Insulin and insulin-like growth factor 1 (IGF-1) have important roles in adipocyte differentiation, glucose tolerance and insulin sensitivity. Here to assess how these pathways can compensate for each other, we created mice with a double tissue-specific knockout of insulin and IGF-1 receptors to eliminate all insulin/IGF-1 signalling in fat. These FIGIRKO mice had markedly decreased white and brown fat mass and were completely resistant to high fat diet-induced obesity and age- and high fat diet-induced glucose intolerance. Energy expenditure was increased in FIGIRKO mice despite a >85% reduction in brown fat mass. However, FIGIRKO mice were unable to maintain body temperature when placed at 4 °C. Brown fat activity was markedly decreased in FIGIRKO mice but was responsive to β3-receptor stimulation. Thus, insulin/IGF-1 signalling has a crucial role in the control of brown and white fat development, and, when disrupted, leads to defective thermogenesis and a paradoxical increase in basal metabolic rate.

Excess adipose tissue is associated with metabolic disease and reduced life span, whereas caloric restriction decreases these risks. Here we show that as mice age, there is downregulation of Dicer and miRNA processing in adipose tissue resulting in decreases of multiple miRNAs. A similar decline of Dicer with age is observed in C. elegans. This is prevented in both species by caloric restriction. Decreased Dicer expression also occurs in preadipocytes from elderly humans and can be produced in cells by exposure to oxidative stress or UV radiation. Knockdown of Dicer in cells results in premature senescence, and fat-specific Dicer knockout renders mice hypersensitive to oxidative stress. Finally, Dicer loss-of-function mutations in worms reduce life span and stress tolerance, while intestinal overexpression of Dicer confers stress resistance. Thus, regulation of miRNA processing in adipose-related tissues plays an important role in longevity and the ability of an organism to respond to environmental stress and age-related disease.

Obesity and type 2 diabetes are associated with mitochondrial dysfunction in adipose tissue, but the role for adipose tissue mitochondria in the development of these disorders is currently unknown. To understand the impact of adipose tissue mitochondria on whole-body metabolism, we have generated a mouse model with disruption of the mitochondrial transcription factor A (TFAM) specifically in fat. F-TFKO adipose tissue exhibit decreased mtDNA copy number, altered levels of proteins of the electron transport chain, and perturbed mitochondrial function with decreased complex I activity and greater oxygen consumption and uncoupling. As a result, F-TFKO mice exhibit higher energy expenditure and are protected from age- and diet-induced obesity, insulin resistance, and hepatosteatosis, despite a greater food intake. Thus, TFAM deletion in the adipose tissue increases mitochondrial oxidation that has positive metabolic effects, suggesting that regulation of adipose tissue mitochondria may be a potential therapeutic target for the treatment of obesity.

Considerable data support the idea that forkhead box O1 (Foxo1) drives the liver transcriptional program during fasting and is then inhibited by thymoma viral proto-oncogene 1 (Akt) after feeding. Here we show that mice with hepatic deletion of Akt1 and Akt2 were glucose intolerant, insulin resistant and defective in their transcriptional response to feeding in the liver. These defects were normalized with concomitant liver-specific deletion of Foxo1. Notably, in the absence of both Akt and Foxo1, mice adapted appropriately to both the fasted and fed state, and insulin suppressed hepatic glucose production normally. A gene expression analysis revealed that deletion of Akt in liver led to the constitutive activation of Foxo1-dependent gene expression, but again, concomitant ablation of Foxo1 restored postprandial regulation, preventing the inhibition of the metabolic response to nutrient intake caused by deletion of Akt. These results are inconsistent with the canonical model of hepatic metabolism in which Akt is an obligate intermediate for proper insulin signaling. Rather, they show that a major role of hepatic Akt is to restrain the activity of Foxo1 and that in the absence of Foxo1, Akt is largely dispensable for insulin- and nutrient-mediated hepatic metabolic regulation in vivo.

A unique mouse model was developed with elevated endogenous GH (2- to 3-fold) and IGF-I (1.2- to 1.4-fold), due to somatotrope-specific Cre-mediated inactivation of IGF-I receptor (IgfIr) and insulin receptor (Insr) genes (IgfIr,Insr(rGHpCre), referred to as HiGH mice). We demonstrate that the metabolic phenotype of HiGH mice is diet dependent and differs from that observed in other mouse models of GH excess due to ectopic heterologous transgene expression or pituitary tumor formation. Elevated endogenous GH promotes lean mass and whole-body lipid oxidation but has minimal effects on adiposity, even in response to diet-induced obesity. When caloric intake is moderated, elevated GH improves glucose clearance, despite low/normal insulin sensitivity, which may be explained in part by enhanced IGF-I and insulin output. However, when caloric intake is in excess, elevated GH promotes hepatic lipid accumulation, insulin resistance, hyperglycemia, and ketosis. The HiGH mouse model represents a useful tool to study the role endogenous circulating GH levels play in regulating health and disease.

We present a consolidated view of the complexity and challenges of designing studies for measurement of energy metabolism in mouse models, including a practical guide to the assessment of energy expenditure, energy intake and body composition and statistical analysis thereof. We hope this guide will facilitate comparisons across studies and minimize spurious interpretations of data. We recommend that division of energy expenditure data by either body weight or lean body weight and that presentation of group effects as histograms should be replaced by plotting individual data and analyzing both group and body-composition effects using analysis of covariance (ANCOVA).

Acetylation has recently emerged as an important mechanism for controlling a broad array of proteins mediating cellular adaptation to metabolic fuels. Acetylation is governed, in part, by SIRTs (sirtuins), class III NAD(+)-dependent deacetylases that regulate lipid and glucose metabolism in liver during fasting and aging. However, the role of acetylation or SIRTs in pathogenic hepatic fuel metabolism under nutrient excess is unknown. In the present study, we isolated acetylated proteins from total liver proteome and observed 193 preferentially acetylated proteins in mice fed on an HFD (high-fat diet) compared with controls, including 11 proteins not previously identified in acetylation studies. Exposure to the HFD led to hyperacetylation of proteins involved in gluconeogenesis, mitochondrial oxidative metabolism, methionine metabolism, liver injury and the ER (endoplasmic reticulum) stress response. Livers of mice fed on the HFD had reduced SIRT3 activity, a 3-fold decrease in hepatic NAD(+) levels and increased mitochondrial protein oxidation. In contrast, neither SIRT1 nor histone acetyltransferase activities were altered, implicating SIRT3 as a dominant factor contributing to the observed phenotype. In Sirt3⁻(/)⁻ mice, exposure to the HFD further increased the acetylation status of liver proteins and reduced the activity of respiratory complexes III and IV. This is the first study to identify acetylation patterns in liver proteins of HFD-fed mice. Our results suggest that SIRT3 is an integral regulator of mitochondrial function and its depletion results in hyperacetylation of critical mitochondrial proteins that protect against hepatic lipotoxicity under conditions of nutrient excess.

Environmental factors, such as the macronutrient composition of the diet, can have a profound impact on risk of diabetes and metabolic syndrome. In the present study we demonstrate how a single, simple dietary factor--leucine--can modify insulin resistance by acting on multiple tissues and at multiple levels of metabolism. Mice were placed on a normal or high fat diet (HFD). Dietary leucine was doubled by addition to the drinking water. mRNA, protein and complete metabolomic profiles were assessed in the major insulin sensitive tissues and serum, and correlated with changes in glucose homeostasis and insulin signaling. After 8 weeks on HFD, mice developed obesity, fatty liver, inflammatory changes in adipose tissue and insulin resistance at the level of IRS-1 phosphorylation, as well as alterations in metabolomic profile of amino acid metabolites, TCA cycle intermediates, glucose and cholesterol metabolites, and fatty acids in liver, muscle, fat and serum. Doubling dietary leucine reversed many of the metabolite abnormalities and caused a marked improvement in glucose tolerance and insulin signaling without altering food intake or weight gain. Increased dietary leucine was also associated with a decrease in hepatic steatosis and a decrease in inflammation in adipose tissue. These changes occurred despite an increase in insulin-stimulated phosphorylation of p70S6 kinase indicating enhanced activation of mTOR, a phenomenon normally associated with insulin resistance. These data indicate that modest changes in a single environmental/nutrient factor can modify multiple metabolic and signaling pathways and modify HFD induced metabolic syndrome by acting at a systemic level on multiple tissues. These data also suggest that increasing dietary leucine may provide an adjunct in the management of obesity-related insulin resistance.