Marco Antonio Carvalho-Filho

Loss-of-Function Mutation in Toll-Like Receptor 4 Prevents Diet-Induced Obesity and Insulin Resistance

Obesity is associated with insulin resistance and a state of abnormal inflammatory response. The Toll-like receptor (TLR)4 has an important role in inflammation and immunity, and its expression has been reported in most tissues of the body, including the insulin-sensitive ones. Because it is activated by lipopolysaccharide and saturated fatty acids, which are inducers of insulin resistance, TLR4 may be a candidate for participation in the cross-talk between inflammatory and metabolic signals. Here, we show that C3H/HeJ mice, which have a loss-of-function mutation in TLR4, are protected against the development of diet-induced obesity. In addition, these mice demonstrate decreased adiposity, increased oxygen consumption, a decreased respiratory exchange ratio, improved insulin sensitivity, and enhanced insulin-signaling capacity in adipose tissue, muscle, and liver compared with control mice during high-fat feeding. Moreover, in these tissues, control mice fed a high-fat diet show an increase in IκB kinase complex and c-Jun NH2-terminal kinase activity, which is prevented in C3H/HeJ mice. In isolated muscles from C3H/HeJ mice, protection from saturated fatty acid–induced insulin resistance is observed. Thus, TLR4 appears to be an important mediator of obesity and insulin resistance and a potential target for the therapy of these highly prevalent medical conditions.

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.

Insulin signalling pathways in aorta and muscle from two animal models of insulin resistance—the obese middle-aged and the spontaneously hypertensive rats

Aims/hypothesisThe aim of this study was to investigate insulin signalling pathways directly in vivo in skeletal muscle and thoracic aorta from obese middle-aged (12-month-old) rats, which have insulin resistance but not cardiovascular disease, and from spontaneously hypertensive rats (SHR), an experimental model of insulin resistance and cardiovascular disease.MethodsWe have used in vivo insulin infusion, followed by tissue extraction, immunoprecipitation and immunoblotting.ResultsObese middle-aged rats and the SHR showed marked insulin resistance, which parallels the reduced effects of this hormone in the insulin signalling cascade in muscle. In aortae from obese middle-aged rats, the PI 3-kinase/Akt pathway is preserved, leading to a normal activation of endothelial nitric oxide synthase. In SHR this pathway is severely blunted, with reductions in eNOS protein concentration and activation. Both animals, however, showed higher concentrations and higher tyrosine phosphorylation of mitogen-activated protein (MAP) kinase isoforms in aortae.Conclusions/interpretationAlterations in the IRS/PI 3-K/Akt pathway in muscle of 12-month-old rats and SHR could be involved in the insulin resistance of these animals. The preservation of this pathway in aorta of 12-month-old rats, apart from increases in MAP kinase protein concentration and activation, could be a factor that contributes to explaining the absence of cardiovascular disease in this animal model. However, in aortae of SHR, the reduced insulin signalling through IRS/PI 3-kinase/Akt/eNOS pathway could contribute to the endothelial dysfunction of this animal.

Targeted Disruption of Inducible Nitric Oxide Synthase Protects Against Aging, S -Nitrosation, and Insulin Resistance in Muscle of Male Mice

Accumulating evidence has demonstrated that S-nitrosation of proteins plays a critical role in several human diseases. Here, we explored the role of inducible nitric oxide synthase (iNOS) in the S-nitrosation of proteins involved in the early steps of the insulin-signaling pathway and insulin resistance in the skeletal muscle of aged mice. Aging increased iNOS expression and S-nitrosation of major proteins involved in insulin signaling, thereby reducing insulin sensitivity in skeletal muscle. Conversely, aged iNOS-null mice were protected from S-nitrosation–induced insulin resistance. Moreover, pharmacological treatment with an iNOS inhibitor and acute exercise reduced iNOS-induced S-nitrosation and increased insulin sensitivity in the muscle of aged animals. These findings indicate that the insulin resistance observed in aged mice is mainly mediated through the S-nitrosation of the insulin-signaling pathway.

Double-Stranded RNA-Activated Protein Kinase Is a Key Modulator of Insulin Sensitivity in Physiological Conditions and in Obesity in Mice

The molecular integration of nutrient- and pathogen-sensing pathways has become of great interest in understanding the mechanisms of insulin resistance in obesity. The double-stranded RNA-dependent protein kinase (PKR) is one candidate molecule that may provide cross talk between inflammatory and metabolic signaling. The present study was performed to determine, first, the role of PKR in modulating insulin action and glucose metabolism in physiological situations, and second, the role of PKR in insulin resistance in obese mice. We used Pkr(-/-) and Pkr(+/+) mice to investigate the role of PKR in modulating insulin sensitivity, glucose metabolism, and insulin signaling in liver, muscle, and adipose tissue in response to a high-fat diet. Our data show that in lean Pkr(-/-) mice, there is an improvement in insulin sensitivity, and in glucose tolerance, and a reduction in fasting blood glucose, probably related to a decrease in protein phosphatase 2A activity and a parallel increase in insulin-induced thymoma viral oncogene-1 (Akt) phosphorylation. PKR is activated in tissues of obese mice and can induce insulin resistance by directly binding to and inducing insulin receptor substrate (IRS)-1 serine307 phosphorylation or indirectly through modulation of c-Jun N-terminal kinase and inhibitor of κB kinase β. Pkr(-/-) mice were protected from high-fat diet-induced insulin resistance and glucose intolerance and showed improved insulin signaling associated with a reduction in c-Jun N-terminal kinase and inhibitor of κB kinase β phosphorylation in insulin-sensitive tissues. PKR may have a role in insulin sensitivity under normal physiological conditions, probably by modulating protein phosphatase 2A activity and serine-threonine kinase phosphorylation, and certainly, this kinase may represent a central mechanism for the integration of pathogen response and innate immunity with insulin action and metabolic pathways that are critical in obesity.

Translational research into gut microbiota: new horizons in obesity treatment

1 , Bruno M. Carvalho 1 , AbstrAct Obesity is a pandemic which has been rapidly developing for three decades. When a popula- tion is submitted to the same nutritional stress, some individuals are less susceptible to diet- induced weight gain and hyperglycemia. This observation suggests that other mechanisms are involved which are not directly related to the human genome. The human gut contains an immense number of microorganisms, collectively known as the microbiota. Evidence that gut microbiota composition can differ between obese and lean humans has led to the speculation that gut microbiota can participate in the pathophysiology of obesity. Different mechanisms have been proposed to explain the link between gut flora and obesity. The first mechanism consists in the role of the gut microbiota to increase energy extraction from indigestible dietary polysaccharides. The second, consists in the role of gut flora to modulate plasma lipopolysac- charide levels which triggers chronic low-grade inflammation leading to obesity and diabetes. A third mechanism proposes that gut microbiota may induce regulation of host genes that modulate how energy is expended and stored. However, further studies are needed to clarify a number of issues related to the relationship between the gut microbiota and obesity.

The Impact of Simulated Medical Consultations on the Empathy Levels of Students at One Medical School

Purpose To examine the impact of simulated medical consultations using standardized patients (SPs) on the empathy levels of fourth- and sixth-year students at the Unicamp medical school in Brazil. Method Throughout 2011 and 2012, the authors conducted this study with two classes of fourth-year (n = 124) and two classes of sixth-year (n = 123) medical students. Students completed the medical student version of the Jefferson Scale of Physician Empathy before and after simulated medical consultations with SPs, followed by an in-depth debriefing dealing with the feelings of the patient about the disease, such as fear, guilt, anger, and abandonment; the feelings of the doctor towards the patient; and other topics as they arose. Results The simulation activity increased the empathy scores of the fourth-year students (from 115.8 to 121.1, P < .001, effect size = 0.61) and of the sixth-year students (from 117.1 to 123.5, P < .001, effect size = 0.64). Conclusions Although the study results were obtained via self-report—a limitation—they suggest that the effective simulation of medical consultations with SPs may improve medical students’ empathy levels. One unexpected result was that this activity, during the debriefing, became a forum for debating topics such as the doctor–patient relationship, the hidden curriculum, negative role models, and emotionally significant experiences of students in medical school. This kind of activity in itself may influence young doctors to become more empathetic and compassionate with their patients and foster a more meaningful way of practicing medicine.

Modulation of double-stranded RNA-activated protein kinase in insulin sensitive tissues of obese humans

Objective: The double‐stranded RNA‐dependent protein kinase (PKR) was recently implicated in regulating molecular integration of nutrient‐ and pathogen‐sensing pathways in obese mice. However, its modulation in human tissues in situations of insulin resistance has not been investigated. The present study was performed to first determine the tissue expression and phosphorylation levels of PKR in the liver, muscle, and adipose tissue in obese humans, and also the modulation of this protein in the adipose tissue of obese patients after bariatric surgery.

[RETRACTION]Translational research into gut microbiota: new horizons in obesity treatment

Obesity is a pandemic which has been rapidly developing for three decades. When a population is submitted to the same nutritional stress, some individuals are less susceptible to diet-induced weight gain and hyperglycemia. This observation suggests that other mechanisms are involved which are not directly related to the human genome. The human gut contains an immense number of microorganisms, collectively known as the microbiota. Evidence that gut microbiota composition can differ between obese and lean humans has led to the speculation that gut microbiota can participate in the pathophysiology of obesity. Different mechanisms have been proposed to explain the link between gut flora and obesity. The first mechanism consists in the role of the gut microbiota to increase energy extraction from indigestible dietary polysaccharides. The second, consists in the role of gut flora to modulate plasma lipopolysaccharide levels which triggers chronic low-grade inflammation leading to obesity and diabetes. A third mechanism proposes that gut microbiota may induce regulation of host genes that modulate how energy is expended and stored. However, further studies are needed to clarify a number of issues related to the relationship between the gut microbiota and obesity.

Profile of the Population Cared for in a Referral Emergency Unit

El objetivo del estudio fue identificar el perfil sociodemografico y las principales quejas de la poblacion adulta atendida en la Unidad de Emergencia de Referencia (UER). El conocimiento de las caracteristicas de la poblacion que frecuenta un servicio de emergencia constituye una herramienta de planificacion de las acciones en salud. Se trata de un estudio descriptivo/ retrospectivo desarrollado en la UER de un hospital de ensenanza en el interior del estado de Sao Paulo. La muestra fue constituida por los boletines de atencion (BAU) del periodo de enero a diciembre de 2.008. El instrumento utilizado fue estructurado con base en los datos del BAU. La busqueda espontanea fue hecha por mujeres jovenes (14 a 54 anos) de los barrios proximos a la UER durante la semana y en el horario de las siete a las diecinueve horas. Las quejas mas frecuentes fueron: cefalea, lumbago, dolores (abdominal, toracica). Se concluye que el perfil de la muestra fue en su mayoria adulto y joven, en edad productiva, de sexo femenino, con demanda espontanea y en el periodo diurno en dias utiles. Las atenciones apuntaron para quejas de baja complejidad.El objetivo del estudio fue identificar el perfil sociodemografico y las principales quejas de la poblacion adulta atendida en la Unidad de Emergencia de Referencia (UER). El conocimiento de las caracteristicas de la poblacion que frecuenta un servicio de emergencia constituye una herramienta de planificacion de las acciones en salud. Se trata de un estudio descriptivo/retrospectivo desarrollado en la UER de un hospital de ensenanza en el interior del estado de Sao Paulo. La muestra fue constituida por los boletines de atencion(BAU) del periodo de enero a diciembre de 2.008. El instrumento utilizado fue estructurado con base en los datos del BAU. La busqueda espontanea fue hecha por mujeres jovenes (14 a 54 anos) de los barrios proximos a la UER durante la semana y en el horario de las siete a las diecinueve horas. Las quejas mas frecuentes fueron: cefalea, lumbago, dolores (abdominal, toracica). Se concluye que el perfil de la muestra fue en su mayoria adulto y joven, en edad productiva, de sexo femenino, con demanda espontanea y en el periodo diurno en dias utiles. Las atenciones apuntaron para quejas de baja complejidad.