Hitomi Ogata

Targeted disruption of AdipoR1 and AdipoR2 causes abrogation of adiponectin binding and metabolic actions

Adiponectin plays a central role as an antidiabetic and antiatherogenic adipokine. AdipoR1 and AdipoR2 serve as receptors for adiponectin in vitro, and their reduction in obesity seems to be correlated with reduced adiponectin sensitivity. Here we show that adenovirus-mediated expression of AdipoR1 and R2 in the liver of Lepr−/− mice increased AMP-activated protein kinase (AMPK) activation and peroxisome proliferator-activated receptor (PPAR)-α signaling pathways, respectively. Activation of AMPK reduced gluconeogenesis, whereas expression of the receptors in both cases increased fatty acid oxidation and lead to an amelioration of diabetes. Alternatively, targeted disruption of AdipoR1 resulted in the abrogation of adiponectin-induced AMPK activation, whereas that of AdipoR2 resulted in decreased activity of PPAR-α signaling pathways. Simultaneous disruption of both AdipoR1 and R2 abolished adiponectin binding and actions, resulting in increased tissue triglyceride content, inflammation and oxidative stress, and thus leading to insulin resistance and marked glucose intolerance. Therefore, AdipoR1 and R2 serve as the predominant receptors for adiponectin in vivo and play important roles in the regulation of glucose and lipid metabolism, inflammation and oxidative stress in vivo.

Overexpression of Monocyte Chemoattractant Protein-1 in Adipose Tissues Causes Macrophage Recruitment and Insulin Resistance *

Adipose tissue expression and circulating concentrations of monocyte chemoattractant protein-1 (MCP-1) correlate positively with adiposity. To ascertain the roles of MCP-1 overexpression in adipose, we generated transgenic mice by utilizing the adipocyte P2 (aP2) promoter (aP2-MCP-1 mice). These mice had higher plasma MCP-1 concentrations and increased macrophage accumulation in adipose tissues, as confirmed by immunochemical, flow cytometric, and gene expression analyses. Tumor necrosis factor-α and interleukin-6 mRNA levels in white adipose tissue and plasma non-esterified fatty acid levels were increased in transgenic mice. aP2-MCP-1 mice showed insulin resistance, suggesting that inflammatory changes in adipose tissues may be involved in the development of insulin resistance. Insulin resistance in aP2-MCP-1 mice was confirmed by hyperinsulinemic euglycemic clamp studies showing that transgenic mice had lower rates of glucose disappearance and higher endogenous glucose production than wild-type mice. Consistent with this, insulin-induced phosphorylations of Akt were significantly decreased in both skeletal muscles and livers of aP2-MCP-1 mice. MCP-1 pretreatment of isolated skeletal muscle blunted insulin-stimulated glucose uptake, which was partially restored by treatment with the MEK inhibitor U0126, suggesting that circulating MCP-1 may contribute to insulin resistance in aP2-MCP-1 mice. We concluded that both paracrine and endocrine effects of MCP-1 may contribute to the development of insulin resistance in aP2-MCP-1 mice.

Adiponectin and AdipoR1 regulate PGC-1α and mitochondria by Ca2+ and AMPK/SIRT1

Adiponectin is an anti-diabetic adipokine. Its receptors possess a seven-transmembrane topology with the amino terminus located intracellularly, which is the opposite of G-protein-coupled receptors. Here we provide evidence that adiponectin induces extracellular Ca2+ influx by adiponectin receptor 1 (AdipoR1), which was necessary for subsequent activation of Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ), AMPK and SIRT1, increased expression and decreased acetylation of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), and increased mitochondria in myocytes. Moreover, muscle-specific disruption of AdipoR1 suppressed the adiponectin-mediated increase in intracellular Ca2+ concentration, and decreased the activation of CaMKK, AMPK and SIRT1 by adiponectin. Suppression of AdipoR1 also resulted in decreased PGC-1α expression and deacetylation, decreased mitochondrial content and enzymes, decreased oxidative type I myofibres, and decreased oxidative stress-detoxifying enzymes in skeletal muscle, which were associated with insulin resistance and decreased exercise endurance. Decreased levels of adiponectin and AdipoR1 in obesity may have causal roles in mitochondrial dysfunction and insulin resistance seen in diabetes.

A small-molecule AdipoR agonist for type 2 diabetes and short life in obesity

Adiponectin secreted from adipocytes binds to adiponectin receptors AdipoR1 and AdipoR2, and exerts antidiabetic effects via activation of AMPK and PPAR-α pathways, respectively. Levels of adiponectin in plasma are reduced in obesity, which causes insulin resistance and type 2 diabetes. Thus, orally active small molecules that bind to and activate AdipoR1 and AdipoR2 could ameliorate obesity-related diseases such as type 2 diabetes. Here we report the identification of orally active synthetic small-molecule AdipoR agonists. One of these compounds, AdipoR agonist (AdipoRon), bound to both AdipoR1 and AdipoR2 in vitro. AdipoRon showed very similar effects to adiponectin in muscle and liver, such as activation of AMPK and PPAR-α pathways, and ameliorated insulin resistance and glucose intolerance in mice fed a high-fat diet, which was completely obliterated in AdipoR1 and AdipoR2 double-knockout mice. Moreover, AdipoRon ameliorated diabetes of genetically obese rodent model db/db mice, and prolonged the shortened lifespan of db/db mice on a high-fat diet. Thus, orally active AdipoR agonists such as AdipoRon are a promising therapeutic approach for the treatment of obesity-related diseases such as type 2 diabetes.

Pioglitazone ameliorates insulin resistance and diabetes by both adiponectin-dependent and -independent pathways.

*Thiazolidinediones have been shown to up-regulate adiponectin expression in white adipose tissue and plasma adiponectin levels, and these up-regulations have been proposed to be a major mechanism of the thiazolidinedione-induced amelioration of insulin resistance linked to obesity. To test this hypothesis, we generated adiponectin knock-out (adipo-/-) ob/ob mice with a C57B/6 background. After 14 days of 10 mg/kg pioglitazone, the insulin resistance and diabetes of ob/ob mice were significantly improved in association with significant up-regulation of serum adiponectin levels. Amelioration of insulin resistance in ob/ob mice was attributed to decreased glucose production and increased AMP-activated protein kinase in the liver but not to increased glucose uptake in skeletal muscle. In contrast, insulin resistance and diabetes were not improved in adipo-/-ob/ob mice. After 14 days of 30 mg/kg pioglitazone, insulin resistance and diabetes of ob/ob mice were again significantly ameliorated, which was attributed not only to decreased glucose production in the liver but also to increased glucose uptake in skeletal muscle. Interestingly, adipo-/-ob/ob mice also displayed significant amelioration of insulin resistance and diabetes, which was attributed to increased glucose uptake in skeletal muscle but not to decreased glucose production in the liver. The serum-free fatty acid and triglyceride levels as well as adipocyte sizes in ob/ob and adipo-/-ob/ob mice were unchanged after 10 mg/kg pioglitazone but were significantly reduced to a similar degree after 30 mg/kg pioglitazone. Moreover, the expressions of TNFα and resistin in adipose tissues of ob/ob and adipo-/-ob/ob mice were unchanged after 10 mg/kg pioglitazone but were decreased after 30 mg/kg pioglitazone. Thus, pioglitazone-induced amelioration of insulin resistance and diabetes may occur adiponectin dependently in the liver and adiponectin independently in skeletal muscle.

Dynamic Functional Relay between Insulin Receptor Substrate 1 and 2 in Hepatic Insulin Signaling during Fasting and Feeding

Insulin receptor substrate (Irs) mediates metabolic actions of insulin. Here, we show that hepatic Irs1 and Irs2 function in a distinct manner in the regulation of glucose homeostasis. The PI3K activity associated with Irs2 began to increase during fasting, reached its peak immediately after refeeding, and decreased rapidly thereafter. By contrast, the PI3K activity associated with Irs1 began to increase a few hours after refeeding and reached its peak thereafter. The data indicate that Irs2 mainly functions during fasting and immediately after refeeding, and Irs1 functions primarily after refeeding. In fact, liver-specific Irs1-knockout mice failed to exhibit insulin resistance during fasting, but showed insulin resistance after refeeding; conversely, liver-specific Irs2-knockout mice displayed insulin resistance during fasting but not after refeeding. We propose the concept of the existence of a dynamic relay between Irs1 and Irs2 in hepatic insulin signaling during fasting and feeding.

Metabolic rate and fuel utilization during sleep assessed by whole-body indirect calorimetry

The purpose of this study was to examine metabolic rate and substrate oxidation during sleep in relation to time of sleep and sleep stage. Twelve male subjects free from sleep-disordered breathing slept for 469 +/- 8.7 (mean +/- SE) minutes until natural awakening in a whole-body indirect calorimeter, and polysomnographic documentation of sleep was recorded. Energy expenditure decreased during the first half of the night, reached a nadir (a 35% decrease), and remained relatively stable until awakening. Similarly, fat oxidation decreased from the onset of sleep. On the other hand, carbohydrate oxidation showed no remarkable changes from the onset of sleep but began to increase before awakening. Because distribution of sleep stages is not uniform throughout the night, with rapid-eye-movement (REM) sleep tending to appear later in the sleep, effect of sleep stage on energy metabolism was isolated by analysis of covariance with time as a covariate. Subsequent comparison of metabolic rate by 1-way analysis of variance with Bonferroni post hoc analysis revealed that energy expenditure during REM sleep was significantly greater than that during sleep stages 2 and 3/4 (stage 2, 25.248 +/- 0.961; stage 3/4, 24.825 +/- 0.935; REM, 25.712 +/- 0.928 kcal kg(-1) fat-free mass d(-1)). Carbohydrate oxidation during REM sleep was significantly greater than that during sleep stage 3/4 (stage 3/4, 12.229 +/- 1.071; REM, 13.986 +/- 1.291 kcal kg(-1) fat-free mass d(-1)). Respiration quotient was statistically different among sleep stages, but Bonferroni post hoc analysis failed to identify significant differences (stage 2, 0.850 +/- 0.010; stage 3/4, 0.846 +/- 0.011; REM, 0.861 +/- 0.013). The increases in energy expenditure and carbohydrate oxidation during REM sleep are consistent with a notion that changes in energy metabolism in brain are manifested as small fluctuations in whole-body energy metabolism during sleep.

Algorithm for transient response of whole body indirect calorimeter: deconvolution with a regularization parameter

A whole body indirect calorimeter provides accurate measurement of energy expenditure over long periods of time, but it has limitations to assess its dynamic changes. The present study aimed to improve algorithms to compute O(2) consumption and CO(2) production by adopting a stochastic deconvolution method, which controls the relative weight of fidelity to the data and smoothness of the estimates. The performance of the new algorithm was compared with that of other algorithms (moving average, trends identification, Kalman filter, and Kalman smoothing) against validation tests in which energy metabolism was evaluated every 1 min. First, an in silico simulation study, rectangular or sinusoidal inputs of gradually decreasing periods (64, 32, 16, and 8 min) were applied, and samples collected from the output were corrupted with superimposed noise. Second, CO(2) was infused into a chamber in gradually decreasing intervals and the CO(2) production rate was estimated by algorithms. In terms of recovery, mean square error, and correlation to the known input signal in the validation tests, deconvolution performed better than the other algorithms. Finally, as a case study, the time course of energy metabolism during sleep, the stages of which were assessed by a standard polysomnogram, was measured in a whole body indirect calorimeter. Analysis of covariance revealed an association of energy expenditure with sleep stage, and energy expenditure computed by deconvolution and Kalman smoothing was more closely associated with sleep stages than that based on trends identification and the Kalman filter. The new algorithm significantly improved the transient response of the whole body indirect calorimeter.

Acute effect of late evening meal on diurnal variation of blood glucose and energy metabolism

SUMMARY OBJECTIVE The notion that late evening meal promotes weight gain is popular, and it may also elicit postprandial hyperglycemia, since glucose tolerance decreases during midnight. Diabetic patients with night-eating symptoms, compared with patients without night-eating behaviors, are more likely to be obese and to have elevated A1c. However, epidemiological analysis adjusted for difference in total energy intake did not identify nighttime eating as the risk of obesity. The present study evaluated the effect of a single loading of late evening meal on diurnal variation of blood glucose and 24-h energy expenditure. METHODS Ten young adults stayed twice in a room-size respiratory chamber for 24 h, in a randomized repeated-measures design. After the entrance to the chamber at 1700 h, the subjects took normal (1900 h) or late (2230 h) evening meal, breakfast and lunch, and remained in the chamber until 1700 h. Time course of blood glucose was measured by continuous glucose monitoring system. RESULTS Late evening meal enhanced postprandial blood glucose response to the evening meal and the subsequent breakfast. Overall 24 h average blood glucose level was also elevated by late evening meal. Late evening meal shifted postprandial increase in energy expenditure toward late at night, but overall 24 h energy expenditure remained almost identical in the two dietary conditions. CONCLUSIONS The present study under controlled sedentary condition supports the notion that a single loading of late evening meal enhances average blood glucose over 24 h, but does not support that late evening meal reduces 24 h energy expenditure.

Effect of breakfast skipping on diurnal variation of energy metabolism and blood glucose

Epidemiological studies suggest an association between breakfast skipping and body weight gain, insulin resistance or type 2 diabetes. Time when meal is consumed affects postprandial increase in energy expenditure and blood glucose, and breakfast skipping may reduce 24 h energy expenditure and elevate blood glucose level. The present study evaluated the effect of breakfast skipping on diurnal variation of energy metabolism and blood glucose. The skipped breakfast was compensated by following big meals at lunch and supper. In a randomized repeated-measure design with or without breakfast, eight males stayed twice in a room-size respiratory chamber. Blood glucose was recorded with a continuous glucose monitoring system. Breakfast skipping did not affect 24 h energy expenditure, fat oxidation and thermic effect of food, but increased overall 24 h average of blood glucose (83 ± 3 vs 89 ± 2 mg/dl, P < 0.05). Unlike 24 h glucose level, 24 h energy expenditure was robust when challenged by breakfast skipping. These observations suggest that changes in glucose homeostasis precede that of energy balance, in the potential sequence caused by breakfast skipping, if this dietary habit has any effect on energy balance.: