mTOR controls cell growth, in part by regulating p70 S6 kinase alpha (p70alpha) and eukaryotic initiation factor 4E binding protein 1 (4EBP1). Raptor is a 150 kDa mTOR binding protein that also binds 4EBP1 and p70alpha. The binding of raptor to mTOR is necessary for the mTOR-catalyzed phosphorylation of 4EBP1 in vitro, and it strongly enhances the mTOR kinase activity toward p70alpha. Rapamycin or amino acid withdrawal increases, whereas insulin strongly inhibits, the recovery of 4EBP1 and raptor on 7-methyl-GTP Sepharose. Partial inhibition of raptor expression by RNA interference (RNAi) reduces mTOR-catalyzed 4EBP1 phosphorylation in vitro. RNAi of C. elegans raptor yields an array of phenotypes that closely resemble those produced by inactivation of Ce-TOR. Thus, raptor is an essential scaffold for the mTOR-catalyzed phosphorylation of 4EBP1 and mediates TOR action in vivo.

Blood glucose levels are maintained by the balance between glucose uptake by peripheral tissues and glucose secretion by the liver. Gluconeogenesis is strongly stimulated during fasting and is aberrantly activated in diabetes mellitus. Here we show that the transcriptional coactivator PGC-1 is strongly induced in liver in fasting mice and in three mouse models of insulin action deficiency: streptozotocin-induced diabetes, ob/ob genotype and liver insulin-receptor knockout. PGC-1 is induced synergistically in primary liver cultures by cyclic AMP and glucocorticoids. Adenoviral-mediated expression of PGC-1 in hepatocytes in culture or in vivo strongly activates an entire programme of key gluconeogenic enzymes, including phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase, leading to increased glucose output. Full transcriptional activation of the PEPCK promoter requires coactivation of the glucocorticoid receptor and the liver-enriched transcription factor HNF-4alpha (hepatic nuclear factor-4alpha) by PGC-1. These results implicate PGC-1 as a key modulator of hepatic gluconeogenesis and as a central target of the insulin-cAMP axis in liver.

Acrp30 is a circulating protein synthesized in adipose tissue. A single injection in mice of purified recombinant Acrp30 leads to a 2-3-fold elevation in circulating Acrp30 levels, which triggers a transient decrease in basal glucose levels. Similar treatment in ob/ob, NOD (non-obese diabetic) or streptozotocin-treated mice transiently abolishes hyperglycemia. This effect on glucose is not associated with an increase in insulin levels. Moreover, in isolated hepatocytes, Acrp30 increases the ability of sub-physiological levels of insulin to suppress glucose production. We thus propose that Acrp30 is a potent insulin enhancer linking adipose tissue and whole-body glucose metabolism.

Studies in invertebrates have led to the identification of a number of genes that regulate lifespan, some of which encode components of the insulin or insulin-like signalling pathways. Examples include the related tyrosine kinase receptors InR (Drosophila melanogaster) and DAF-2 (Caenorhabditis elegans) that are homologues of the mammalian insulin-like growth factor type 1 receptor (IGF-1R). To investigate whether IGF-1R also controls longevity in mammals, we inactivated the IGF-1R gene in mice (Igf1r). Here, using heterozygous knockout mice because null mutants are not viable, we report that Igf1r(+/-) mice live on average 26% longer than their wild-type littermates (P < 0.02). Female Igf1r(+/-) mice live 33% longer than wild-type females (P < 0.001), whereas the equivalent male mice show an increase in lifespan of 16%, which is not statistically significant. Long-lived Igf1r(+/-) mice do not develop dwarfism, their energy metabolism is normal, and their nutrient uptake, physical activity, fertility and reproduction are unaffected. The Igf1r(+/-) mice display greater resistance to oxidative stress, a known determinant of ageing. These results indicate that the IGF-1 receptor may be a central regulator of mammalian lifespan.

Omega-3 fatty acids (omega-3 FAs), DHA and EPA, exert anti-inflammatory effects, but the mechanisms are poorly understood. Here, we show that the G protein-coupled receptor 120 (GPR120) functions as an omega-3 FA receptor/sensor. Stimulation of GPR120 with omega-3 FAs or a chemical agonist causes broad anti-inflammatory effects in monocytic RAW 264.7 cells and in primary intraperitoneal macrophages. All of these effects are abrogated by GPR120 knockdown. Since chronic macrophage-mediated tissue inflammation is a key mechanism for insulin resistance in obesity, we fed obese WT and GPR120 knockout mice a high-fat diet with or without omega-3 FA supplementation. The omega-3 FA treatment inhibited inflammation and enhanced systemic insulin sensitivity in WT mice, but was without effect in GPR120 knockout mice. In conclusion, GPR120 is a functional omega-3 FA receptor/sensor and mediates potent insulin sensitizing and antidiabetic effects in vivo by repressing macrophage-induced tissue inflammation.

Methods for assessment, e.g., anthropometric indicators and imaging techniques, of several phenotypes of human obesity, with special reference to abdominal fat content, have been evaluated. The correlation of fat distribution with age, gender, total body fat, energy balance, adipose tissue lipoprotein lipase and lipolytic activity, adipose tissue receptors, and genetic characteristics are discussed. Several secreted or expressed factors in the adipocyte are evaluated in the context of fat tissue localization. The body fat distribution and the metabolic profile in nonobese and obese individuals is discussed relative to lipolysis, antilypolysis and lipogenesis, insulin sensitivity, and glucose, lipid, and protein metabolism. Finally, the endocrine regulation of abdominal visceral fat in comparison with the adipose tissue localized in other areas is presented.

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of elevated liver enzymes in patients of developed countries. We determined the long-term clinical and histological courses of such patients. In a cohort study, 129 consecutively enrolled patients diagnosed with biopsy-proven NAFLD were reevaluated. Survival and causes of death were compared with a matched reference population. Living NAFLD patients were offered repeat liver biopsy and clinical and biochemical investigation. Mean follow-up (SD) was 13.7 (1.3) years. Mortality was not increased in patients with steatosis. Survival of patients with nonalcoholic steatohepatitis (NASH) was reduced (P = .01). These subjects more often died from cardiovascular (P = .04) and liver-related (P = .04) causes. Seven patients (5.4%) developed end-stage liver disease, including 3 patients with hepatocellular carcinoma. The absence of periportal fibrosis at baseline had a negative predictive value of 100% in predicting liver-related complications. At follow-up, 69 of 88 patients had diabetes or impaired glucose tolerance. Progression of liver fibrosis occurred in 41%. These subjects more often had a weight gain exceeding 5 kg (P = .02), they were more insulin resistant (P = .04), and they exhibited more pronounced hepatic fatty infiltration (P = .03) at follow-up. In conclusion, NAFLD with elevated liver enzymes is associated with a clinically significant risk of developing end-stage liver disease. Survival is lower in patients with NASH. Most NAFLD patients will develop diabetes or impaired glucose tolerance in the long term. Progression of liver fibrosis is associated with more pronounced insulin resistance and significant weight gain.

In obesity and type 2 diabetes, expression of the GLUT4 glucose transporter is decreased selectively in adipocytes. Adipose-specific Glut4 (also known as Slc2a4) knockout (adipose-Glut4(-/-)) mice show insulin resistance secondarily in muscle and liver. Here we show, using DNA arrays, that expression of retinol binding protein-4 (RBP4) is elevated in adipose tissue of adipose-Glut4(-/-) mice. We show that serum RBP4 levels are elevated in insulin-resistant mice and humans with obesity and type 2 diabetes. RBP4 levels are normalized by rosiglitazone, an insulin-sensitizing drug. Transgenic overexpression of human RBP4 or injection of recombinant RBP4 in normal mice causes insulin resistance. Conversely, genetic deletion of Rbp4 enhances insulin sensitivity. Fenretinide, a synthetic retinoid that increases urinary excretion of RBP4, normalizes serum RBP4 levels and improves insulin resistance and glucose intolerance in mice with obesity induced by a high-fat diet. Increasing serum RBP4 induces hepatic expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) and impairs insulin signalling in muscle. Thus, RBP4 is an adipocyte-derived 'signal' that may contribute to the pathogenesis of type 2 diabetes. Lowering RBP4 could be a new strategy for treating type 2 diabetes.

White adipose tissue is no longer considered an inert tissue mainly devoted to energy storage but is emerging as an active participant in regulating physiologic and pathologic processes, including immunity and inflammation. Macrophages are components of adipose tissue and actively participate in its activities. Furthermore, cross-talk between lymphocytes and adipocytes can lead to immune regulation. Adipose tissue produces and releases a variety of proinflammatory and anti-inflammatory factors, including the adipokines leptin, adiponectin, resistin, and visfatin, as well as cytokines and chemokines, such as TNF-alpha, IL-6, monocyte chemoattractant protein 1, and others. Proinflammatory molecules produced by adipose tissue have been implicated as active participants in the development of insulin resistance and the increased risk of cardiovascular disease associated with obesity. In contrast, reduced leptin levels might predispose to increased susceptibility to infection caused by reduced T-cell responses in malnourished individuals. Altered adipokine levels have been observed in a variety of inflammatory conditions, although their pathogenic role has not been completely clarified.

A signaling pathway was delineated by which insulin-like growth factor 1 (IGF-1) promotes the survival of cerebellar neurons. IGF-1 activation of phosphoinositide 3-kinase (PI3-K) triggered the activation of two protein kinases, the serine-threonine kinase Akt and the p70 ribosomal protein S6 kinase (p70(S6K)). Experiments with pharmacological inhibitors, as well as expression of wild-type and dominant-inhibitory forms of Akt, demonstrated that Akt but not p70(S6K) mediates PI3-K-dependent survival. These findings suggest that in the developing nervous system, Akt is a critical mediator of growth factor-induced neuronal survival.

Obesity is highly associated with insulin resistance and is the biggest risk factor for non-insulin-dependent diabetes mellitus. The molecular basis of this common syndrome, however, is poorly understood. It has been suggested that tumour necrosis factor (TNF)-alpha is a candidate mediator of insulin resistance in obesity, as it is overexpressed in the adipose tissues of rodents and humans and it blocks the action of insulin in cultured cells and whole animals. To investigate the role of TNF-alpha in obesity and insulin resistance, we have generated obese mice with a targeted null mutation in the gene encoding TNF-alpha and those encoding the two receptors for TNF-alpha. The absence of TNF-alpha resulted in significantly improved insulin sensitivity in both diet-induced obesity and that resulting for the ob/ob model of obesity. The TNFalpha-deficient obese mice had lower levels of circulating free fatty acids, and were protected from the obesity-related reduction in the insulin receptor signalling in muscle and fat tissues. These results indicate that TNF-alpha is an important mediator of insulin resistance in obesity through its effects on several important sites of insulin action.

Although metformin is widely used for the treatment of non-insulin-dependent diabetes, its mode of action remains unclear. Here we provide evidence that its primary site of action is through a direct inhibition of complex 1 of the respiratory chain. Metformin(50 microM) inhibited mitochondrial oxidation of glutamate+malate in hepatoma cells by 13 and 30% after 24 and 60 h exposure respectively, but succinate oxidation was unaffected. Metformin also caused time-dependent inhibition of complex 1 in isolated mitochondria, whereas in sub-mitochondrial particles inhibition was immediate but required very high metformin concentrations (K(0.5),79 mM). These data are compatible with the slow membrane-potential-driven accumulation of the positively charged drug within the mitochondrial matrix leading to inhibition of complex 1. Metformin inhibition of gluconeogenesis from L-lactate in isolated rat hepatocytes was also time- and concentration-dependent, and accompanied by changes in metabolite levels similar to those induced by other inhibitors of gluconeogenesis acting on complex 1. Freeze-clamped livers from metformin-treated rats exhibited similar changes in metabolite concentrations. We conclude that the drug's pharmacological effects are mediated, at least in part, through a time-dependent, self-limiting inhibition of the respiratory chain that restrains hepatic gluconeogenesis while increasing glucose utilization in peripheral tissues. Lactic acidosis, an occasional side effect, canal so be explained in this way.

We show that high doses of salicylates reverse hyperglycemia, hyperinsulinemia, and dyslipidemia in obese rodents by sensitizing insulin signaling. Activation or overexpression of the IkappaB kinase beta (IKKbeta) attenuated insulin signaling in cultured cells, whereas IKKbeta inhibition reversed insulin resistance. Thus, IKKbeta, rather than the cyclooxygenases, appears to be the relevant molecular target. Heterozygous deletion (Ikkbeta+/-) protected against the development of insulin resistance during high-fat feeding and in obese Lep(ob/ob) mice. These findings implicate an inflammatory process in the pathogenesis of insulin resistance in obesity and type 2 diabetes mellitus and identify the IKKbeta pathway as a target for insulin sensitization.

Obesity and its associated comorbidities are among the most prevalent and challenging conditions confronting the medical profession in the 21st century. A major metabolic consequence of obesity is insulin resistance, which is strongly associated with the deposition of triglycerides in the liver. Hepatic steatosis can either be a benign, noninflammatory condition that appears to have no adverse sequelae or can be associated with steatohepatitis: a condition that can result in end-stage liver disease, accounting for up to 14% of liver transplants in the US. Here we highlight recent advances in our understanding of the molecular events contributing to hepatic steatosis and nonalcoholic steatohepatitis.

Tumor necrosis factor-alpha (TNF-alpha) is an important mediator of insulin resistance in obesity and diabetes through its ability to decrease the tyrosine kinase activity of the insulin receptor (IR). Treatment of cultured murine adipocytes with TNF-alpha was shown to induce serine phosphorylation of insulin receptor substrate 1 (IRS-1) and convert IRS-1 into an inhibitor of the IR tyrosine kinase activity in vitro. Myeloid 32D cells, which lack endogenous IRS-1, were resistant to TNF-alpha-mediated inhibition of IR signaling, whereas transfected 32D cells that express IRS-1 were very sensitive to this effect of TNF-alpha. An inhibitory form of IRS-1 was observed in muscle and fat tissues from obese rats. These results indicate that TNF-alpha induces insulin resistance through an unexpected action of IRS-1 to attenuate insulin receptor signaling.

Insulin sensitivity (euglycemic clamp, insulin infusion rate: 40 mU. m(-2). min(-1)) was studied in 30 subjects with biopsy-proven nonalcoholic fatty liver disease (NAFLD), normal glucose tolerance, and a BMI <30 kg/m(2). Of those 30 subjects, 9 had pure fatty liver and 21 had evidence of steatohepatitis. In addition, 10 patients with type 2 diabetes under good metabolic control and 10 healthy subjects were studied. Most NAFLD patients had central fat accumulation, increased triglycerides and uric acid, and low HDL cholesterol, irrespective of BMI. Glucose disposal during the clamp was reduced by nearly 50% in NAFLD patients, as well as in patients with normal body weight, to an extent similar to that of the type 2 diabetic patients. Basal free fatty acids were increased, whereas insulin-mediated suppression of lipolysis was less effective (-69% in NAFLD vs. -84% in control subjects; P = 0.003). Postabsorptive hepatic glucose production (HGP), measured by [6,6-(2)H(2)]glucose, was normal. In response to insulin infusion, HGP decreased by only 63% of basal in NAFLD vs. 84% in control subjects (P = 0.002). Compared with type 2 diabetic patients, NAFLD patients were characterized by lower basal HGP, but with similarly reduced insulin-mediated suppression of HGP. There was laboratory evidence of iron overload in many NAFLD patients, but clinical, histological, and biochemical data (including insulin sensitivity) were not correlated with iron status. Four subjects were heterozygous for mutation His63Asp of the HFE gene of familiar hemochromatosis. We concluded that NAFLD, in the presence of normoglycemia and normal or moderately increased body weight, is characterized by clinical and laboratory data similar to those found in diabetes and obesity. NAFLD may be considered an additional feature of the metabolic syndrome, with specific hepatic insulin resistance.

BACKGROUND

Sleep duration may be an important regulator of body weight and metabolism. An association between short habitual sleep time and increased body mass index (BMI) has been reported in large population samples. The potential role of metabolic hormones in this association is unknown.

RESULTS

Study participants were 1,024 volunteers from the Wisconsin Sleep Cohort Study, a population-based longitudinal study of sleep disorders. Participants underwent nocturnal polysomnography and reported on their sleep habits through questionnaires and sleep diaries. Following polysomnography, morning, fasted blood samples were evaluated for serum leptin and ghrelin (two key opposing hormones in appetite regulation), adiponectin, insulin, glucose, and lipid profile. Relationships among these measures, BMI, and sleep duration (habitual and immediately prior to blood sampling) were examined using multiple variable regressions with control for confounding factors. A U-shaped curvilinear association between sleep duration and BMI was observed. In persons sleeping less than 8 h (74.4% of the sample), increased BMI was proportional to decreased sleep. Short sleep was associated with low leptin (p for slope = 0.01), with a predicted 15.5% lower leptin for habitual sleep of 5 h versus 8 h, and high ghrelin (p for slope = 0.008), with a predicted 14.9% higher ghrelin for nocturnal (polysomnographic) sleep of 5 h versus 8 h, independent of BMI.

CONCLUSIONS

Participants with short sleep had reduced leptin and elevated ghrelin. These differences in leptin and ghrelin are likely to increase appetite, possibly explaining the increased BMI observed with short sleep duration. In Western societies, where chronic sleep restriction is common and food is widely available, changes in appetite regulatory hormones with sleep curtailment may contribute to obesity.

OBJECTIVE

Prevalence of nonalcoholic fatty liver disease (NAFLD) has not been well established. The purpose of this study was to prospectively define the prevalence of both NAFLD and nonalcoholic steatohepatitis (NASH).

METHODS

Outpatients 18 to 70 years old were recruited from Brooke Army Medical Center. All patients completed a baseline questionnaire and ultrasound. If fatty liver was identified, then laboratory data and a liver biopsy were obtained.

RESULTS

Four hundred patients were enrolled. Three hundred and twenty-eight patients completed the questionnaire and ultrasound. Mean age (range, 28-70 years) was 54.6 years (7.35); 62.5% Caucasian, 22% Hispanic, and 11.3% African American; 50.9% female; mean body mass index (BMI) (calculated as kg/m(2)) was 29.8 (5.64); and diabetes and hypertension prevalence 16.5% and 49.7%, respectively. Prevalence of NAFLD was 46%. NASH was confirmed in 40 patients (12.2% of total cohort, 29.9% of ultrasound positive patients). Hispanics had the highest prevalence of NAFLD (58.3%), then Caucasians (44.4%) and African Americans (35.1%). NAFLD patients were more likely to be male (58.9%), older (P = .004), hypertensive (P < .00005), and diabetic (P < .00005). They had a higher BMI (P < .0005), ate fast food more often (P = .049), and exercised less (P = 0.02) than their non-NAFLD counterparts. Hispanics had a higher prevalence of NASH compared with Caucasians (19.4% vs 9.8%; P = .03). Alanine aminotransferase, aspartate aminotransferase, BMI, insulin, Quantitative Insulin-Sensitivity Check Index, and cytokeratin-18 correlated with NASH. Among the 54 diabetic patients, NAFLD was found in 74% and NASH in 22.2%.

CONCLUSIONS

Prevalence of NAFLD and NASH is higher than estimated previously. Hispanics and patients with diabetes are at greatest risk for both NAFLD and NASH.

OBJECTIVE

This report documents that the gastric bypass operation provides long-term control for obesity and diabetes.

BACKGROUND

Obesity and diabetes, both notoriously resistant to medical therapy, continue to be two of our most common and serious diseases.

METHODS

Over the last 14 years, 608 morbidly obese patients underwent gastric bypass, an operation that restricts caloric intake by (1) reducing the functional stomach to approximately 30 mL, (2) delaying gastric emptying with a c. 0.8 to 1.0 cm gastric outlet, and (3) excluding foregut with a 40 to 60 cm Roux-en-Y gastrojejunostomy. Even though many of the patients were seriously ill, the operation was performed with a perioperative mortality and complication rate of 1.5% and 8.5%, respectively. Seventeen of the 608 patients (< 3%) were lost to follow-up.

RESULTS

Gastric bypass provides durable weight control. Weights fell from a preoperative mean of 304.4 lb (range, 198 to 615 lb) to 192.2 lb (range, 104 to 466) by 1 year and were maintained at 205.4 lb (range, 107 to 512 lb) at 5 years, 206.5 lb (130 to 388 lb) at 10 years, and 204.7 lb (158 to 270 lb) at 14 years. The operation provides long-term control of non-insulin-dependent diabetes mellitus (NIDDM). In those patients with adequate follow-up, 121 of 146 patients (82.9%) with NIDDM and 150 of 152 patients (98.7%) with glucose impairment maintained normal levels of plasma glucose, glycosylated hemoglobin, and insulin. These antidiabetic effects appear to be due primarily to a reduction in caloric intake, suggesting that insulin resistance is a secondary protective effect rather than the initial lesion. In addition to the control of weight and NIDDM, gastric bypass also corrected or alleviated a number of other comorbidities of obesity, including hypertension, sleep apnea, cardiopulmonary failure, arthritis, and infertility. Gastric bypass is now established as an effective and safe therapy for morbid obesity and its associated morbidities. No other therapy has produced such durable and complete control of diabetes mellitus.

Insulin resistance is a complex metabolic disorder that defies explanation by a single etiological pathway. Accumulation of ectopic lipid metabolites, activation of the unfolded protein response (UPR) pathway, and innate immune pathways have all been implicated in the pathogenesis of insulin resistance. However, these pathways are also closely linked to changes in fatty acid uptake, lipogenesis, and energy expenditure that can impact ectopic lipid deposition. Ultimately, these cellular changes may converge to promote the accumulation of specific lipid metabolites (diacylglycerols and/or ceramides) in liver and skeletal muscle, a common final pathway leading to impaired insulin signaling and insulin resistance.

Gut peptides, exemplified by glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are secreted in a nutrient-dependent manner and stimulate glucose-dependent insulin secretion. Both GIP and GLP-1 also promote beta cell proliferation and inhibit apoptosis, leading to expansion of beta cell mass. GLP-1, but not GIP, controls glycemia via additional actions on glucose sensors, inhibition of gastric emptying, food intake and glucagon secretion. Furthermore, GLP-1, unlike GIP, potently stimulates insulin secretion and reduces blood glucose in human subjects with type 2 diabetes. This article summarizes current concepts of incretin action and highlights the potential therapeutic utility of GLP-1 receptor agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors for the treatment of type 2 diabetes.

The C-C motif chemokine receptor-2 (CCR2) regulates monocyte and macrophage recruitment and is necessary for macrophage-dependent inflammatory responses and the development of atherosclerosis. Although adipose tissue expression and circulating concentrations of CCL2 (also known as MCP1), a high-affinity ligand for CCR2, are elevated in obesity, the role of CCR2 in metabolic disorders, including insulin resistance, hepatic steatosis, and inflammation associated with obesity, has not been studied. To determine what role CCR2 plays in the development of metabolic phenotypes, we studied the effects of Ccr2 genotype on the development of obesity and its associated phenotypes. Genetic deficiency in Ccr2 reduced food intake and attenuated the development of obesity in mice fed a high-fat diet. In obese mice matched for adiposity, Ccr2 deficiency reduced macrophage content and the inflammatory profile of adipose tissue, increased adiponectin expression, ameliorated hepatic steatosis, and improved systemic glucose homeostasis and insulin sensitivity. In mice with established obesity, short-term treatment with a pharmacological antagonist of CCR2 lowered macrophage content of adipose tissue and improved insulin sensitivity without significantly altering body mass or improving hepatic steatosis. These data suggest that CCR2 influences the development of obesity and associated adipose tissue inflammation and systemic insulin resistance and plays a role in the maintenance of adipose tissue macrophages and insulin resistance once obesity and its metabolic consequences are established.

The recently discovered orexigenic peptide ghrelin is produced primarily by the stomach and circulates in blood at levels that increase during prolonged fasting in rats. When administered to rodents at supraphysiological doses, ghrelin activates hypothalamic neuropeptide Y/agouti gene-related protein neurons and increases food intake and body weight. These findings suggest that ghrelin may participate in meal initiation. As a first step to investigate this hypothesis, we sought to determine whether circulating ghrelin levels are elevated before the consumption of individual meals in humans. Ghrelin, insulin, and leptin were measured by radioimmunoassay in plasma samples drawn 38 times throughout a 24-h period in 10 healthy subjects provided meals on a fixed schedule. Plasma ghrelin levels increased nearly twofold immediately before each meal and fell to trough levels within 1 h after eating, a pattern reciprocal to that of insulin. Intermeal ghrelin levels displayed a diurnal rhythm that was exactly in phase with that of leptin, with both hormones rising throughout the day to a zenith at 0100, then falling overnight to a nadir at 0900. Ghrelin levels sampled during the troughs before and after breakfast correlated strongly with 24-h integrated area under the curve values (r = 0.873 and 0.954, respectively), suggesting that these convenient, single measurements might serve as surrogates for 24-h profiles to estimate overall ghrelin levels. Circulating ghrelin also correlated positively with age (r = 0.701). The clear preprandial rise and postprandial fall in plasma ghrelin levels support the hypothesis that ghrelin plays a physiological role in meal initiation in humans.