Berberine has been shown to have antidiabetic properties, although its mode of action is not known. Here, we have investigated the metabolic effects of berberine in two animal models of insulin resistance and in insulin-responsive cell lines. Berberine reduced body weight and caused a significant improvement in glucose tolerance without altering food intake in db/db mice. Similarly, berberine reduced body weight and plasma triglycerides and improved insulin action in high-fat-fed Wistar rats. Berberine downregulated the expression of genes involved in lipogenesis and upregulated those involved in energy expenditure in adipose tissue and muscle. Berberine treatment resulted in increased AMP-activated protein kinase (AMPK) activity in 3T3-L1 adipocytes and L6 myotubes, increased GLUT4 translocation in L6 cells in a phosphatidylinositol 3 kinase-independent manner, and reduced lipid accumulation in 3T3-L1 adipocytes. These findings suggest that berberine displays beneficial effects in the treatment of diabetes and obesity at least in part via stimulation of AMPK activity.

OBJECTIVE-Berberine (BBR) activates AMP-activated protein kinase (AMPK) and improves insulin sensitivity in rodent models of insulin resistance. We investigated the mechanism of activation of AMPK by BBR and explored whether derivatization of BBR could improve its in vivo efficacy. RESEARCH DESIGN AND METHODS-AMPK phosphorylation was examined in L6 myotubes and LKB1 Ϫ/Ϫ cells, with or without the Ca 2ϩ /calmodulin-dependent protein kinase kinase (CAMKK) inhibitor STO-609. Oxygen consumption was measured in L6 myotubes and isolated muscle mitochondria. The effect of a BBR derivative, dihydroberberine (dhBBR), on adiposity and glucose metabolism was examined in rodents fed a high-fat diet. RESULTS-We have made the following novel observations: 1) BBR dose-dependently inhibited respiration in L6 myotubes and muscle mitochondria, through a specific effect on respiratory complex I, similar to that observed with metformin and rosiglitazone; 2) activation of AMPK by BBR did not rely on the activity of either LKB1 or CAMKK␤, consistent with major regulation at the level of the AMPK phosphatase; and 3) a novel BBR derivative, dhBBR, was identified that displayed improved in vivo efficacy in terms of counteracting increased adiposity, tissue triglyceride accumulation, and insulin resistance in high-fat-fed rodents. This effect is likely due to enhanced oral bioavailability. CONCLUSIONS-Complex I of the respiratory chain represents a major target for compounds that improve whole-body insulin sensitivity through increased AMPK activity. The identification of a novel derivative of BBR with improved in vivo efficacy highlights the potential importance of BBR as a novel therapy for the treatment of type 2 diabetes.

A significant contributor to the rising rates of human obesity is an increase in energy intake. The ‘protein leverage hypothesis’ proposes that a dominant appetite for protein in conjunction with a decline in the ratio of protein to fat and carbohydrate in the diet drives excess energy intake and could therefore promote the development of obesity. Our aim was to test the ‘protein leverage hypothesis’ in lean humans by disguising the macronutrient composition of foods offered to subjects under ad libitum feeding conditions. Energy intakes and hunger ratings were measured for 22 lean subjects studied over three 4-day periods of in-house dietary manipulation. Subjects were restricted to fixed menus in random order comprising 28 foods designed to be similar in palatability, availability, variety and sensory quality and providing 10%, 15% or 25% energy as protein. Nutrient and energy intake was calculated as the product of the amount of each food eaten and its composition. Lowering the percent protein of the diet from 15% to 10% resulted in higher (+12±4.5%, p = 0.02) total energy intake, predominantly from savoury-flavoured foods available between meals. This increased energy intake was not sufficient to maintain protein intake constant, indicating that protein leverage is incomplete. Urinary urea on the 10% and 15% protein diets did not differ statistically, nor did they differ from habitual values prior to the study. In contrast, increasing protein from 15% to 25% did not alter energy intake. On the fourth day of the trial, however, there was a greater increase in the hunger score between 1–2 h after the 10% protein breakfast versus the 25% protein breakfast (1.6±0.4 vs 25%: 0.5±0.3, p = 0.005). In our study population a change in the nutritional environment that dilutes dietary protein with carbohydrate and fat promotes overconsumption, enhancing the risk for potential weight gain.

We apply nutritional geometry, a framework for modelling the interactive effects of nutrients on animals, to help understand the role of modern environments in the obesity pandemic. Evidence suggests that humans regulate the intake of protein energy (PE) more strongly than non-protein energy (nPE), and consequently will over-and under-ingest nPE on diets with low or high PE, respectively. This pattern of macronutrient regulation has led to the protein leverage hypothesis, which proposes that the rise in obesity has been caused partly by a shift towards diets with reduced PE:nPE ratios relative to the set point for protein regulation. We discuss potential causes of this mismatch, including environmentally induced reductions in the protein density of the human diet and factors that might increase the regulatory set point for protein and hence exacerbate protein leverage. Economics -the high price of protein compared with fats and carbohydrates -is one factor that might contribute to the reduction of dietary protein concentrations. The possibility that rising atmospheric CO 2 levels could also play a role through reducing the PE:nPE ratios in plants and animals in the human food chain is discussed. Factors that reduce protein efficiency, for example by increasing the use of ingested amino acids in energy metabolism (hepatic gluconeogenesis), are highlighted as potential drivers of increased set points for protein regulation. We recommend that a similar approach is taken to understand the rise of obesity in other species, and identify some key gaps in the understanding of nutrient regulation in companion animals.

2002;10:401-407. Objective: Obesity is a major risk factor for the development of type 2 diabetes. Tumor necrosis factor (TNF)-␣ is a candidate gene for the development of both obesity and insulin resistance. We investigated whether a common polymorphism in the promoter region (Ϫ308 G/A) of the TNF-␣ gene was associated with increased risk for the development of insulin resistance and cardiovascular disease in an obese Australian population. Research Methods and Procedures: Obese, non-diabetic subjects (146 women and 34 men) were genotyped with polymerase chain reaction-restriction fragment length polymorphism techniques, and anthropometric and biochemical measurements were analyzed. A homeostasis model assessment (HOMA) score was used to gauge the level of insulin resistance. Results: The frequencies of the G allele and the A allele were 0.759 and 0.241, respectively. Subjects homozygous for the A allele had higher fasting insulin levels (226 vs. 131 pM; p Ͻ 0.001), higher HOMA scores (10.2 vs. 5.3; p Ͻ 0.001), higher systolic blood pressure (143 vs. 129 mm Hg; p ϭ 0.02), and lower high-density lipoprotein (HDL) cholesterol (1.13 vs. 1.25 mM; p ϭ 0.04) than did subjects homozygous for the G allele. Whereas an association between insulin resistance and body mass index or waist circumference was seen in all subjects, a highly significant negative correlation of HDL cholesterol to HOMA scores (r ϭ Ϫ0.710; p Ͻ 0.001) occurred in subjects with the A allele only. Discussion: The Ϫ308 G/A TNF-␣ gene variant conveys an increased risk for the development of insulin resistance in obese subjects. The presence of low HDL cholesterol levels further increases the risks associated with insulin resistance in carriers of the A allele.

Increased energy intakes are contributing to overweight and obesity. Growing evidence supports the role of protein appetite in driving excess intake when dietary protein is diluted (the protein leverage hypothesis). Understanding the interactions between dietary macronutrient balance and nutrient-specific appetite systems will be required for designing dietary interventions that work with, rather than against, basic regulatory physiology. Data were collected from 38 published experimental trials measuring ad libitum intake in subjects confined to menus differing in macronutrient composition. Collectively, these trials encompassed considerable variation in percent protein (spanning 8-54% of total energy), carbohydrate (1.6-72%) and fat (11-66%). The data provide an opportunity to describe the individual and interactive effects of dietary protein, carbohydrate and fat on the control of total energy intake. Percent dietary protein was negatively associated with total energy intake (F = 6.9, P < 0.0001) irrespective of whether carbohydrate (F = 0, P = 0.7) or fat (F = 0, P = 0.5) were the diluents of protein. The analysis strongly supports a role for protein leverage in lean, overweight and obese humans. A better appreciation of the targets and regulatory priorities for protein, carbohydrate and fat intake will inform the design of effective and health-promoting weight loss diets, food labelling policies, food production systems and regulatory frameworks.

Objective: Protein leverage plays a role in driving increased energy intakes that may promote weight gain. The influence of the protein to carbohydrate ratio (P:C) in diets of C57BL/6J mice on total energy intake, fat storage, and thermogenesis was investigated. Design and Methods: Male mice (9 weeks old) were provided ad libitum access to one of five isocaloric diets that differed in P:C. Food intake was recorded for 12 weeks. After 16 weeks, white adipose tissue (WAT) and brown adipose tissue (BAT) deposits were dissected, weighed, and the expression levels of key metabolic regulators were determined in BAT. In a separate cohort, body surface temperature was measured in response to 25 diets differing in protein, fat, and carbohydrate content. Results: Mice on low P:C diets (9:72 and 17:64) had greater total energy intake and increased WAT and BAT stores. Body surface temperature increased with total energy intake and with protein, fat, and carbohydrate, making similar contributions per kJ ingested. Expression of three key regulators of thermogenesis were downregulated in BAT in mice on the lowest P:C diet. Conclusions: Low-protein diets induced sustained hyperphagia and a generalized expansion of fat stores. Increased body surface temperature on low P:C diets was consistent with diet-induced thermogenesis (DIT) as a means to dissipate excess ingested energy on such diets, although this was not sufficient to prevent development of increased adiposity. Whether BAT was involved in DIT is not clear. Increased BAT mass on low P:C diets might suggest so, but patterns of thermogenic gene expression do not support a role for BAT in DIT, although they might reflect failure of thermogenic function with prolonged exposure to a low P:C diet.Obesity (2013) 21, 85-92.

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