Genome-Wide DNA Methylation Analysis Identifies Novel Hypomethylated Non-Pericentromeric Genes with Potential Clinical Implications in ICF Syndrome

Introduction and Results Immunodeficiency, centromeric instability and facial anomalies syndrome (ICF) is a rare autosomal recessive disease, characterized by severe hypomethylation in pericentromeric regions of chromosomes (1, 16 and 9), marked immunodeficiency and facial anomalies. The majority of ICF patients present mutations in the DNMT3B gene, affecting the DNA methyltransferase activity of the protein. In the present study, we have used the Infinium 450K DNA methylation array to evaluate the methylation level of 450,000 CpGs in lymphoblastoid cell lines and untrasformed fibroblasts derived from ICF patients and healthy donors. Our results demonstrate that ICF-specific DNMT3B variants A603T/STP807ins and V699G/R54X cause global DNA hypomethylation compared to wild-type protein. We identified 181 novel differentially methylated positions (DMPs) including subtelomeric and intrachromosomic regions, outside the classical ICF-related pericentromeric hypomethylated positions. Interestingly, these sites were mainly located in intergenic regions and inside the CpG islands. Among the identified hypomethylated CpG-island associated genes, we confirmed the overexpression of three selected genes, BOLL, SYCP2 and NCRNA00221, in ICF compared to healthy controls, which are supposed to be expressed in germ line and silenced in somatic tissues. Conclusions In conclusion, this study contributes in clarifying the direct relationship between DNA methylation defect and gene expression impairment in ICF syndrome, identifying novel direct target genes of DNMT3B. A high percentage of the DMPs are located in the subtelomeric regions, indicating a specific role of DNMT3B in methylating these chromosomal sites. Therefore, we provide further evidence that hypomethylation in specific non-pericentromeric regions of chromosomes might be involved in the molecular pathogenesis of ICF syndrome. The detection of DNA hypomethylation at BOLL, SYCP2 and NCRNA00221 may pave the way for the development of specific clinical biomarkers with the aim to facilitate the identification of ICF patients

A two-gene epigenetic signature for the prediction of response to neoadjuvant chemotherapy in triple-negative breast cancer patients

Background: pathological complete response (pCR) after neoadjuvant chemotherapy (NAC) in triple-negative breast cancer (TNBC) varies between 30 and 40% approximately. To provide further insight into the prediction of pCR, we evaluated the role of an epigenetic methylation-based signature. Methods: epigenetic assessment of DNA extracted from biopsy archived samples previous to NAC from TNBC patients was performed. Patients included were categorized according to previous response to NAC in responder (pCR or residual cancer burden, RCB = 0) or non-responder (non-pCR or RCB > 0) patients. A methyloma study was performed in a discovery cohort by the Infinium HumanMethylation450 BeadChip (450K array) from Illumina. The epigenetic silencing of those methylated genes in the discovery cohort were validated by bisulfite pyrosequencing (PyroMark Q96 System version 2.0.6, Qiagen) and qRT-PCR in an independent cohort of TN patients and in TN cell lines. Results: twenty-four and 30 patients were included in the discovery and validation cohorts, respectively. In the discovery cohort, nine genes were differentially methylated: six presented higher methylation in non-responder patients (LOC641519, LEF1, HOXA5, EVC2, TLX3, CDKL2) and three greater methylation in responder patients (FERD3L, CHL1, and TRIP10). After validation, a two-gene (FER3L and TRIP10) epigenetic score predicted RCB = 0 with an area under the ROC curve (AUC) = 0.905 (95% CI = 0.805-1.000). Patients with a positive epigenetic two-gene score showed 78.6% RCB = 0 versus only 10.7% RCB = 0 if signature were negative. Conclusions: these results suggest that pCR in TNBC could be accurately predicted with an epigenetic signature of FERD3L and TRIP10 genes. Further prospective validation of these findings is warranted

Effect of A Very Low-Calorie Ketogenic Diet on Food and Alcohol Cravings, Physical and Sexual Activity, Sleep Disturbances, and Quality of Life in Obese Patients

Psychological well-being and hunger and food control are two relevant factors involved in the success of weight-loss therapy in treating obesity. Thus, this study aims to evaluate food and alcohol cravings, physical and sexual activity, sleep, and life quality (QoL) in obese patients following a very low-calorie ketogenic (VLCK) diet, as well as the role of weight lost and ketosis on these parameters. A battery of psychological test was performed in twenty obese patients (12 females, 47.2 +/- 10.2 year and BMI of 35.5 +/- 4.4) through the course of a 4-month VLCK diet on four subsequent visits: baseline, maximum ketosis, reduced ketosis, and endpoint. Each subject acted as their own control. Relevantly, the dietary-induced changes in body composition (7.7 units of BMI lost, 18 kg of fat mass (1.2 kg of visceral fat mass)) were associated with a statistically significant improvement in food craving scores, physical activity, sleepiness, and female sexual function. Overall, these results also translated in a notable enhancement in QoL of the treated obese patients. Therefore, the rapid and sustained weight and fat mass (FM) loss induced by the VLCK diet is associated with good food control and improvements in the psychological well-being parameters in obese subjects, which could contribute to the long-term success of this therapy

Higher baseline irisin concentrations are associated with greater reductions in glycemia and insulinemia after weight loss in obese subjects

Irisin is assumed to be a relevant link between muscle and weight maintenance as well as to mediate exercise benefits on health. The aim of this study was to assess the possible associations between irisin levels and glucose homeostasis in obese subjects with metabolic syndrome (MetS) following an energy-restricted treatment. Ninety-six adults with excessive body weight and MetS features underwent a hypocaloric dietary pattern for 8 weeks, within the RESMENA randomized controlled trial (www.clinicaltrials.gov; NCT01087086). After the intervention, dietary restriction significantly reduced body weight and evidenced a dietary-induced decrease in circulating levels of irisin in parallel with improvements on glucose homeostasis markers. Interestingly, participants with higher irisin values at baseline (above the median) showed a greater reduction on glucose (P=0.022) and insulin (P=0.021) concentrations as well as on the homeostasis model assessment index (P=0.008) and triglycerides (P=0.006) after the dietary intervention, compared with those presenting low-irisin baseline values (below the median). Interestingly, a positive correlation between irisin and carbohydrate intake was found at the end of the experimental period. In conclusion, irisin appears to be involved in glucose metabolism regulation after a dietary-induced weight loss

Oxidative Stress Induced by Excess of Adiposity Is Related to a Downregulation of Hepatic SIRT6 Expression in Obese Individuals

Sirt6 is a member of the sirtuin family involved in physiological and pathological processes including aging, cancer, obesity, diabetes, and energy metabolism. This study is aimed at evaluating the relationship between liver SIRT6 gene expression and the oxidative stress network depending on adiposity levels in Zucker rats, an animal model of metabolic syndrome. We observed that liver-specific SIRT6 expression is reduced in an in vivo model of spontaneous obesity and metabolic syndrome. We also observed that SIRT6 expression in the liver is positively associated with SIRT1 and GST-M2 expressions, two proteins involved in antioxidant protection pathways and inversely related to body weight and plasmatic oxidative status. Interestingly, the SIRT6 expression is upregulated after energy restriction-induced weight loss concomitantly with an improvement in oxidative stress markers. These results suggest that SIRT6 may be a potential therapeutic target for the treatment of obesity and associated metabolic disorders, such as liver disease.Centro de Investigacion Biomedica en Red de Fisiopatologia de la Obesidad y Nutricion (CIBERobn)Instituto de Salud Carlos IIIEuropean Regional Development Fund (FEDER

Weight regain after a diet-induced loss is predicted by higher baseline leptin and lower ghrelin plasma levels

CONTEXT: Appetite-related hormones may play an important role in weight regain after obesity therapy. OBJECTIVE: Our objective was to investigate the potential involvement of ghrelin, leptin, and insulin plasma levels in weight regain after a therapeutic hypocaloric diet. DESIGN: A group of obese/overweight volunteers (49 women and 55 men; 35 ± 7 yr; 30.7 ± 2.4 kg/m(2)) followed an 8-wk hypocaloric diet (-30% energy expenditure) and were evaluated again 32 wk after treatment. Body weight as well as plasma fasting ghrelin, leptin, and insulin concentrations were measured at three points (wk 0, 8, and 32). RESULTS: After the 8-wk hypocaloric diet, the average weight loss was -5.0 ± 2.2% (P < 0.001). Plasma leptin and insulin concentrations decreased significantly, whereas ghrelin levels did not markedly change. In the group regaining more than 10% of the weight loss, leptin levels were higher (P < 0.01), whereas ghrelin levels were lower (P < 0.05). No differences were observed in insulin levels. Weight regain at wk 32 was negatively correlated with ghrelin and positively associated with leptin levels at baseline (wk 0) and endpoint (wk 8). These outcomes showed a gender-specific influence, being statistically significant among men for ghrelin and between women for leptin. Moreover, a decrease in ghrelin after an 8-wk hypocaloric diet was related to an increased risk for weight regain (odds ratio = 3.109; P = 0.008) whereas a greater reduction in leptin (odds ratio = 0.141; P = 0.001) was related to weight-loss maintenance. CONCLUSIONS: Subjects with higher plasma leptin and lower ghrelin levels at baseline could be more prone to regain lost weight, and hormones levels could be proposed as biomarkers for predicting obesity-treatment outcomes

"Food Addiction" in Patients with Eating Disorders is Associated with Negative Urgency and Difficulties to Focus on Long-Term Goals

Objectives: The present study aimed to investigate if eating disorder patients differ in specific personality traits depending on a positive screening of food addiction (FA) and to find a model to predict FA in eating disorder patients using measures of personality and impulsivity. Methods : Two hundred seventy eight patients, having an eating disorder, self-reported on FA, impulsivity, personality, eating and general psychopathology. Patients were then split into two groups, depending on a positive or negative result on the FA screening. Analysis of variance was used to compare means between the two groups. Stepwise binary logistic regression was used to obtain a predictive model for the presence of FA. Results: Patients with FA had lower self-directedness, and more negative urgency and lack of perseverance than patients not reporting addictive eating. The probability of FA can be predicted by high negative urgency, high reward dependence, and low lack of premeditation. Conclusion: Eating disorder patients who have more problems to pursue tasks to the end and to focus on long-term goals seem to be more likely to develop addictive eating patterns

Genome-wide DNA methylation pattern in visceral adipose tissue differentiates insulin-resistant from insulin-sensitive obese subject

Elucidating the potential mechanisms involved in the detrimental effect of excess body weight on insulin action is an important priority in counteracting obesity-associated diseases. The present study aimed to disentangle the epigenetic basis of insulin resistance by performing a genome-wide epigenetic analysis in visceral adipose tissue (VAT) from morbidly obese patients depending on the insulin sensitivity evaluated by the clamp technique. The global human methylome screening performed in VAT from 7 insulin-resistant (IR) and 5 insulin-sensitive (IS) morbidly obese patients (discovery cohort) analyzed using the Infinium HumanMethylation450 BeadChip array identified 982 CpG sites able to perfectly separate the IR and IS samples. The identified sites represented 538 unique genes, 10% of which were diabetes-associated genes. The current work identified novel IR-related genes epigenetically regulated in VAT, such as COL9A1, COL11A2, CD44, MUC4, ADAM2, IGF2BP1, GATA4, TET1, ZNF714, ADCY9, TBX5, and HDACM. The gene with the largest methylation fold-change and mapped by 5 differentially methylated CpG sites located in island/shore and promoter region was ZNF714. This gene presented lower methylation levels in IR than in IS patients in association with increased transcription levels, as further reflected in a validation cohort (n = 24; 11 IR and 13 IS). This study reveals, for the first time, a potential epigenetic regulation involved in the dysregulation of VAT that could predispose patients to insulin resistance and future type 2 diabetes in morbid obesity, providing a potential therapeutic target and biomarkers for counteracting this process

'Food Addiction' in Patients with Eating Disorders is Associated with Negative Urgency and Difficulties to Focus on Long-Term Goals

Objectives: The present study aimed to investigate if eating disorder patients differ in specific personality traits depending on a positive screening of food addiction (FA) and to find a model to predict FA in eating disorder patients using measures of personality and impulsivity. Methods: Two hundred seventy eight patients, having an eating disorder, self-reported on FA, impulsivity, personality, eating and general psychopathology. Patients were then split into two groups, depending on a positive or negative result on the FA screening. Analysis of variance was used to compare means between the two groups. Stepwise binary logistic regression was used to obtain a predictive model for the presence of FA. Results: Patients with FA had lower self-directedness, and more negative urgency and lack of perseverance than patients not reporting addictive eating. The probability of FA can be predicted by high negative urgency, high reward dependence, and low lack of premeditation. Conclusion: Eating disorder patients who have more problems to pursue tasks to the end and to focus on long-term goals seem to be more likely to develop addictive eating patterns

Comparative Effects of Pterostilbene and Its Parent Compound Resveratrol on Oxidative Stress and Inflammation in Steatohepatitis Induced by High-Fat High-Fructose Feeding

Different studies have revealed that oxidative stress and inflammation are crucial in NAFLD (Non-alcoholic fatty liver disease). The aim of this study is to analyze whether pterostilbene and resveratrol are able to either avoid or delay the progression of non-alcoholic liver steatosis towards steatohepatitis. This has been performed by examining their effects on oxidative stress, inflammation, fibrosis and pre-carcinogenic stages. Rats were distributed into five experimental groups and were fed with either a standard diet or a high-fat high-fructose diet, supplemented or not with pterostilbene (15 or 30 mg/kg/d) or resveratrol (30 mg/kg/d), for 8 weeks. Liver histological analysis was carried out by haematoxylin–eosin staining. Serum and hepatic oxidative stress-related parameters were assessed using spectrophotometry, and the expression of genes related to inflammation, fibrosis and cancer by qRT-PCR. The dietary model used in this study led to the development of steatohepatitis, where rats displayed oxidative stress, inflammation and ballooning, although not fibrosis. It also modified the expression of hepatocarcinoma-related genes. The results show, for the first time, that pterostilbene was able to partially prevent these alterations, with the exception of changes in hepatocarcinoma-related genes, mainly at 30 mg/kg/d. Pterostilbene was more effective than its parent compound resveratrol, probably due to its high bioavailability and higher anti-oxidant and anti-inflammatory activities, attributable to its different chemical structure.This study was supported by grants from the Ministerio de Economía y Competitividad (AGL-2015-65719-R), Fondo Europeo de Desarrollo Regional (FEDER), the Instituto de Salud Carlos III (CIBERobn) under Grant CB12/03/30007 and the University of the Basque Country under Grant GIU18-173