Analysis of platelets from a diet-induced obesity rat model: elucidating platelet dysfunction in obesity

Obesity is one of the main health problems in industrialized countries. The contribution of multiple factors developed in obesity can hardly be modeled in vitro. In this context, the development of animal models mimicking human obesity could be essential. The aim of the present study was to compare platelets from a diet-induced obesity (DIO) rat model with their lean control group in order to elucidate platelet dysfunction mechanisms in obesity and correlate the results with previous data from morbid obese patients. In parallel, we also established a blood collection and platelet isolation methodology to study the DIO rat model at biochemical and functional level. Optimal blood collection was obtained from vena cava and platelet isolation was based on a serial of centrifugations avoiding platelet activation. Our results show that the DIO rat model simulate obesity pathologically since weight gain, fasting glucose and platelet counts are increased in obese rats. Interestingly, platelet levels of the active form of Src (pTyr(419)) showed a tendency to increase in DIO rats pointing towards a potential dysfunction in Src family kinases-related signalling pathways in obesity. Moreover, platelets from DIO rats adhere more to collagen compared with the control group, pointing towards Glycoprotein VI (GPVI) as one of the dysregulated receptors in obesity, in agreement with our recent studies in humans. These results confirm that obesity, in line with human studies, present a platelet dysregulation, and highlight the relevance of considering novel antithrombotic drug targets in these patients, such as GPVI

Gut Microbiota Composition in Male Rat Models under Different Nutritional Status and Physical Activity and Its Association with Serum Leptin and Ghrelin Levels

Background: Several evidences indicate that gut microbiota is involved in the control of host energy metabolism. Objective: To evaluate the differences in the composition of gut microbiota in rat models under different nutritional status and physical activity and to identify their associations with serum leptin and ghrelin levels. Methods: In a case control study, forty male rats were randomly assigned to one of these four experimental groups: ABA group with food restriction and free access to exercise; control ABA group with food restriction and no access to exercise; exercise group with free access to exercise and feed ad libitum and ad libitum group without access to exercise and feed ad libitum. The fecal bacteria composition was investigated by PCR-denaturing gradient gel electrophoresis and real-time qPCR. Results: In restricted eaters, we have found a significant increase in the number of Proteobacteria, Bacteroides, Clostridium, Enterococcus, Prevotella and M. smithii and a significant decrease in the quantities of Actinobacteria, Firmicutes, Bacteroidetes, B. coccoides-E. rectale group, Lactobacillus and Bifidobacterium with respect to unrestricted eaters. Moreover, a significant increase in the number of Lactobacillus, Bifidobacterium and B. coccoides-E. rectale group was observed in exercise group with respect to the rest of groups. We also found a significant positive correlation between the quantity of Bifidobacterium and Lactobacillus and serum leptin levels, and a significant and negative correlation among the number of Clostridium, Bacteroides and Prevotella and serum leptin levels in all experimental groups. Furthermore, serum ghrelin levels were negatively correlated with the quantity of Bifidobacterium, Lactobacillus and B. coccoides-Eubacterium rectale group and positively correlated with the number of Bacteroides and Prevotella. Conclusions: Nutritional status and physical activity alter gut microbiota composition affecting the diversity and similarity. This study highlights the associations between gut microbiota and appetite-regulating hormones that may be important in terms of satiety and host metabolism

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

Vesicles Shed by Pathological Murine Adipocytes Spread Pathology: Characterization and Functional Role of Insulin Resistant/Hypertrophied Adiposomes

Extracellular vesicles (EVs) have recently emerged as a relevant way of cell to cell communication, and its analysis has become an indirect approach to assess the cell/tissue of origin status. However, the knowledge about their nature and role on metabolic diseases is still very scarce. We have established an insulin resistant (IR) and two lipid (palmitic/oleic) hypertrophied adipocyte cell models to isolate EVs to perform a protein cargo qualitative and quantitative Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH) analysis by mass spectrometry. Our results show a high proportion of obesity and IR-related proteins in pathological EVs; thus, we propose a panel of potential obese adipose tissue EV-biomarkers. Among those, lipid hypertrophied vesicles are characterized by ceruloplasmin, mimecan, and perilipin 1 adipokines, and those from the IR by the striking presence of the adiposity and IR related transforming growth factor-beta-induced protein ig-h3 (TFGBI). Interestingly, functional assays show that IR and hypertrophied adipocytes induce differentiation/hypertrophy and IR in healthy adipocytes through secreted EVs. Finally, we demonstrate that lipid atrophied adipocytes shed EVs promote macrophage inflammation by stimulating IL-6 and TNFα expression. Thus, we conclude that pathological adipocytes release vesicles containing representative protein cargo of the cell of origin that are able to induce metabolic alterations on healthy cells probably exacerbating the disease once establishedThis research was funded by Instituto de Salud Carlos III-FEDER, grant number PI16/01212S

Effect of a Very-Low-Calorie Ketogenic Diet on Circulating Myokine Levels Compared with the Effect of Bariatric Surgery or a Low-Calorie Diet in Patients with Obesity

: The preservation of muscle mass and muscle function after weight loss therapy is currently a considerable challenge in the fight against obesity. Muscle mass secretes proteins called myokines that have relevant functions in the regulation of metabolism and health. This study was aimed to evaluate whether a very low-calorie ketogenic (VLCK) diet may modulate myokine levels, in addition to changes in body composition, compared to a standard, balanced low-calorie (LC) diet or bariatric surgery in patients with obesity. Body composition, ketosis, insulin sensitivity and myokines were evaluated in 79 patients with overweight/obesity after a therapy to lose weight with a VLCK diet, a LC diet or bariatric surgery. The follow-up was 6 months. The weight loss therapies induced changes in myokine levels in association with changes in body composition and biochemical parameters. The effects on circulating myokine levels compared to those at baseline were stronger after the VLCK diet than LC diet or bariatric surgery. Differences reached statistical significance for IL-8, MMP2 and irisin. In conclusion, nutritional interventions or bariatric surgery to lose weight induces changes in circulating myokine levels, being this effect potentially most notable after following a VLCK diet

Effect of a Very-Low-Calorie Ketogenic Diet on Circulating Myokine Levels Compared with the Effect of Bariatric Surgery or a Low-Calorie Diet in Patients with Obesity

The preservation of muscle mass and muscle function after weight loss therapy is currently a considerable challenge in the fight against obesity. Muscle mass secretes proteins called myokines that have relevant functions in the regulation of metabolism and health. This study was aimed to evaluate whether a very low-calorie ketogenic (VLCK) diet may modulate myokine levels, in addition to changes in body composition, compared to a standard, balanced low-calorie (LC) diet or bariatric surgery in patients with obesity. Body composition, ketosis, insulin sensitivity and myokines were evaluated in 79 patients with overweight/obesity after a therapy to lose weight with a VLCK diet, a LC diet or bariatric surgery. The follow-up was 6 months. The weight loss therapies induced changes in myokine levels in association with changes in body composition and biochemical parameters. The effects on circulating myokine levels compared to those at baseline were stronger after the VLCK diet than LC diet or bariatric surgery. Differences reached statistical significance for IL-8, MMP2 and irisin. In conclusion, nutritional interventions or bariatric surgery to lose weight induces changes in circulating myokine levels, being this effect potentially most notable after following a VLCK dietThis work was supported by the PronoKal Group® and grants from the Fondo de Investigacion Sanitaria, PI17/01287 research projects and CIBERobn (CB06/03/0003, CB12/03/30007, CB12/03/30002, CB06/03/0018) from the Instituto de Salud Carlos III (ISCIII)-Subdireccion General de Evaluacion y Fomento de la Investigación; Fondo Europeo de Desarrollo Regional (FEDER) and the Health Department of the Government of Navarra (48/2009), Spain and Linea Especial “Nutrition, Obesity and Health” (University of Navarra LE/97). Ana B Crujeiras is funded by a research contract “Miguel Servet” (CP17/00088) from the ISCIII, co-financed by the European Regional Development Fund (FEDER)S

ZNF577 Methylation Levels in Leukocytes From Women With Breast Cancer Is Modulated by Adiposity, Menopausal State, and the Mediterranean Diet

The methylation levels of ZNF577 in breast tumors has been previously identified as a possible epigenetic mark of breast cancer associated with obesity. The aim of the current study was to investigate differences in methylation levels of ZNF577 depending on obesity, menopausal state and dietary pattern in blood leukocytes, a non-invasive sample. The methylation levels of ZNF577 of two CpG sites (CpGs) located in promoter and island previously identified as differentially methylated according to adiposity and menopausal state by 450 k array (cg10635122, cg03562414) were evaluated by pyrosequencing in DNA from the blood leukocytes of breast cancer patients [n = 90; n = 64 (71.1%) overweight/obesity and n = 26 (28.9%) normal-weight] and paired tumor tissue biopsies (n = 8 breast cancer patients with obesity; n = 3/5 premenopausal/postmenopausal women). Differences in methylation levels were evaluated at each CpGs individually and at the mean of the two evaluated CpGs. Adherence to the Mediterranean diet was evaluated using the MEDAS-validated questionnaire, and the consumption of food groups of interest was also evaluated using the recommended intakes of the Sociedad Espanola de Nutricion Comunitaria. The methylation levels of ZNF577 were correlated between paired leukocytes and breast tumor biopsies (r = 0.62; p = 0.001). Moreover, higher methylation was found in leukocytes from patients with obesity (p = 0.002) and postmenopausal patients (p = 0.022) than patients with normal-weight or premenopausal, respectively. After adjusting for the body mass index and age, higher levels of ZNF577 methylation were also found in women with greater adherence to the Mediterranean diet (p = 0.017) or specific foods. Relevantly, the methylation levels of ZNF577 showed a good ability for fish consumption detection [area under the ROC curve (AUC) = 0.72; p = 0.016]. In conclusion, the association between methylation of ZNF577 and adiposity, menopausal state, and adherence to the Mediterranean diet can be detected in the blood leukocytes. The results guarantee the need of performing further studies in longer longitudinal cohorts in order to elucidate the role of ZNF577 methylation in the association between breast cancer, adiposity and dietary patterns

Altered pathways in methylome and transcriptome longitudinal analysis of normal weight and bariatric surgery women

DNA methylation could provide a link between environmental, genetic factors and weight control and can modify gene expression pattern. This study aimed to identify genes, which are differentially expressed and methylated depending on adiposity state by evaluating normal weight women and obese women before and after bariatric surgery (BS). We enrolled 24 normal weight (BMI: 22.5 +/- 1.6 kg/m(2)) and 24 obese women (BMI: 43.3 +/- 5.7 kg/m(2)) submitted to BS. Genome-wide methylation analysis was conducted using Infinium Human Methylation 450 BeadChip (threshold for significant CpG sites based on delta methylation level with a minimum value of 5%, a false discovery rate correction (FDR) of q /=2.0 was used to detect differentially expressed probes). The integrative analysis of both array data identified four genes (i.e. TPP2, PSMG6, ARL6IP1 and FAM49B) with higher methylation and lower expression level in pre-surgery women compared to normal weight women: and two genes (i.e. ZFP36L1 and USP32) that were differentially methylated after BS. These methylation changes were in promoter region and gene body. All genes are related to MAPK cascade, NIK/NF-kappaB signaling, cellular response to insulin stimulus, proteolysis and others. Integrating analysis of DNA methylation and gene expression evidenced that there is a set of genes relevant to obesity that changed after BS. A gene ontology analysis showed that these genes were enriched in biological functions related to adipogenesis, orexigenic, oxidative stress and insulin metabolism pathways. Also, our results suggest that although methylation plays a role in gene silencing, the majority of effects were not correlated

Recombinant human leptin treatment in genetic lipodystrophic syndromes: the long-term Spanish experience

Lipodystrophies are a group of diseases mainly characterized by a loss of adipose tissue and frequently associated with insulin resistance, hypertriglyceridemia, and hepatic steatosis. In uncommon lipodystrophies, these complications frequently are difficult to control with conventional therapeutic approaches. This retrospective study addressed the effectiveness of recombinant methionyl leptin (metreleptin) for improving glucose metabolism, lipid profile, and hepatic steatosis in patients with genetic lipodystrophic syndromes. We studied nine patients (five females and four males) with genetic lipodystrophies [seven with Berardinelli-Seip syndrome, one with atypical progeroid syndrome, and one with type 2 familial partial lipodystrophy (FPLD)]. Six patients were children under age 9 years, and all patients had baseline triglycerides levels >2.26 mmol/L and hepatic steatosis; six had poorly controlled diabetes mellitus. Metreleptin was self-administered subcutaneously daily at a final dose that ranged between 0.05 and 0.24 mg/(kg day) [median: 0.08 mg/(kg day)] according to the body weight. The duration of treatment ranged from 9 months to 5 years, 9 months (median: 3 years). Plasma glucose, hemoglobin A1c (Hb A1c), lipid profile, plasma insulin and leptin, and hepatic enzymes were evaluated at baseline and at least every 6 months. Except for the patient with FPLD, metreleptin replacement significantly improved metabolic control (Hb A1c: from 10.4 to 7.1 %, p < 0.05). Plasma triglycerides were reduced 76 % on average, and hepatic enzymes decreased more than 65 %. This study extends knowledge about metreleptin replacement in genetic lipodystrophies, bearing out its effectiveness for long periods of time

Equality, Equity, and Diversity: Educational Solutions in the Basque Country

Public education is one of the greatest achievements of European countries during the twentieth century. While schooling systems neither exclusively form citizens, nor are they sufficient to alleviate all inequalities, education plays an increasingly important strategic role in relieving social problems and promoting the civic and ethical upbringing of our children. Researchers and professors at the UPV/EHU have had the privilege to design and implement important educational projects in conjunction with government of Autonomous Community of the Basque Country, which has the authority over education in its territory. This book presents the timely (in most cases since 2000) observations, research, and programs that have resulted from this cooperation. Our stress—in both our theoretical and analytical dimensions—has been on the importance of diversity, the promotion of social and human values, and respect for basic human rights. In addition, we describe the cooperation that must be fostered—and the various needs met—between all educational "agents": academic researchers, administrators, teachers, parents, and the community at large to promote equality and fairness in our society.This book was published with generous financial support from the Basque Government.Introduction: Alfonso Unceta and Concepción Medrano ? Part 1 Education in the Basque Country ? 1. Education Provision in the Basque Country by Alfonso Unceta and Andrés Davila ? 2. Addressing Basque Diversity in the Classroom: Measures to Avoid Excluding At-Risk Youth by Begoña Martínez Domínguez ? 3. Improving Social Interaction: Experimentally Validated Proposals for Psycho-educational Intervention by Maite Garaigodobil and Jone Aliri ? 4. Socialization to Prevent Gender Violence in the Basque Country by Maria José Alonso Olea, Aitor Gómez González, and Nekane Beloki Arizti ? 5. Resolution and Transformation of At-School Conflicts by Ramón Alzate Sáez de Heredia, Lucía Gorbeña, and Cristina Merino ? Part 2 Socioeducational Context in the Basque Country ? 6. Learning Communities: A Basque Egalitarian Educational Project by Maite Arandia Loroño, Isabel Martínez Domínguez, and Iñaki Santa Cruz Ayo ? 7. Migrants en Route: Community Socioeducational Action by Miguel Arriaga Landeta and Begoña Abad Miguélez ? 8. Educating from the Family: A Proposal to Connect Homes and Institutions by Enrique Arranz Freijo, Fernando Olabarrieta Artetxe, Juan Luis Martín Ayala ? 9. The Development of Values and the Media by Concepción Medrano, Ana Aierbe, and Juan Ignacio Martínez de Morentin ? Index ? List of Contributor