabelled: Mitochondrial fatty acid oxidation (mFAO) is considered to be essential for driving gluconeogenesis (GNG) during fasting. However, quantitative in vivo data on de novo synthesis of glucose-6-phosphate upon acute inhibition of mFAO are lacking. We assessed hepatic glucose metabolism in vivo after acute inhibition of mFAO by 30 mg kg(-1) 2-tetradecylglycidic acid (TDGA) in hypoketotic hypoglycemic male C57BL/6J mice by the infusion of [U-(13)C]glucose, [2-(13)C]glycerol, [1-(2)H]galactose, and paracetamol for 6 hours, which was followed by mass isotopomer distribution analysis in blood glucose and urinary paracetamol-glucuronide. During TDGA treatment, endogenous glucose production was unaffected (127 +/- 10 versus 118 +/- 7 micromol kg(-1) minute(-1), control versus TDGA, not significant), but the metabolic clearance rate of glucose was significantly enhanced (15.9 +/- 0.9 versus 26.3 +/- 1.1 mL kg(-1) minute(-1), control versus TDGA,P < 0.05). In comparison with control mice, de novo synthesis of glucose-6-phosphate (G6P) was slightly decreased in TDGA-treated mice (108 +/- 19 versus 85 +/- 6 micromol kg(-1) minute(-1), control versus TDGA, P < 0.05). Recycling of glucose was decreased upon TDGA treatment (26 +/- 14 versus 12 +/- 4 micromol kg(-1) minute(-1), control versus TDGA, P < 0.05). Hepatic messenger RNA (mRNA) levels of genes encoding enzymes involved in de novo G6P synthesis were unaltered, whereas glucose-6-phosphate hydrolase mRNA expressions were increased in TDGA-treated mice. Glucokinase and pyruvate kinase mRNA levels were significantly decreased, whereas pyruvate dehydrogenase kinase isozyme 4 expression was increased 30-fold; this suggested decreased glycolytic activity., Conclusion: Acute pharmacological inhibition of mFAO using TDGA had no effect on endogenous glucose production and only a marginal effect on de novo G6P synthesis. Hence, fully active mFAO is not essential for maintenance of hepatic GNG in vivo in fasted mice.

The dietary fiber guar gum has beneficial effects on obesity, hyperglycemia and hypercholesterolemia in both humans and rodents. The major products of colonic fermentation of dietary fiber, the short-chain fatty acids (SCFAs), have been suggested to play an important role. Recently, we showed that SCFAs protect against the metabolic syndrome via a signaling cascade that involves peroxisome proliferator-activated receptor (PPAR) γ repression and AMP-activated protein kinase (AMPK) activation. In this study we investigated the molecular mechanism via which the dietary fiber guar gum protects against the metabolic syndrome. C57Bl/6J mice were fed a high-fat diet supplemented with 0% or 10% of the fiber guar gum for 12 weeks and effects on lipid and glucose metabolism were studied. We demonstrate that, like SCFAs, also guar gum protects against high-fat diet-induced metabolic abnormalities by PPARγ repression, subsequently increasing mitochondrial uncoupling protein 2 expression and AMP/ATP ratio, leading to the activation of AMPK and culminating in enhanced oxidative metabolism in both liver and adipose tissue. Moreover, guar gum markedly increased peripheral glucose clearance, possibly mediated by the SCFA-induced colonic hormone glucagon-like peptide-1. Overall, this study provides novel molecular insights into the beneficial effects of guar gum on the metabolic syndrome and strengthens the potential role of guar gum as a dietary-fiber intervention. [ABSTRACT FROM AUTHOR]

Apart from impaired β-oxidation, Pparα-deficient (Pparα-/-) mice suffer from hypoglycemia during prolonged fasting, suggesting alterations in hepatic glucose metabolism. We compared hepatic glucose metabolism in vivo in wild type (WT) and Pparα-/- mice after a short term fast, applying novel isotopic methods. After a 9-h fast, mice were infused with [U-13C]glucose, [2-13C]glycerol, [1-²H]galactose, and paracetamol for 6 h, and blood and urine was collected in timed intervals. Plasma glucose concentrations remained constant and were not different between the groups. Hepatic glycogen content was 69 ± 11 and 90 ± 31 μmol/g liver after 15 h of fasting in WT and Pparα-/- mice, respectively. The gluconeogenic flux toward glucose 6-phosphate was not different between the groups (i.e. 157 ± 9 and 153 ± 9 μmol/kg/min in WT and Ppara-/- mice, respectively). The gluconeogenic flux toward plasma glucose, however, was decreased in PPARα-/- mice (i.e. 142 ± 9 versus 124 ± 13 μmol/kg/min) (p < 0.05), accounting for the observed decrease (−15%) in hepatic glucose production in Pparα-/- mice. Expression of the gene encoding glucose-6-phosphate hydrolase (G6ph) was lower in the PPARα-/- mice compared with WT mice. In conclusion, Pparα-/- mice were able to maintain a normal total gluconeogenic flux to glucose 6-phosphate during moderate fasting, despite their inability to up-regulate β-oxidation. However, this gluconeogenic flux was directed more toward glycogen, leading to a decreased hepatic glucose output. This was associated with a down-regulation of the expression of G6ph in PPARα-deficient mice. [ABSTRACT FROM AUTHOR]

Inappropriately elevated hepatic glucose production is considered a major contributor to hyperglycemia in type II diabetes. Increased long-chain fatty acid oxidation is held essential to drive glucose production during fasting. Complete inhibition of long-chain fatty acid oxidation has been explored as a therapeutic option but resulted in hypertrophic cardiomyopathy due to lipid storage. We assessed the effects of inhibition of fatty acid oxidation at the level of medium-chain acyl-CoA dehydrogenase (MCAD) by spiropentaneacetic acid (SPA) on hepatic glucose production in short-term fasted male C57BL/6J mice by infusing [U-13C]glucose, [2-13C]glycerol, [1-2H]galactose and paracetamol for 6 hours, which started immediately after injection of SPA, followed by mass isotopomer distribution analysis on blood glucose and urinary paracetamol-glucuronide. SPA-treatment caused mild hypoketotic hypoglycaemia. Under these conditions, the metabolic clearance rate of glucose was not affected (15.5 ± 0.8 vs. 16.1 ± 0.7 ml⋅kg-1⋅min-1, control vs. SPA). Compared to control mice, the rate of de novo synthesis of glucose-6-phosphate (G6P) was slightly, albeit significantly, decreased in SPA-treated mice (111 ± 8 vs. 84 ± 6 µmol⋅kg-1⋅min-1, control vs. SPA, p<0.05), associated with a corresponding decrease of endogenous glucose production (109 ± 7 vs. 82 ± 4 µmol⋅kg -1⋅min-1, control vs. SPA, p<0.05). Furthermore, only minor changes were observed in fluxes associated with glucose cycling or glycogen metabolism, which were not significant. As a consequence, partitioning of newly synthesised G6P to blood glucose was not changed (69 ± 2 vs. 67 ±. 2%, control vs. SPA, NS). Hepatic mRNA levels of genes encoding enzymes involved in de novo G6P synthesis (Pepck, Pc and Fop-1) were unaltered in SPA-treated mice, while G6P-hydrolase mRNA expressions were increased in SPA-treated mice. In conclusion, acute inhibition of MCAD using SPA only slightly suppressed gluconeogenesis, which resulted in decreased endogenous glucose production because partitioning of newly synthesised G6P to blood glucose did not change. [ABSTRACT FROM AUTHOR]

Inappropriately high hepatic glucose production is a major contributor to hyperglycemia in type II diabetes,. Since long-chain fatty acid oxidation (1cFAO) is considered to be essential for driving gluconeogenesis, inhibition of 1cFAO has been explored as a therapeutic option to diminish glucose production. We assessed hepatic glucose metabolism in vivo after acute inhibition of 1cFAO by 2-tetradecylglycidic acid (TDGA) and subsequent short-term fasting in male C57BL/6J mice and control mice. We infused [U-13C]-glucose, [2-13C]glycerol, [1-2H]galactose and paracetamol for 6 hours, followed by mass isotopomer distribution analysis on blood glucose and urinary paracetamol-glucuronide. Under hypoketotic hypoglycaemic conditions induced by TDGA-treatment, endogenous glucose production was unaffected (127±10 vs. 118 ± 7 µmol⋅kg-1.⋅min-1, control vs. TDGA, NS), but metabolic clearance rate of glucose was enhanced (16 ± 1 vs. 26 ±1 ml⋅kg-l.⋅min-1, control vs. TDGA, p<0.05). Compared to control mice, the rate of de novo synthesis of glucose-6-phosphate (G6P) was only slightly decreased in TDGA-treated mice (108 ± 19 vs. 85 ± 6 µmol.⋅kg-1⋅min- 1, control vs. TDGA, p<0.05). Recycling of glucose was decreased upon TDGA-treatment (26 ±14 vs. 12 ±4 µmol.⋅kg -1⋅min-1, control vs. TDGA, p<0.05) due to decreased glucose phosphorylation (39 ± 8 vs. 16 ± 2 µmol⋅kg-1⋅min-1, control s. TDGA, p<0.05). Furthermore, glycogen synthesis decreased significantly (56 ± 5 vs. 33 ±1 µmol⋅kg-1⋅min-1, control vs. TDGA, p<0.05), resulting in an increased partitioning of newly synthesised G6P to blood glucose. Hepatic mRNA levels of genes encoding for enzymes involved in de novo G6P-synthesis were unaltered, while G6P-hydrolase mRNA expressions were increased in TDGA-treated mice. Glucokinase and pyruvate kinase mRNA levels were significantly decreased and Pdk-4 expression was increased 30-fold, suggesting decreased glycolytic activity. In conclusion, acute pharmacological inhibition of 1cFAO using TDGA in short-term fasted mice suppressed de novo G6P-synthesis only marginally, while endogenous glucose production remained unaffected. Hence, it appears that a fully active 1cFAO is not essential for maintenance of hepatic GNG in vivo, but affects predominantly metabolic handling of G6P. [ABSTRACT FROM AUTHOR]

Hyperglycemia in type II diabetes is partly due to an inappropriately elevated hepatic glucose production. Long-chain fatty acid oxidation (1cFAO) is held essential to drive glucose production during fasting. Treatment of type II diabetes by inhibition of 1cFAO has been unsuccessful, because it is accompanied by hypertrophic cardiomyopathy due to lipid accumulation. Medium-chain acyl-CoA dehydrogenase (MCAD) is responsible for the first step in medium-chain fatty acid oxidation (mcFAO) essential for complete 1cFAO. As proof of principal to treat type II diabetes by MCAD inhibition, we assessed hepatic glucose metabolism in vivo in short-term fasted male MCAD-deficient (MCADD) mice and their WT littermates, by infusing [U-13C]glucose, [2-13C]glycerol, [1-2H]galactose, and paracetamol for 6 hours, followed by mass isotopomer distribution analysis on blood glucose and urinary paracetamol-glucuronide. Upon short-term fasting, glucose concentration in blood were comparable in both groups (6.2 ± 0.4 vs. 5.8 ± 0.6 mmol/L, control vs. MCADD mice, NS). Endogenous glucose production tended to be decreased in MCADD mice compared to controls (143 ± 14 vs. 113 ± 12 p-mol⋅kg-l⋅min-1, control vs. MCADD, NS). Metabolic clearance rate of glucose did not differ between the groups (18 ± 2 vs. 20 ± 3 ml⋅kg-1⋅min-1, control vs. MCADD, NS). De novo synthesis of glucose-6-phosphate (G6P) was not affected in MCADD mice (124 ± 23 vs. 124 ± 19 µmol⋅kg-1⋅min-1, control vs. MCADD, NS), as was glucose cycling (31 ± 10 vs. 27 ± 5 µmol⋅kg-1⋅min-1, control vs. MCADD, NS). In contrast, fluxes associated with glycogen metabolism were changed, with increased flux into glycogen (47 ± 10 vs. 70 ± 14 µmol⋅kg-1⋅min-1, control vs. MCADD, p<0.05) and decreased flux out of glycogen (59 ± 14 vs. 31 ± 15 µmol⋅kg-1⋅min-1, control vs. MCADD, p<0.05). Hepatic mRNA levels of genes encoding for enzymes involved in de novo G6P synthesis, glucose production, glucose phosphorylation and glycogen metabolism were unaltered in MCADD mice. In conclusion, short-term fasting did not affect gluconeogenesis in MCADD mice but surprisingly, changed fluxes associated with hepatic glycogen metabolism. Our data indicate that inhibition of mcFAO is not a therapeutic option in type II diabetes. [ABSTRACT FROM AUTHOR]