Simple methodology for the quantitative analysis of fatty acids in human red blood cells

In the last years, there has been an increasing interest in evaluating possible relations between fatty acid (FA) patterns and the risk for chronic diseases. Due to the long life span (120 days) of red blood cells (RBCs), their FA profile reflects a longer term dietary intake and was recently suggested to be used as an appropriate biomarker to investigate correlations between FA metabolism and diseases. Therefore, the aim of this work was to develop and validate a simple and fast methodology for the quantification of a broad range of FAs in RBCs using gas chromatography with flame ionization detector, as a more common and affordable equipment suitable for biomedical and nutritional studies including a large number of samples. For this purpose, different sample preparation protocols were tested and compared, including a classic two-step method (Folch method) with modifications and different one-step methods, in which lipid extraction and derivatization were performed simultaneously. For the one-step methods, different methylation periods and the inclusion of a saponification reaction were evaluated. Differences in absolute FA concentrations were observed among the tested methods, in particular for some metabolically relevant FAs such as trans elaidic acid and eicosapentaenoic acid. The one-step method with saponification and 60 min of methylation time was selected since it allowed the identification of a higher number of FAs, and was further submitted to in-house validation. The proposed methodology provides a simple, fast and accurate tool to quantitatively analyse FAs in human RBCs, useful for clinical and nutritional studies.This work received financial support from the European Union (FEDER funds through COMPETE) and National Funds (FCT, Fundação para a Ciência e Tecnologia) through project PTDC/SAU-ENB/116929/2010 and EXPL/EMS-SIS/2215/2013. ROR acknowledges PhD scholarship SFRH/BD/97658/2013 attributed by FCT (Fundação para a Ciência e Tecnologia).info:eu-repo/semantics/publishedVersio

Valence-dependent influence of serotonin depletion on model-based choice strategy.

Human decision-making arises from both reflective and reflexive mechanisms, which underpin goal-directed and habitual behavioural control. Computationally, these two systems of behavioural control have been described by different learning algorithms, model-based and model-free learning, respectively. Here, we investigated the effect of diminished serotonin (5-hydroxytryptamine) neurotransmission using dietary tryptophan depletion (TD) in healthy volunteers on the performance of a two-stage decision-making task, which allows discrimination between model-free and model-based behavioural strategies. A novel version of the task was used, which not only examined choice balance for monetary reward but also for punishment (monetary loss). TD impaired goal-directed (model-based) behaviour in the reward condition, but promoted it under punishment. This effect on appetitive and aversive goal-directed behaviour is likely mediated by alteration of the average reward representation produced by TD, which is consistent with previous studies. Overall, the major implication of this study is that serotonin differentially affects goal-directed learning as a function of affective valence. These findings are relevant for a further understanding of psychiatric disorders associated with breakdown of goal-directed behavioural control such as obsessive-compulsive disorders or addictions.This research was funded by Wellcome Trust Grants awarded to VV (Intermediate WT Fellowship) and Programme Grant (089589/Z/09/Z) awarded to TWR, BJE, ACR, JWD and BJS. It was conducted at the Behavioural and Clinical Neuroscience Institute, which is supported by a joint award from the Medical Research Council and Wellcome Trust (G00001354). YW was supported by the Fyssen Foundation. SP is supported by Marie Curie Intra-European Fellowship (FP7-People-2012-IEF).This is the final version of the article. It first appeared from NPG via http://dx.doi.org/10.1038/mp.2015.4

Inverse relationship of food and alcohol intake to sleep measures in obesity.

BACKGROUND: Short sleep and weight gain are inversely related. Sleep deprivation acutely increases food intake but little is known about eating behavior in chronically sleep-deprived, obese individuals. OBJECTIVE: To characterize the relationship between sleep, food intake and alcohol consumption under free-living conditions in obese, chronically sleep-deprived individuals. DESIGN: Cross-sectional study of a cohort of obese men and premenopausal women. SUBJECTS: A total of 118 obese subjects (age: 40.3±6.7 years; 91 females/27 males; body mass index 38.7±6.4 kg m(-2)). MEASUREMENTS: Energy, macronutrient, alcohol and caffeine intake assessed by 3-day food records. Sleep duration estimated by actigraphy. Respiratory disturbance index assessed by a portable device. RESULTS: SUBJECTS slept 360.7±50.2 min per night and had a total energy intake of 2279.1±689 kcal per day. Sleep duration and energy intake were inversely related (r=-0.230, P=0.015). By extrapolation, each 30-min deficit per day in sleep duration would translate to an ∼83 kcal per day increase in energy intake. In addition, sleep apnea was associated with a shift from carbohydrate to fat intake. Alcohol intake in subjects consuming >3.5 g of alcohol per day (N=41) was inversely related to sleep duration (r=-0.472, P=0.002). CONCLUSIONS: Shorter sleep duration and obstructive sleep apnea are associated with higher energy, fat and alcohol intakes in obese individuals. The importance of this study relies on the population studied, obese subjects with chronic sleep deprivation. These novel findings apply to the large segment of the US population who are obese and sleep-deprived

Canagliflozin triggers the FGF23/1,25-dihydroxyvitamin D/PTH axis in healthy volunteers in a randomized crossover study

BACKGROUND. Sodium glucose cotransporter-2 (SGLT2) inhibitors are the most recently approved class of drugs for type 2 diabetes and provide both glycemic efficacy and cardiovascular risk reduction. A number of safety issues have been identified, including treatment-emergent bone fractures. To understand the overall clinical profile, these safety issues must be balanced against an attractive efficacy profile. Our study was designed to investigate pathophysiological mechanisms mediating treatment-emergent adverse effects on bone health. METHODS. We conducted a single-blind randomized crossover study in hospitalized healthy adults (n = 25) receiving either canagliflozin (300 mg/d) or placebo for 5 days. The primary end-point was the drug-induced change in AUC for plasma intact fibroblast growth factor 23 (FGF23) immunoactivity between 24 and 72 hours. RESULTS. Canagliflozin administration increased placebo-subtracted mean levels of serum phosphorus (+16%), plasma FGF23 (+20%), and plasma parathyroid hormone (PTH) (+25%), while decreasing the level of 1,25-dihydroxyvitamin D (–10%). There was substantial interindividual variation in the magnitude of each of these pharmacodynamic responses. The increase in plasma FGF23 was correlated with the increase in serum phosphorus, and the decrease in plasma 1,25-dihydroxyvitamin D was correlated with the increase in plasma FGF23. CONCLUSIONS. Canagliflozin induced a prompt increase in serum phosphorus, which triggers downstream changes in FGF23, 1,25-dihydroxyvitamin D, and PTH, with potential to exert adverse effects on bone health. These pharmacodynamic data provide a foundation for future research to elucidate pathophysiological mechanisms of adverse effects on bone health, with the objective of devising therapeutic strategies to mitigate the drug-associated fracture risk. TRIAL REGISTRATION. ClinicalTrial.gov (NCT02404870). FUNDING. Supported by the Intramural Program of NIDDK

Early effects of roflumilast on insulin sensitivity in adults with prediabetes and overweight/obesity involve age-associated fat mass loss – results of an exploratory study

Ijeoma M Muo,1 Sandra D MacDonald,2 Ritu Madan,3 Sung-Jun Park,1 Ahmed M Gharib,4 Pedro E. Martinez,5 Mary F Walter,3 Shanna B Yang,6 Justin A Rodante,7 Amber B Courville,6 Peter J Walter,8 Hongyi Cai,8 Michael Glicksman,3 Gioia M Guerrieri,5 Rivka R Ben-Dor,9 Ronald Ouwerkerk,4 Stephanie Mao,1 Jay H Chung1 1Laboratory of Obesity and Aging Research NHLBI, National Institutes of Health, Bethesda, MD 20892, USA; 2NHLBI Pulmonary Branch, Laboratory of Chronic Airway Infections, National Institutes of Health, Bethesda, MD 20892, USA; 3Diabetes Endocrinology and Obesity Branch, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA; 4Biomedical and Metabolic Imaging Branch NIDDK, National Institutes of Health, Bethesda, MD 20892, USA; 5Section on Behavioral Endocrinology, NIMH, National Institutes of Health, Bethesda, MD 20892, USA; 6Clinical Center Nutrition Department, National Institutes of Health, Bethesda, MD 20892, USA; 7Laboratory of Inflammation and Cardiometabolic Diseases, NHLBI, National Institutes of Health, Bethesda, MD 20892, USA; 8Mass Spectrometry Clinical Core, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA; 9NIMH, National Institutes of Health, Bethesda, MD 20892, USA Purpose: Roflumilast (Daliresp, Daxas) is a FDA-approved phosphodiesterase 4 (PDE4) inhibitor for the treatment of moderate-to-severe chronic obstructive pulmonary disease. In mice and in limited human studies, this oral medication can cause weight loss and improve insulin sensitivity. We set out to determine the mechanism of its effect on insulin sensitivity. Patients and methods: Eight adults with overweight/obesity and prediabetes received roflumilast for 6 weeks. Before and after roflumilast, subjects underwent tests of insulin sensitivity, mixed meal test, body composition, markers of inflammation, and mitochondria function. Dietary intake and physical activity were also assessed. Our primary outcome was the change in peripheral insulin sensitivity, as assessed by the hyper-insulinemic euglycemic clamp. Results: This study was underpowered for the primary outcome. Pre- and post-roflumilast mean peripheral insulin sensitivity were 48.7 and 70.0 mg/g fat free mass/minute, respectively, (P-value=0.18), respectively. Among the mixed meal variables, roflumilast altered glucagon-like peptide 1 (GLP-1) hormone the most, although the average effect was not statistically significant (P=0.18). Roflumilast induced a trend toward significance in 1) decreased energy intake (from 11,095 KJ to 8,4555 KJ, P=0.07), 2) decreased fat mass (from 34.53 to 32.97 kg, P=0.06), 3) decreased total and LDL cholesterol (P=0.06 for both variables), and 4) increased plasma free fatty acids (from 0.40 to 0.50 mEq/L, P=0.09) The interval changes in adiposity and free fatty acid were significantly associated with the subject’s age (P-value range=<0.001 to 0.02 for the correlations). Inflammatory and adhesion markers, though unchanged, significantly correlated with one another and with incretin hormones only after roflumilast. Conclusion: We demonstrate, for the first time in humans, increasing percentage of fat mass loss from roflumilast with increasing age in adults with prediabetes and overweight/obesity. We also demonstrate novel associations among roflumilast-induced changes in incretin hormones, inflammatory markers, peripheral insulin sensitivity, and adiposity. We conclude that roflumilast’s early effects on insulin sensitivity is indirect and likely mediated through roflumilast’s prioritization of lipid over glucose handling.Clinical trials registration: NCT01862029. Keywords: phosphodiesterase 4, obesity, diabetes, inflammation, incretins, aging &nbsp

Diet-Induced Changes in n-3- and n-6-Derived Endocannabinoids and Reductions in Headache Pain and Psychological Distress.

Omega-3 and omega-6 fatty acids are biosynthetic precursors of endocannabinoids with antinociceptive, anxiolytic, and neurogenic properties. We recently reported that targeted dietary manipulation-increasing omega-3 fatty acids while reducing omega-6 linoleic acid (the H3-L6 intervention)-reduced headache pain and psychological distress among chronic headache patients. It is not yet known whether these clinical improvements were due to changes in endocannabinoids and related mediators derived from omega-3 and omega-6 fatty acids. We therefore used data from this trial (N = 55) to investigate 1) whether the H3-L6 intervention altered omega-3- and omega-6-derived endocannabinoids in plasma and 2) whether diet-induced changes in these bioactive lipids were associated with clinical improvements. The H3-L6 intervention significantly increased the omega-3 docosahexaenoic acid derivatives 2-docosahexaenoylglycerol (+65%, P < .001) and docosahexaenoylethanolamine (+99%, P < .001) and reduced the omega-6 arachidonic acid derivative 2-arachidonoylglycerol (-25%, P = .001). Diet-induced changes in these endocannabinoid derivatives of omega-3 docosahexaenoic acid, but not omega-6 arachidonic acid, correlated with reductions in physical pain and psychological distress. These findings demonstrate that targeted dietary manipulation can alter endocannabinoids derived from omega-3 and omega-6 fatty acids in humans and suggest that 2-docosahexaenoylglycerol and docosahexaenoylethanolamine could have physical and/or psychological pain modulating properties. TRIAL REGISTRATION: ClinicalTrials.gov (NCT01157208) PERSPECTIVE: This article demonstrates that targeted dietary manipulation can alter endocannabinoids derived from omega-3 and omega-6 fatty acids and that these changes are related to reductions in headache pain and psychological distress. These findings suggest that dietary interventions could provide an effective, complementary approach for managing chronic pain and related condition