Javier Campión

Individuality and epigenetics in obesity.

Excessive weight gain arises from the interactions among environmental factors, genetic predisposition and the individual behavior. However, it is becoming evident that interindividual differences in obesity susceptibility depend also on epigenetic factors. Epigenetics studies the heritable changes in gene expression that do not involve changes to the underlying DNA sequence. These processes include DNA methylation, covalent histone modifications, chromatin folding and, more recently described, the regulatory action of miRNAs and polycomb group complexes. In this review, we focus on experimental evidences concerning dietary factors influencing obesity development by epigenetic mechanisms, reporting treatment doses and durations. Moreover, we present a bioinformatic analysis of promoter regions for the search of future epigenetic biomarkers of obesity, including methylation pattern analyses of several obesity‐related genes (epiobesigenes), such as FGF2, PTEN, CDKN1A and ESR1, implicated in adipogenesis, SOCS1/SOCS3, in inflammation, and COX7A1 LPL, CAV1, and IGFBP3, in intermediate metabolism and insulin signalling. The identification of those individuals that at an early age could present changes in the methylation profiles of specific genes could help to predict their susceptibility to later develop obesity, which may allow to prevent and follow‐up its progress, as well as to research and develop newer therapeutic approaches.

Weight Gain Induced by High-Fat Feeding Involves Increased Liver Oxidative Stress

Objective: To assess the effects of high‐fat feeding on white adipose tissue gene expression and liver oxidative stress.

High fat diet-induced obesity modifies the methylation pattern of leptin promoter in rats.

Leptin is an adipokine involved in body weight and food intake regulation whose promoter region presents CpG islands that could be subject to dynamic methylation. This methylation process could be affected by environmental (e.g. diet) or endogenous (e.g., adipocyte differentiation, inflammation, hypoxia) factors, and could influence adipocyte leptin gene expression. The aim of this article was to study whether a high-energy diet may affect leptin gene promoter methylation in rats. A group of eleven male Wistar rats were assigned into two dietary groups, one fed on a control diet for 11 weeks and the other on a high-fat cafeteria diet. Rats fed a high-energy diet become overweight and hyperleptin emic as compared to the controls. DNA isolated from retroperitoneal adipocytes was treated with bisulfite and a distal portion of leptin promoter (from −694 to −372 bp) including 13 CpG sites was amplified by PCR and sequenced. The studied promoter portion was slightly more methylated in the cafeteria-fed animals, which was statistically significant (p<0.05) for one of the CpG sites (located at the position −443). In obese rats, such methy lation was associated to lower circulating leptin levels, suggesting that this position could be important in the regulation of leptin gene expression, probably by being a target sequence of different transcription factors. Our findings reveal, for the first time, that leptin methylation pattern can be influenced by diet-induced obesity, and suggest that epigenetic mechanisms could be involved in obesity by regulating the expression of important epiobesigenic genes.ResumenLa leptina es una adipoquina implicada en la regulación del peso corporal y la ingesta energética cuya región promotora presenta islas CpG que podrían ser metiladas dinámicamente. Este proceso de metilación podría verse afectado por factores ambientales, como la dieta, o endógenos, como la diferenciación adipocitaria, inflamación o hipoxia, y podría influir en la expresión de leptina por parte de los adipocitos. El objetivo de este artículo es estudiar si una dieta alta en grasa podría afectar a la metilación del promotor de la leptina en ratas. Un grupo de once ratas Wistar macho fue dividido en dos subgrupos, uno alimentado con dieta control durante 11 semanas y el otro con dieta alta en grasa (dieta de cafetería). Las ratas alimentadas con la dieta rica en grasa presentaron sobrepeso e hiperleptinemia. El ADN aislado de los adipocitos retroperitoneales fue tratado con bisulfito y una porción distal del promotor de la leptina (de la base-694 a la — 372), conteniendo 13 sitios CpG, fue amplificada por PCR y secuenciada. Esta región del promotor apareció ligeramente más metilada en los animales alimentados con dieta de cafetería, lo cuál fue especialmente significativo (p <0,05) para uno de los sitios CpG (en la posición-443). En las ratas obesas, la metilación se asoció a una disminución de los niveles de leptina circulante, lo que sugiere que esta posición podría ser importante en la regulación de la expresión génica de esta adipoquina, probablemente por ser una secuencia diana de diferentes factores de trnascripción. Nuestos resultados, por primera vez, ponen de manifiesto que el patrón de metilación del promotor de la leptina puede estar influido por la obesidad inducida por la dieta, y sugieren que los mecanismos epigenéticos podrían estar implicados en la reciente pandemia de obesidad mediante la regulación de la expresión de importantes genes epiobesigénicos.

A dual epigenomic approach for the search of obesity biomarkers: DNA methylation in relation to diet-induced weight loss

Epigenetics could help to explain individual differences in weight loss after an energy‐restriction intervention. Here, we identify novel potential epigenetic biomarkers of weight loss, comparing DNA methylation patterns of high and low responders to a hypocaloric diet. Twenty‐five overweight or obese men participated in an 8‐wk caloric restriction intervention. DNA was isolated from peripheral blood mononuclear cells and treated with bisulfite. The basal and endpoint epigenetic differences between high and low responders were analyzed by methylation microarray, which was also useful in comparing epigenetic changes due to the nutrition intervention. Subsequently, MALDI‐TOF mass spectrometry was used to validate several relevant CpGs and the surrounding regions. DNA methylation levels in several CpGs located in the ATP10A and CD44 genes showed statistical baseline differences depending on the weight‐loss outcome. At the treatment endpoint, DNA methylation levels of several CpGs on the WT1 promoter were statistically more methylated in the high than in the low responders. Finally, different CpG sites from WT1 and ATP10A were significantly modified as a result of the intervention. In summary, hypocaloricdiet‐induced weight loss in humans could alter DNA methylation status of specific genes. Moreover, baseline DNA methylation patterns may be used as epigenetic markers that could help to predict weight loss.—Milagro, F. I., Campion, J., Cordero, P., Goyenechea, E., Gómez‐Uriz, A. M., Abete, I., Zulet, M. A., Martínez, J. A. A dual epigenomic approach for the search of obesity biomarkers: DNA methylation in relation to diet‐induced weight loss. FASEB J. 25, 1378–1389 (2011). www.fasebj.org

Antidiabetic effects of natural plant extracts via inhibition of carbohydrate hydrolysis enzymes with emphasis on pancreatic alpha amylase

Introduction: The increasing prevalence of type 2 diabetes mellitus and the negative clinical outcomes observed with the commercially available anti-diabetic drugs have led to the investigation of new therapeutic approaches focused on controlling postprandrial glucose levels. The use of carbohydrate digestive enzyme inhibitors from natural resources could be a possible strategy to block dietary carbohydrate absorption with less adverse effects than synthetic drugs. Areas covered: This review covers the latest evidence regarding in vitro and in vivo studies in relation to pancreatic alpha-amylase inhibitors of plant origin, and presents bioactive compounds of phenolic nature that exhibit anti-amylase activity. Expert opinion: Pancreatic alpha-amylase inhibitors from traditional plant extracts are a promising tool for diabetes treatment. Many studies have confirmed the alpha-amylase inhibitory activity of plants and their bioactive compounds in vitro, but few studies corroborate these findings in rodents and very few in humans. Thus, despite some encouraging results, more research is required for developing a valuable anti-diabetic therapy using pancreatic alpha-amylase inhibitors of plant origin.

Natural Inhibitors of Pancreatic Lipase as New Players in Obesity Treatment

Obesity is a multifactorial disease characterized by an excessive weight for height due to an enlarged fat deposition such as adipose tissue, which is attributed to a higher calorie intake than the energy expenditure. The key strategy to combat obesity is to prevent chronic positive impairments in the energy equation. However, it is often difficult to maintain energy balance, because many available foods are high-energy yielding, which is usually accompanied by low levels of physical activity. The pharmaceutical industry has invested many efforts in producing antiobesity drugs; but only a lipid digestion inhibitor obtained from an actinobacterium is currently approved and authorized in Europe for obesity treatment. This compound inhibits the activity of pancreatic lipase, which is one of the enzymes involved in fat digestion. In a similar way, hundreds of extracts are currently being isolated from plants, fungi, algae, or bacteria and screened for their potential inhibition of pancreatic lipase activity. Among them, extracts isolated from common foodstuffs such as tea, soybean, ginseng, yerba mate, peanut, apple, or grapevine have been reported. Some of them are polyphenols and saponins with an inhibitory effect on pancreatic lipase activity, which could be applied in the management of the obesity epidemic.

CLOCK, PER2 and BMAL1 DNA methylation: association with obesity and metabolic syndrome characteristics and monounsaturated fat intake.

The circadian clock system instructs 24-h rhythmicity on gene expression in essentially all cells, including adipocytes, and epigenetic mechanisms may participate in this regulation. The aim of this research was to investigate the influence of obesity and metabolic syndrome (MetS) features in clock gene methylation and the involvement of these epigenetic modifications in the outcomes. Sixty normal-weight, overweight and obese women followed a 16-weeks weight reduction program. DNA methylation levels at different CpG sites of CLOCK, BMAL1 and PER2 genes were analyzed by Sequenoms MassARRAY in white blood cells obtained before the treatment. Statistical differences between normal-weight and overweight + obese subjects were found in the methylation status of different CpG sites of CLOCK (CpGs 1, 5-6, 8 and 11-14) and, with lower statistical significance, in BMAL1 (CpGs 6-7, 8, 15 and 16-17). The methylation pattern of different CpG sites of the three genes showed significant associations with anthropometric parameters such as body mass index and adiposity, and with a MetS score. Moreover, the baseline methylation levels of CLOCK CpG 1 and PER2 CpGs 2-3 and 25 correlated with the magnitude of weight loss. Interestingly, the percentage of methylation of CLOCK CpGs 1 and 8 showed associations with the intake of monounsaturated and polyunsaturated fatty acids. This study demonstrates for the first time an association between methylation status of CpG sites located in clock genes (CLOCK, BMAL1 and PER2) with obesity, MetS and weight loss. Moreover, the methylation status of different CpG sites in CLOCK and PER2 could be used as biomarkers of weight-loss success, particularly CLOCK CPGs 5-6. (Author correspondence: garaulet@um.es)

TNF‐α Promoter Methylation as a Predictive Biomarker for Weight‐loss Response

Tumor necrosis factor‐α (TNF‐α) is a proinflammatory cytokine which is commonly elevated in obese subjects and whose promoter is susceptible to be regulated by cytosine methylation. The aim of this research was to analyze whether epigenetic regulation of human TNF‐α promoter by cytosine methylation could be involved in the predisposition to lose body weight after following a balanced hypocaloric diet. Twenty‐four patients (12 women/12 men) with excessive body weight‐for‐height (BMI: 30.5 ± 0.32 kg/m2; age: 34 ± 4 years old) followed an 8‐week energy‐restricted diet. Blood mononuclear cell DNA, isolated before the nutritional intervention, was treated with bisulfite and a region of TNF‐α gene promoter (from −360 to +50 bp) was sequenced. Obese men with successful weight loss (≥5% of initial body weight) showed lower levels of total TNF‐α promoter methylation (r = 0.74; P = 0.021), especially in the positions −170 bp (r = 0.75, P = 0.005) and −120 bp (r = 0.70, P = 0.011). Baseline TNF‐α circulating levels were positively associated with total promoter methylation (r = 0.84, P = 0.005) and methylation at position −245 bp (r = 0.75, P = 0.020). TNF‐α promoter methylation could be a good inflammation marker predicting the hypocaloric diet‐induced weight‐loss, and constitutes a first step toward personalized nutrition based on epigenetic criteria.

Differential DNA methylation patterns between high and low responders to a weight loss intervention in overweight or obese adolescents: the EVASYON study

In recent years, epigenetic markers emerged as a new tool to understand the influence of lifestyle factors on obesity phenotypes. Adolescence is considered an important epigenetic window over a humans lifetime. The objective of this work was to explore baseline changes in DNA methylation that could be associated with a better weight loss response after a multidisciplinary intervention program in Spanish obese or overweight adolescents. Overweight or obese adolescents (n=107) undergoing 10 wk of a multidisciplinary intervention for weight loss were assigned as high or low responders to the treatment. A methylation microarray was performed to search for baseline epigenetic differences between the 2 groups (12 subjects/group), and MALDI‐TOF mass spectrometry was used to validate (n=107) relevant CpG sites and surrounding regions. After validation, 5 regions located in or near AQP9, DUSP22, HIPK3, TNNT1, and TNNI3 genes showed differential methylation levels between high and low responders to the multidisciplinary weight loss intervention. Moreover, a calculated methylation score was significantly associated with changes in weight, BMI‐SDS, and body fat mass loss after the treatment. In summary, we have identified 5 DNA regions that are differentially methylated depending on weight loss response. These methylation changes may help to better understand the weight loss response in obese adolescents.—Moleres, A., Campión, J., Milagro, F. I., Marcos, A., Campoy, C., Garagorri, J. M., Gómez‐Martínez, S., Martínez, J. A., Azcona‐Sanjulián, M. C., Martí, A. Differential DNA methylation patterns between high and low responders to a weight loss intervention in overweight or obese adolescents: the EVASYON study. FASEB J. 27, 2504–2512 (2013). www.fasebj.org

Epigenetics and Obesity

The etiology of obesity is multifactorial, involving complex interactions among the genetic makeup, neuroendocrine status, fetal programming, and different unhealthy environmental factors, such as sedentarism or inadequate dietary habits. Among the different mechanisms causing obesity, epigenetics, defined as the study of heritable changes in gene expression that occur without a change in the DNA sequence, has emerged as a very important determinant. Experimental evidence concerning dietary factors influencing obesity development through epigenetic mechanisms has been described. Thus, identification of those individuals who present with changes in DNA methylation profiles, certain histone modifications, or other epigenetically related processes could help to predict their susceptibility to gain or lose weight. Indeed, research concerning epigenetic mechanisms affecting weight homeostasis may play a role in the prevention of excessive fat deposition, the prediction of the most appropriate weight reduction plan, and the implementation of newer therapeutic approaches.