José Rodrigo Pauli

High-fat diet induces apoptosis of hypothalamic neurons.

Consumption of dietary fats is amongst the most important environmental factors leading to obesity. In rodents, the consumption of fat-rich diets blunts leptin and insulin anorexigenic signaling in the hypothalamus by a mechanism dependent on the in situ activation of inflammation. Since inflammatory signal transduction can lead to the activation of apoptotic signaling pathways, we evaluated the effect of high-fat feeding on the induction of apoptosis of hypothalamic cells. Here, we show that consumption of dietary fats induce apoptosis of neurons and a reduction of synaptic inputs in the arcuate nucleus and lateral hypothalamus. This effect is dependent upon diet composition, and not on caloric intake, since pair-feeding is not sufficient to reduce the expression of apoptotic markers. The presence of an intact TLR4 receptor, protects cells from further apoptotic signals. In diet-induced inflammation of the hypothalamus, TLR4 exerts a dual function, on one side activating pro-inflammatory pathways that play a central role in the development of resistance to leptin and insulin, and on the other side restraining further damage by controlling the apoptotic activity.

Unsaturated fatty acids revert diet-induced hypothalamic inflammation in obesity.

Background In experimental models, hypothalamic inflammation is an early and determining factor in the installation and progression of obesity. Pharmacological and gene-based approaches have proven efficient in restraining inflammation and correcting the obese phenotypes. However, the role of nutrients in the modulation of hypothalamic inflammation is unknown. Methodology/Principal Findings Here we show that, in a mouse model of diet-induced obesity, partial substitution of the fatty acid component of the diet by flax seed oil (rich in C18:3) or olive oil (rich in C18:1) corrects hypothalamic inflammation, hypothalamic and whole body insulin resistance, and body adiposity. In addition, upon icv injection in obese rats, both ω3 and ω9 pure fatty acids reduce spontaneous food intake and body mass gain. These effects are accompanied by the reversal of functional and molecular hypothalamic resistance to leptin/insulin and increased POMC and CART expressions. In addition, both, ω3 and ω9 fatty acids inhibit the AMPK/ACC pathway and increase CPT1 and SCD1 expression in the hypothalamus. Finally, acute hypothalamic injection of ω3 and ω9 fatty acids activate signal transduction through the recently identified GPR120 unsaturated fatty acid receptor. Conclusions/Significance Unsaturated fatty acids can act either as nutrients or directly in the hypothalamus, reverting diet-induced inflammation and reducing body adiposity. These data show that, in addition to pharmacological and genetic approaches, nutrients can also be attractive candidates for controlling hypothalamic inflammation in obesity.

IL-6 and IL-10 Anti-Inflammatory Activity Links Exercise to Hypothalamic Insulin and Leptin Sensitivity through IKKβ and ER Stress Inhibition

Physical activity confers beneficial metabolic effects by inducing anti-inflammatory activity in the hypothalamus region of the brain in rodents, resulting in a reorganization of the set point of nutritional balance and reduced insulin and leptin resistance.

A Central Role for Neuronal AMP-Activated Protein Kinase (AMPK) and Mammalian Target of Rapamycin (mTOR) in High-Protein Diet–Induced Weight Loss

OBJECTIVE—A high-protein diet (HPD) is known to promote the reduction of body fat, but the mechanisms underlying this change are unclear. AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) function as majors regulators of cellular metabolism that respond to changes in energy status, and recent data demonstrated that they also play a critical role in systemic energy balance. Here, we sought to determine whether the response of the AMPK and mTOR pathways could contribute to the molecular effects of an HPD. RESEARCH DESIGN AND METHODS—Western blotting, confocal microscopy, chromatography, light microscopy, and RT-PCR assays were combined to explore the anorexigenic effects of an HPD. RESULTS—An HPD reduced food intake and induced weight loss in both normal rats and ob/ob mice. The intracerebroventricular administration of leucine reduced food intake, and the magnitude of weight loss and reduction of food intake in a leucine-supplemented diet are similar to that achieved by HPD in normal rats and in ob/ob mice, suggesting that leucine is a major component of the effects of an HPD. Leucine and HPD decrease AMPK and increase mTOR activity in the hypothalamus, leading to inhibition of neuropeptide Y and stimulation of pro-opiomelanocortin expression. Consistent with a cross-regulation between AMPK and mTOR to control food intake, our data show that the activation of these enzymes occurs in the same specific neuronal subtypes. CONCLUSIONS—These findings provide support for the hypothesis that AMPK and mTOR interact in the hypothalamus to regulate feeding during HPD in a leucine-dependent manner.

Reversal of diet-induced insulin resistance with a single bout of exercise in the rat: the role of PTP1B and IRS-1 serine phosphorylation

Lifestyle interventions including exercise programmes are cornerstones in the prevention of obesity‐related diabetes. In this study, we demonstrate that a single bout of exercise inhibits high‐fat diet‐induced insulin resistance. Diet‐induced obesity (DIO) increased the expression and activity of the protein tyrosine phosphatase 1B (PTP1B) and attenuated insulin signalling in gastrocnemius muscle of rats, a phenomenon which was reversed by a single session of exercise. In addition, DIO was observed to lead to serine phosphorylation of insulin receptor substrate 1 (IRS‐1), which was also reversed by exercise in muscle in parallel with a reduction in c‐Jun N‐terminal kinase (JNK) activity. Thus, acute exercise increased the insulin sensitivity during high‐fat feeding in obese rats. Overall, these results provide new insights into the mechanism by which exercise restores insulin sensitivity.

Interleukin-10 is a protective factor against diet-induced insulin resistance in liver☆

BACKGROUND/AIMS The anti-inflammatory cytokine, interleukin-10 (IL-10), is known to exert a protective role in hepatic damage caused by viruses, alcohol, autoimmunity and a number of experimental aggressors. Recently, a protective role for IL-10 has also been proposed in diet-induced hepatic dysfunction. However, studies about the mechanisms involved in this process are controversial. The objective of this study was to evaluate the role of endogenous IL-10 in the development of hepatic insulin resistance, associated with diet-induced fatty liver disease. METHODS Male Swiss mice treated for eight weeks with a high-fat diet became diabetic and developed non-alcoholic fatty liver disease, which is characterized by increased hepatic fat deposition and liver infiltration by F4/80 positive cells. This was accompanied by an increased hepatic expression of TNF-alpha, IL-6, IL-1beta and IL-10, and by an impaired insulin signal transduction through the insulin receptor/IRS1-IRS2/PI3-kinase/Akt/FOXO1 signaling pathway. RESULTS Upon endogenous IL-10 inhibition for 5 days, using two distinct methods, a neutralizing anti-IL-10 antibody and an antisense oligonucleotide against IL-10, increased hepatic expression of the inflammatory markers TNF-alpha, IL-6, IL-1beta and F4/80 was observed. This was accompanied by a significant negative modulation of insulin signal transduction through insulin receptor/IRS1-IRS2/PI3-kinase/Akt/FOXO1, and by the stimulation of hepatic signaling proteins involved in gluconeogenesis and lipid synthesis. CONCLUSIONS Thus, in an animal model of diet-induced fatty liver disease, the inhibition of IL-10 promotes the increased expression of inflammatory cytokines, the worsening of insulin signaling and the activation of gluconeogenic and lipidogenic pathways.

Inhibition of toll-like receptor 2 expression improves insulin sensitivity and signaling in muscle and white adipose tissue of mice fed a high-fat diet

The aims of the present study were to investigate the expression of toll-like receptor 2 (TLR2) in muscle and white adipose tissue (WAT) of diet-induced obesity (DIO) mice, and also the effects of its inhibition, with the use of TLR2 antisense oligonucleotide (ASON), on insulin sensitivity and signaling. The expression of TLR2 was increased in muscle and WAT of DIO mice, compared with those that received standard chow. Inhibition of TLR2 in DIO mice, by TLR2 ASON, improved insulin sensitivity and signaling in muscle and WAT. In addition, data show that the inhibition of TLR2 expression prevents the activation of IKBKB, MAPK8, and serine phosphorylation of IRS1 in DIO mice, suggesting that TLR2 is a key modulator of the crosstalk between inflammatory and metabolic pathways. We, therefore, suggest that a selective interference with TLR2 presents an attractive opportunity for the treatment of insulin resistance in obesity and type 2 diabetes.

Physical exercise reduces circulating lipopolysaccharide and TLR4 activation and improves insulin signaling in tissues of DIO rats.

OBJECTIVE Insulin resistance in diet-induced obesity (DIO) is associated with a chronic systemic low-grade inflammation, and Toll–like receptor 4 (TLR4) plays an important role in the link among insulin resistance, inflammation, and obesity. The current study aimed to analyze the effect of exercise on TLR4 expression and activation in obese rats and its consequences on insulin sensitivity and signaling. RESEARCH DESIGN AND METHODS The effect of chronic and acute exercise was investigated on insulin sensitivity, insulin signaling, TLR4 activation, c-Jun NH2-terminal kinase (JNK) and IκB kinase (IKKβ) activity, and lipopolysaccharide (LPS) serum levels in tissues of DIO rats. RESULTS The results showed that chronic exercise reduced TLR4 mRNA and protein expression in liver, muscle, and adipose tissue. However, both acute and chronic exercise blunted TLR4 signaling in these tissues, including a reduction in JNK and IKKβ phosphorylation and IRS-1 serine 307 phosphorylation, and, in parallel, improved insulin-induced IR, IRS-1 tyrosine phosphorylation, and Akt serine phosphorylation, and reduced LPS serum levels. CONCLUSIONS Our results show that physical exercise in DIO rats, both acute and chronic, induces an important suppression in the TLR4 signaling pathway in the liver, muscle, and adipose tissue, reduces LPS serum levels, and improves insulin signaling and sensitivity. These data provide considerable progress in our understanding of the molecular events that link physical exercise to an improvement in inflammation and insulin resistance.

Acute physical exercise reverses S-nitrosation of the insulin receptor, insulin receptor substrate 1 and protein kinase B/Akt in diet-induced obese Wistar rats

Early evidence demonstrates that exogenous nitric oxide (NO) and the NO produced by inducible nitric oxide synthase (iNOS) can induce insulin resistance. Here, we investigated whether this insulin resistance, mediated by S‐nitrosation of proteins involved in early steps of the insulin signal transduction pathway, could be reversed by acute physical exercise. Rats on a high‐fat diet were subjected to swimming for two 3 h‐long bouts, separated by a 45 min rest period. Two or 16 h after the exercise protocol the rats were killed and proteins from the insulin signalling pathway were analysed by immunoprecipitation and immunoblotting. We demonstrated that a high‐fat diet led to an increase in the iNOS protein level and S‐nitrosation of insulin receptor β (IRβ), insulin receptor substrate 1 (IRS1) and Akt. Interestingly, an acute bout of exercise reduced iNOS expression and S‐nitrosation of proteins involved in the early steps of insulin action, and improved insulin sensitivity in diet‐induced obesity rats. Furthermore, administration of GSNO (NO donor) prevents this improvement in insulin action and the use of an inhibitor of iNOS (l‐N6‐(1‐iminoethyl)lysine; l‐NIL) simulates the effects of exercise on insulin action, insulin signalling and S‐nitrosation of IRβ, IRS1 and Akt. In summary, a single bout of exercise reverses insulin sensitivity in diet‐induced obese rats by improving the insulin signalling pathway, in parallel with a decrease in iNOS expression and in the S‐nitrosation of IR/IRS1/Akt. The decrease in iNOS protein expression in the muscle of diet‐induced obese rats after an acute bout of exercise was accompanied by an increase in AMP‐activated protein kinase (AMPK) activity. These results provide new insights into the mechanism by which exercise restores insulin sensitivity.

Inhibition of UCP2 expression reverses diet-induced diabetes mellitus by effects on both insulin secretion and action

Recent characterization of the ability of uncoupling protein 2 (UCP2) to reduce ATP production and inhibit insulin secretion by pancreatic β‐cells has placed this mitochondrial protein as a candidate target for therapeutics in diabetes mellitus. In the present study we evaluate the effects of short‐term treatment of two animal models of type 2 diabetes mellitus with an antisense oligonucleotide to UCP2. In both models, Swiss mice (made obese and diabetic by a hyperlipidic diet) and ob/ob mice, the treatment resulted in a significant improvement in the hyperglyce‐mic syndrome. This effect was due not only to an improvement of insulin secretion, but also to improved peripheral insulin action. In isolated pancreatic islets, the partial inhibition of UCP2 increased ATP content, followed by increased glucose‐stimulated insulin secretion. This was not accompanied by increased expression of enzymes involved in protection against oxida‐tive stress. The evaluation of insulin action in peripheral tissues revealed that the inhibition of UCP2 expression significantly improved insulin signal trans‐duction in adipose tissue. In conclusion, short‐term inhibition of UCP2 expression ameliorates the hyper‐glycemic syndrome in two distinct animal models of obesity and diabetes. Metabolic improvement is due to a combined effect on insulin‐producing pancreatic islets and in at least one peripheral tissue that acts as a target for insulin.—De Souza, C. T., Araújo, E. P., Stoppiglia, L. F., Pauli, J. R., Ropelle, E., Rocco, S. A., Marin, R. M., Franchini, K. G., Carvalheira, J. B., Saad, M. J., Boschero, A. C., Carneiro, E. M., Velloso, L. A. Inhibition of UCP2 expression reverses diet‐induced diabetes mellitus by effects on both insulin secretion and action. FASEB J. 21, 1153–1163 (2007)