A nutritional biomarker score of the Mediterranean diet and incident type 2 diabetes: Integrated analysis of data from the MedLey randomised controlled trial and the EPIC-InterAct case-cohort study.

BACKGROUND: Self-reported adherence to the Mediterranean diet has been modestly inversely associated with incidence of type 2 diabetes (T2D) in cohort studies. There is uncertainty about the validity and magnitude of this association due to subjective reporting of diet. The association has not been evaluated using an objectively measured biomarker of the Mediterranean diet. METHODS AND FINDINGS: We derived a biomarker score based on 5 circulating carotenoids and 24 fatty acids that discriminated between the Mediterranean or habitual diet arms of a parallel design, 6-month partial-feeding randomised controlled trial (RCT) conducted between 2013 and 2014, the MedLey trial (128 participants out of 166 randomised). We applied this biomarker score in an observational study, the European Prospective Investigation into Cancer and Nutrition (EPIC)-InterAct case-cohort study, to assess the association of the score with T2D incidence over an average of 9.7 years of follow-up since the baseline (1991 to 1998). We included 22,202 participants, of whom 9,453 were T2D cases, with relevant biomarkers from an original case-cohort of 27,779 participants sampled from a cohort of 340,234 people. As a secondary measure of the Mediterranean diet, we used a score estimated from dietary-self report. Within the trial, the biomarker score discriminated well between the 2 arms; the cross-validated C-statistic was 0.88 (95% confidence interval (CI) 0.82 to 0.94). The score was inversely associated with incident T2D in EPIC-InterAct: the hazard ratio (HR) per standard deviation of the score was 0.71 (95% CI: 0.65 to 0.77) following adjustment for sociodemographic, lifestyle and medical factors, and adiposity. In comparison, the HR per standard deviation of the self-reported Mediterranean diet was 0.90 (95% CI: 0.86 to 0.95). Assuming the score was causally associated with T2D, higher adherence to the Mediterranean diet in Western European adults by 10 percentiles of the score was estimated to reduce the incidence of T2D by 11% (95% CI: 7% to 14%). The study limitations included potential measurement error in nutritional biomarkers, unclear specificity of the biomarker score to the Mediterranean diet, and possible residual confounding. CONCLUSIONS: These findings suggest that objectively assessed adherence to the Mediterranean diet is associated with lower risk of T2D and that even modestly higher adherence may have the potential to reduce the population burden of T2D meaningfully. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (ANZCTR) ACTRN12613000602729 https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=363860.

Genetic regulation of liver lipids in a mouse model of insulin resistance and hepatic steatosis.

To elucidate the contributions of specific lipid species to metabolic traits, we integrated global hepatic lipid data with other omics measures and genetic data from a cohort of about 100 diverse inbred strains of mice fed a high-fat/high-sucrose diet for 8 weeks. Association mapping, correlation, structure analyses, and network modeling revealed pathways and genes underlying these interactions. In particular, our studies lead to the identification of Ifi203 and Map2k6 as regulators of hepatic phosphatidylcholine homeostasis and triacylglycerol accumulation, respectively. Our analyses highlight mechanisms for how genetic variation in hepatic lipidome can be linked to physiological and molecular phenotypes, such as microbiota composition.

Associations of circulating C-reactive proteins, APOE ε4, and brain markers for Alzheimer's disease in healthy samples across the lifespan.

The apolipoprotein E gene ε4 allele (APOE ε4) and higher circulating level of C-reactive protein (CRP) have been extensively investigated as risk factors for Alzheimer's disease (AD). Paradoxically, APOE ε4 has been associated with lower levels of blood CRP in middle-aged and older populations. However, few studies have investigated this intriguing relation and its impact on neurological markers for AD in younger ages, nor across the whole lifespan. Here, we examine associations of blood CRP levels, APOE ε4, and biomarkers for AD in a cognitively healthy lifespan cohort (N up to 749; 20-81 years of age) and replicate the findings in UK Biobank (N = 304 322; 37-72 years of age), the developmental ABCD study (N = 10 283; 9-11 years of age), and a middle-aged sample (N = 339; 40-65 years of age). Hippocampal volume, brain amyloid-ß (Aß) plaque levels, cerebrospinal fluid (CSF) levels of Aß and tau species, and neurofilament protein light protein (NFL) were used as AD biomarkers in subsamples. In addition, we examined the genetic contribution to the variation of CRP levels over different CRP ranges using polygenic scores for CRP (PGS-CRP). Our results show APOE ε4 consistently associates with low blood CRP levels across all age groups (p < 0.05). Strikingly, both ε4 and PGS-CRP associated mainly with blood CRP levels within the low range (<5mg/L). We then show both APOE ε4 and high CRP levels associate with smaller hippocampus volumes across the lifespan (p < 0.025). APOE ε4 was associated with high Aß plaque levels in the brain (FDR-corrected p = 8.69x10-4), low levels of CSF Aß42 (FDR-corrected p = 6.9x10-2), and lower ratios of Aß42 to Aß40 (FDR-corrected p = 5.08x10-5). Blood CRP levels were weakly correlated with higher ratio of CSF Aß42 to Aß40 (p = 0.03, FDR-corrected p = 0.4). APOE ε4 did not correlate with blood concentrations of another 9 inflammatory cytokines, and none of these cytokines correlated with AD biomarkers. CONCLUSION: The inverse correlation between APOEε 4 and blood CRP levels existed before any pathological AD biomarker was observed, and only in the low CRP level range. Thus, we suggest to investigate whether APOEε 4 can confer risk by being associated with a lower inflammatory response to daily exposures, possibly leading to greater accumulation of low-grade inflammatory stress throughout life. A lifespan perspective is needed to understand this relationship concerning risk of developing AD.

Personalised nutrition advice reduces intake of discretionary foods and beverages: findings from the Food4Me randomised controlled trial.

BACKGROUND: The effect of personalised nutrition advice on discretionary foods intake is unknown. To date, two national classifications for discretionary foods have been derived. This study examined changes in intake of discretionary foods and beverages following a personalised nutrition intervention using these two classifications. METHODS: Participants were recruited into a 6-month RCT across seven European countries (Food4Me) and were randomised to receive generalised dietary advice (control) or one of three levels of personalised nutrition advice (based on diet [L1], phenotype [L2] and genotype [L3]). Dietary intake was derived from an FFQ. An analysis of covariance was used to determine intervention effects at month 6 between personalised nutrition (overall and by levels) and control on i) percentage energy from discretionary items and ii) percentage contribution of total fat, SFA, total sugars and salt to discretionary intake, defined by Food Standards Scotland (FSS) and Australian Dietary Guidelines (ADG) classifications. RESULTS: Of the 1607 adults at baseline, n = 1270 (57% female) completed the intervention. Percentage sugars from FSS discretionary items was lower in personalised nutrition vs control (19.0 ± 0.37 vs 21.1 ± 0.65; P = 0.005). Percentage energy (31.2 ± 0.59 vs 32.7 ± 0.59; P = 0.031), percentage total fat (31.5 ± 0.37 vs 33.3 ± 0.65; P = 0.021), SFA (36.0 ± 0.43 vs 37.8 ± 0.75; P = 0.034) and sugars (31.7 ± 0.44 vs 34.7 ± 0.78; P < 0.001) from ADG discretionary items were lower in personalised nutrition vs control. There were greater reductions in ADG percentage energy and percentage total fat, SFA and salt for those randomised to L3 vs L2. CONCLUSIONS: Compared with generalised dietary advice, personalised nutrition advice achieved greater reductions in discretionary foods intake when the classification included all foods high in fat, added sugars and salt. Future personalised nutrition approaches may be used to target intake of discretionary foods. TRIAL REGISTRATION: Clinicaltrials.gov NCT01530139 . Registered 9 February 2012.

The association between circulating 25-hydroxyvitamin D metabolites and type 2 diabetes in European populations: A meta-analysis and Mendelian randomisation analysis.

BACKGROUND: Prior research suggested a differential association of 25-hydroxyvitamin D (25(OH)D) metabolites with type 2 diabetes (T2D), with total 25(OH)D and 25(OH)D3 inversely associated with T2D, but the epimeric form (C3-epi-25(OH)D3) positively associated with T2D. Whether or not these observational associations are causal remains uncertain. We aimed to examine the potential causality of these associations using Mendelian randomisation (MR) analysis. METHODS AND FINDINGS: We performed a meta-analysis of genome-wide association studies for total 25(OH)D (N = 120,618), 25(OH)D3 (N = 40,562), and C3-epi-25(OH)D3 (N = 40,562) in participants of European descent (European Prospective Investigation into Cancer and Nutrition [EPIC]-InterAct study, EPIC-Norfolk study, EPIC-CVD study, Ely study, and the SUNLIGHT consortium). We identified genetic variants for MR analysis to investigate the causal association of the 25(OH)D metabolites with T2D (including 80,983 T2D cases and 842,909 non-cases). We also estimated the observational association of 25(OH)D metabolites with T2D by performing random effects meta-analysis of results from previous studies and results from the EPIC-InterAct study. We identified 10 genetic loci associated with total 25(OH)D, 7 loci associated with 25(OH)D3 and 3 loci associated with C3-epi-25(OH)D3. Based on the meta-analysis of observational studies, each 1-standard deviation (SD) higher level of 25(OH)D was associated with a 20% lower risk of T2D (relative risk [RR]: 0.80; 95% CI 0.77, 0.84; p < 0.001), but a genetically predicted 1-SD increase in 25(OH)D was not significantly associated with T2D (odds ratio [OR]: 0.96; 95% CI 0.89, 1.03; p = 0.23); this result was consistent across sensitivity analyses. In EPIC-InterAct, 25(OH)D3 (per 1-SD) was associated with a lower risk of T2D (RR: 0.81; 95% CI 0.77, 0.86; p < 0.001), while C3-epi-25(OH)D3 (above versus below lower limit of quantification) was positively associated with T2D (RR: 1.12; 95% CI 1.03, 1.22; p = 0.006), but neither 25(OH)D3 (OR: 0.97; 95% CI 0.93, 1.01; p = 0.14) nor C3-epi-25(OH)D3 (OR: 0.98; 95% CI 0.93, 1.04; p = 0.53) was causally associated with T2D risk in the MR analysis. Main limitations include the lack of a non-linear MR analysis and of the generalisability of the current findings from European populations to other populations of different ethnicities. CONCLUSIONS: Our study found discordant associations of biochemically measured and genetically predicted differences in blood 25(OH)D with T2D risk. The findings based on MR analysis in a large sample of European ancestry do not support a causal association of total 25(OH)D or 25(OH)D metabolites with T2D and argue against the use of vitamin D supplementation for the prevention of T2D.

Characteristics of participants who benefit most from personalised nutrition: findings from the pan-European Food4Me randomised controlled trial.

Little is known about who would benefit from Internet-based personalised nutrition (PN) interventions. This study aimed to evaluate the characteristics of participants who achieved greatest improvements (i.e. benefit) in diet, adiposity and biomarkers following an Internet-based PN intervention. Adults (n 1607) from seven European countries were recruited into a 6-month, randomised controlled trial (Food4Me) and randomised to receive conventional dietary advice (control) or PN advice. Information on dietary intake, adiposity, physical activity (PA), blood biomarkers and participant characteristics was collected at baseline and month 6. Benefit from the intervention was defined as ≥5 % change in the primary outcome (Healthy Eating Index) and secondary outcomes (waist circumference and BMI, PA, sedentary time and plasma concentrations of cholesterol, carotenoids and omega-3 index) at month 6. For our primary outcome, benefit from the intervention was greater in older participants, women and participants with lower HEI scores at baseline. Benefit was greater for individuals reporting greater self-efficacy for 'sticking to healthful foods' and who 'felt weird if [they] didn't eat healthily'. Participants benefited more if they reported wanting to improve their health and well-being. The characteristics of individuals benefiting did not differ by other demographic, health-related, anthropometric or genotypic characteristics. Findings were similar for secondary outcomes. These findings have implications for the design of more effective future PN intervention studies and for tailored nutritional advice in public health and clinical settings.

Infant cholesterol and glycated haemoglobin concentrations vary widely-Associations with breastfeeding, infant diet and maternal biomarkers.

AIM: Elevated total cholesterol (TC) and glycated haemoglobin (HbA1c) are risk factors for cardiovascular disease; however, little is known about their determinants in infants. We aimed to describe TC and HbA1c concentrations in infants aged 8-14 months and explore the relation between infant TC, HbA1c, breastfeeding, infant diet, and maternal TC and HbA1c. METHODS: In this cross-sectional pilot study, mothers of infants aged 6 and 12 months were invited to complete a food frequency questionnaire and to take home-based dried blood spot samples from themselves and their infants. RESULTS: Among the 143 included infants, the mean (SD, range) concentration was 4.1 (0.8, 2.3-6.6) mmol/L for TC and 4.9 (0.4, 3.7-6.0)% for HbA1c. There was no significant difference between age groups and sexes. There was a positive relation between TC concentrations of all infants and mothers (B = 0.30 unadjusted, B = 0.32 adjusted, P < .001 for both) and a negative relation between infant TC and intake of unsaturated fatty acids in the oldest age group (B = -0.09, P = .03 unadjusted, B = -0.08, P = .06 adjusted). Infant HbA1c was not significantly related to diet or maternal HbA1c. CONCLUSION: TC and HbA1c concentrations varied widely among infants aged 8-14 months. Infant TC was associated with macronutrient intake and maternal TC.

Genetic, dietary, and sex-specific regulation of hepatic ceramides and the relationship between hepatic ceramides and IR.

Elevated hepatic ceramide levels have been implicated in both insulin resistance (IR) and hepatic steatosis. To understand the factors contributing to hepatic ceramide levels in mice of both sexes, we have quantitated ceramides in a reference population of mice, the Hybrid Mouse Diversity Panel that has been previously characterized for a variety of metabolic syndrome traits. We observed significant positive correlations between Cer(d18:1/16:0) and IR/hepatic steatosis, consistent with previous findings, although the relationship broke down between sexes, as females were less insulin resistant, but had higher Cer(d18:1/16:0) levels than males. The sex difference was due in part to testosterone-mediated repression of ceramide synthase 6. One ceramide species, Cer(d18:1/20:0), was present at higher levels in males and was associated with IR only in males. Clear evidence of gene-by-sex and gene-by-diet interactions was observed, including sex-specific genome-wide association study results. Thus, our studies show clear differences in how hepatic ceramides are regulated between the sexes, which again suggests that the physiological roles of certain hepatic ceramides differ between the sexes.

Capturing health and eating status through a nutritional perception screening questionnaire (NPSQ9) in a randomised internet-based personalised nutrition intervention: the Food4Me study.

BACKGROUND: National guidelines emphasize healthy eating to promote wellbeing and prevention of non-communicable diseases. The perceived healthiness of food is determined by many factors affecting food intake. A positive perception of healthy eating has been shown to be associated with greater diet quality. Internet-based methodologies allow contact with large populations. Our present study aims to design and evaluate a short nutritional perception questionnaire, to be used as a screening tool for assessing nutritional status, and to predict an optimal level of personalisation in nutritional advice delivered via the Internet. METHODS: Data from all participants who were screened and then enrolled into the Food4Me proof-of-principle study (n = 2369) were used to determine the optimal items for inclusion in a novel screening tool, the Nutritional Perception Screening Questionnaire-9 (NPSQ9). Exploratory and confirmatory factor analyses were performed on anthropometric and biochemical data and on dietary indices acquired from participants who had completed the Food4Me dietary intervention (n = 1153). Baseline and intervention data were analysed using linear regression and linear mixed regression, respectively. RESULTS: A final model with 9 NPSQ items was validated against the dietary intervention data. NPSQ9 scores were inversely associated with BMI (ß = -0.181, p < 0.001) and waist circumference (Β = -0.155, p < 0.001), and positively associated with total carotenoids (ß = 0.198, p < 0.001), omega-3 fatty acid index (ß = 0.155, p < 0.001), Healthy Eating Index (HEI) (ß = 0.299, p < 0.001) and Mediterranean Diet Score (MDS) (ß = 0. 279, p < 0.001). Findings from the longitudinal intervention study showed a greater reduction in BMI and improved dietary indices among participants with lower NPSQ9 scores. CONCLUSIONS: Healthy eating perceptions and dietary habits captured by the NPSQ9 score, based on nine questionnaire items, were associated with reduced body weight and improved diet quality. Likewise, participants with a lower score achieved greater health improvements than those with higher scores, in response to personalised advice, suggesting that NPSQ9 may be used for early evaluation of nutritional status and to tailor nutritional advice. TRIAL REGISTRATION: NCT01530139 .

Vitamin D supplementation and vitamin D status in children of immigrant background in Norway.

OBJECTIVE: Sufficient vitamin D status during infancy is important for child health and development. Several initiatives for improving vitamin D status among immigrant children have been implemented in Norway. The present study aimed to evaluate the vitamin D status and its determinants in children of immigrant background in Oslo. DESIGN: Cross-sectional study. SETTING: Child health clinics in Oslo. SUBJECTS: Healthy children with immigrant background (n 102) aged 9-16 months were recruited at the routine one-year check-up from two child health clinics with high proportions of immigrant clients. Blood samples were collected using the dried blood spot technique and analysed for serum 25-hydroxyvitamin D (s-25(OH)D) concentration using LC-MS/MS. RESULTS: Mean s-25(OH)D was 52·3 (sd 16·7) nmol/l, with only three children below 25 nmol/l and none below 12·5 nmol/l. There was no significant gender, ethnic or seasonal variation in s-25(OH)D. However, compared with breast-fed children, s-25(OH)D concentration was significantly higher among children who were about 1 year of age and not breast-fed. About 38 % of the children were anaemic, but there was no significant correlation between s-25(OH)D and Hb (Pearson correlation, r=0·1, P=0·33). CONCLUSIONS: Few children in the study had vitamin D deficiency, but about 47 % of the children in the study population were under the recommended s-25(OH)D sufficiency level of ≥50 nmol/l.

Vitamin D status in pre-school children in rural Nepal.

OBJECTIVE: Vitamin D plays a major role in Ca and bone metabolism, and its extraskeletal functions are being appraised. Although inadequate vitamin D concentrations have been reported in populations worldwide, too little is known about vitamin D status and its determinants among children in developing countries. We aimed to determine vitamin D status and its determinants in Nepalese children of pre-school age. DESIGN: A community-based, cross-sectional study. SETTING: Rural Nepal at latitude 27.39° N. SUBJECTS: Healthy children (n 280) aged 12-60 months, selected randomly from the records of a vitamin A supplementation programme. Blood samples were collected using the dried blood spot technique and analysed for serum 25-hydroxyvitamin D (s-25(OH)D) concentration using liquid chromatography-tandem mass spectrometry. Ca intake and background variables were assessed with a structured questionnaire. RESULTS: Hypovitaminosis D, defined as s-25(OH)D concentration less than 50 nmol/l, was found in 91.1% of the children. S-25(OH)D concentration was not related to gender, socio-economic indicators, sun exposure or nutritional status. Currently breast-fed children had higher s-25(OH)D concentrations (36.4 (sd 13.2) nmol/l) than those who were not (28.6 (sd 9.8) nmol/l, P<0.001). Adjustment for sociodemographic factors did not alter the results. CONCLUSION: There is widespread vitamin D deficiency among pre-school children in a rural area of Nepal. In our sample, sociodemographic factors did not affect the vitamin D status of children, but prolonged breast-feeding was associated with higher s-25(OH)D concentrations. Further research is required to investigate the health consequences of poor vitamin D status for this population.

Association Between Prediagnostic Serum 25-Hydroxyvitamin D Concentration and Glioma.

There are no previous studies of the association between prediagnostic serum vitamin D concentration and glioma. Vitamin D has immunosuppressive properties; as does glioma. It was, therefore, our hypothesis that elevated vitamin D concentration would increase glioma risk. We conducted a nested case-control study using specimens from the Janus Serum Bank cohort in Norway. Blood donors who were subsequently diagnosed with glioma (n = 592), between 1974 and 2007, were matched to donors without glioma (n = 1112) on date and age at blood collection and sex. We measured 25-hydroxyvitamin D [25(OH)D], an indicator of vitamin D availability, using liquid chromatography coupled with mass spectrometry. Seasonally adjusted odds ratios (ORs) and 95% confidence intervals (95% CIs) were estimated for each control quintile of 25(OH)D using conditional logistic regression. Among men diagnosed with high grade glioma >56, we found a negative trend (P = .04). Men diagnosed ≤ 56 showed a borderline positive trend (P = .08). High levels (>66 nmol/L) of 25(OH)D in men >56 were inversely related to high grade glioma from ≥2 yr before diagnosis (OR = 0.59; 95% CI = 0.38, 0.91) to ≥15 yr before diagnosis (OR = 0.61; 95% CI = 0.38,0.96). Our findings are consistent long before glioma diagnosis and are therefore unlikely to reflect preclinical disease.

Diet-induced Weight Loss and Phenotypic Flexibility Among Healthy Overweight Adults: A Randomized Trial.

BACKGROUND: The capacity of an individual to respond to changes in food intake so that postprandial metabolic perturbations are resolved, and metabolism returns to its pre-prandial state, is called phenotypic flexibility. This ability may be a more important indicator of current health status than metabolic markers in a fasting state. AIM: In this parallel randomized controlled trial study, an energy-restricted healthy diet and 2 dietary challenges were used to assess the effect of weight loss on phenotypic flexibility. METHODS: Seventy-two volunteers with overweight and obesity underwent a 12-wk dietary intervention. The participants were randomized to a weight loss group (WLG) with 20% less energy intake or a weight-maintenance group (WMG). At weeks 1 and 12, participants were assessed for body composition by MRI. Concurrently, markers of metabolism and insulin sensitivity were obtained from the analysis of plasma metabolome during 2 different dietary challenges-an oral glucose tolerance test (OGTT) and a mixed-meal tolerance test. RESULTS: Intended weight loss was achieved in the WLG (-5.6 kg, P < 0.0001) and induced a significant reduction in total and regional adipose tissue as well as ectopic fat in the liver. Amino acid-based markers of insulin action and resistance such as leucine and glutamate were reduced in the postprandial phase of the OGTT in the WLG by 11.5% and 28%, respectively, after body weight reduction. Weight loss correlated with the magnitude of changes in metabolic responses to dietary challenges. Large interindividual variation in metabolic responses to weight loss was observed. CONCLUSION: Application of dietary challenges increased sensitivity to detect metabolic response to weight loss intervention. Large interindividual variation was observed across a wide range of measurements allowing the identification of distinct responses to the weight loss intervention and mechanistic insight into the metabolic response to weight loss.

Dried blood spot (DBS) sample collection for determination of the oxidative stress biomarker 8-epi-PGF(2α) in humans using liquid chromatography/tandem mass spectrometry.

RATIONALE: F2-isoprostanes are a series of prostaglandin F2-like compounds that are formed by free-radical-catalyzed peroxidation of arachidonic acid (ARA). Several F2-isoprostanes, but in particular 8-epi-PGF(2α), are widely used as oxidative stress biomarkers. In this study we have developed an analytical tool for finger-tip blood sampling and analysis of 8-epi-PGF(2α) from dried blood spots (DBS). METHODS: We have applied solid-phase extraction (SPE) and liquid chromatography/tandem mass spectrometry (LC/MS/MS) for the extraction, separation and detection of 8-epi-PGF(2α) in DBS and have studied the stability of this marker using the DBS collection platform. RESULTS: The mass limit of detection (mLOD) for 8-epi-PGF(2α) extracted from DBS samples was 1.5 pg while the concentration limit of detection (cLOD) and concentration limit of quantitation (cLOQ) were 6 pg/mL and 18 pg/mL, respectively. All values based on triplicate analysis. Sufficient stability of 8-epi-PGF(2α) in DBS was achieved by preconditioning DBS paper with vitamin E and BHT. CONCLUSIONS: The developed method is sensitive, specific, robust, efficient, and can accurately measure endogenous concentrations of 8-epi-PGF(2α) in DBS. Thus, it offers an analytical approach to measure 8-epi-PGF(2α) by a novel sample collection technique that is less invasive and costly than conventional techniques.

Plasma Metabolic Signatures of Healthy Overweight Subjects Challenged With an Oral Glucose Tolerance Test.

Insulin secretion following ingestion of a carbohydrate load affects a multitude of metabolic pathways that simultaneously change direction and quantity of interorgan fluxes of sugars, lipids and amino acids. In the present study, we aimed at identifying markers associated with differential responses to an OGTT a population of healthy adults. By use of three metabolite profiling platforms, we assessed these postprandial responses of a total of 202 metabolites in plasma of 72 healthy volunteers undergoing comprehensive phenotyping and of which half enrolled into a weight-loss program over a three-month period. A standard oral glucose tolerance test (OGTT) served as dietary challenge test to identify changes in postprandial metabolite profiles. Despite classified as healthy according to WHO criteria, two discrete clusters (A and B) were identified based on the postprandial glucose profiles with a balanced distribution of volunteers based on gender and other measures. Cluster A individuals displayed 26% higher postprandial glucose levels, delayed glucose clearance and increased fasting plasma concentrations of more than 20 known biomarkers of insulin resistance and diabetes previously identified in large cohort studies. The volunteers identified by canonical postprandial responses that form cluster A may be called pre-pre-diabetics and defined as "at risk" for development of insulin resistance. Moreover, postprandial changes in selected fatty acids and complex lipids, bile acids, amino acids, acylcarnitines and sugars like mannose revealed marked differences in the responses seen in cluster A and cluster B individuals that sustained over the entire challenge test period of 240 min. Almost all metabolites, including glucose and insulin, returned to baseline values at the end of the test (at 240 min), except a variety of amino acids and here those that have been linked to diabetes development. Analysis of the corresponding metabolite profile in a fasting blood sample may therefore allow for early identification of these subjects at risk for insulin resistance without the need to undergo an OGTT.

Interactions of Carbohydrate Intake and Physical Activity with Regulatory Genes Affecting Glycaemia: A Food4Me Study Analysis.

INTRODUCTION: Carbohydrate intake and physical activity are related to glucose homeostasis, both being influenced by individual genetic makeup. However, the interactions between these 2 factors, as affected by genetics, on glycaemia have been scarcely reported. OBJECTIVE: We focused on analysing the interplay between carbohydrate intake and physical activity levels on blood glucose, taking into account a genetic risk score (GRS), based on SNPs related to glucose/energy metabolism. METHODS: A total of 1,271 individuals from the Food4Me cohort, who completed the nutritional intervention, were evaluated at baseline. We collected dietary information by using an online-validated food frequency questionnaire, a questionnaire on physical activity, blood biochemistry by analysis of dried blood spots, and by analysis of selected SNPs. Fifteen out of 31 SNPs, with recognized participation in carbohydrate/energy metabolism, were included in the component analyses. The GRS included risk alleles involved in the control of glycaemia or energy-yielding processes. RESULTS: Data concerning anthropometric, clinical, metabolic, dietary intake, physical activity, and genetics related to blood glucose levels showed expected trends in European individuals of comparable sex and age, being categorized by lifestyle, BMI, and energy/carbohydrate intakes, in this Food4Me population. Blood glucose was inversely associated with physical activity level (ß = -0.041, p = 0.013) and positively correlated with the GRS values (ß = 0.015, p = 0.047). Interestingly, an interaction affecting glycaemia, concerning physical activity level with carbohydrate intake, was found (ß = -0.060, p = 0.033), which also significantly depended on the genetic background (GRS). CONCLUSIONS: The relationships of carbohydrate intake and physical activity are important in understanding glucose homeostasis, where a role for the genetic background should be ascribed.

Plasma Vitamin C and Type 2 Diabetes: Genome-Wide Association Study and Mendelian Randomization Analysis in European Populations.

OBJECTIVE: Higher plasma vitamin C levels are associated with lower type 2 diabetes risk, but whether this association is causal is uncertain. To investigate this, we studied the association of genetically predicted plasma vitamin C with type 2 diabetes. RESEARCH DESIGN AND METHODS: We conducted genome-wide association studies of plasma vitamin C among 52,018 individuals of European ancestry to discover novel genetic variants. We performed Mendelian randomization analyses to estimate the association of genetically predicted differences in plasma vitamin C with type 2 diabetes in up to 80,983 case participants and 842,909 noncase participants. We compared this estimate with the observational association between plasma vitamin C and incident type 2 diabetes, including 8,133 case participants and 11,073 noncase participants. RESULTS: We identified 11 genomic regions associated with plasma vitamin C (P < 5 × 10-8), with the strongest signal at SLC23A1, and 10 novel genetic loci including SLC23A3, CHPT1, BCAS3, SNRPF, RER1, MAF, GSTA5, RGS14, AKT1, and FADS1. Plasma vitamin C was inversely associated with type 2 diabetes (hazard ratio per SD 0.88; 95% CI 0.82, 0.94), but there was no association between genetically predicted plasma vitamin C (excluding FADS1 variant due to its apparent pleiotropic effect) and type 2 diabetes (1.03; 95% CI 0.96, 1.10). CONCLUSIONS: These findings indicate discordance between biochemically measured and genetically predicted plasma vitamin C levels in the association with type 2 diabetes among European populations. The null Mendelian randomization findings provide no strong evidence to suggest the use of vitamin C supplementation for type 2 diabetes prevention.

Hormone-sensitive lipase (HSL) is also a retinyl ester hydrolase: evidence from mice lacking HSL.

Here, we investigated the importance of hormone-sensitive lipase (HSL) as a retinyl ester hydrolase (REH). REH activity was measured in vitro using recombinant HSL and retinyl palmitate. The expression of retinoic acid (RA)-regulated genes and retinoid metabolites were measured in high-fat diet fed HSL-null mice using real-time quantitative PCR and triple-stage liquid chromatography/tandem mass spectrometry, respectively. Age- and gender-matched wild-type littermates were used as controls. The REH activity of rat HSL was found to be higher than that against the hitherto best known HSL substrate, i.e., diacylglycerols. REH activity in white adipose tissue (WAT) of HSL-null mice was completely blunted and accompanied by increased levels of retinyl esters and decreased levels of retinol, retinaldehyde and all-trans RA. Accordingly, genes known to be positively regulated by RA were down-regulated in HSL-null mice, including pRb and RIP140, key factors promoting differentiation into the white over the brown adipocyte lineage. Dietary RA supplementation partly restored WAT mass and the expression of RA-regulated genes in WAT of HSL-null mice. These findings demonstrate the importance of HSL as an REH of adipose tissue and suggest that HSL via this action provides RA and other retinoids for signaling events that are crucial for adipocyte differentiation and lineage commitment.

Association of plasma biomarkers of fruit and vegetable intake with incident type 2 diabetes: EPIC-InterAct case-cohort study in eight European countries.

OBJECTIVE: To investigate the association of plasma vitamin C and carotenoids, as indicators of fruit and vegetable intake, with the risk of type 2 diabetes. DESIGN: Prospective case-cohort study. SETTING: Populations from eight European countries. PARTICIPANTS: 9754 participants with incident type 2 diabetes, and a subcohort of 13 662 individuals from the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort of 340 234 participants: EPIC-InterAct case-cohort study. MAIN OUTCOME MEASURE: Incident type 2 diabetes. RESULTS: In a multivariable adjusted model, higher plasma vitamin C was associated with a lower risk of developing type 2 diabetes (hazard ratio per standard deviation 0.82, 95% confidence interval 0.76 to 0.89). A similar inverse association was shown for total carotenoids (hazard ratio per standard deviation 0.75, 0.68 to 0.82). A composite biomarker score (split into five equal groups), comprising vitamin C and individual carotenoids, was inversely associated with type 2 diabetes with hazard ratios 0.77, 0.66, 0.59, and 0.50 for groups 2-5 compared with group 1 (the lowest group). Self-reported median fruit and vegetable intake was 274 g/day, 396 g/day, and 508 g/day for participants in categories defined by groups 1, 3, and 5 of the composite biomarker score, respectively. One standard deviation difference in the composite biomarker score, equivalent to a 66 (95% confidence interval 61 to 71) g/day difference in total fruit and vegetable intake, was associated with a hazard ratio of 0.75 (0.67 to 0.83). This would be equivalent to an absolute risk reduction of 0.95 per 1000 person years of follow up if achieved across an entire population with the characteristics of the eight European countries included in this analysis. CONCLUSIONS: These findings indicate an inverse association between plasma vitamin C, carotenoids, and their composite biomarker score, and incident type 2 diabetes in different European countries. These biomarkers are objective indicators of fruit and vegetable consumption, and suggest that diets rich in even modestly higher fruit and vegetable consumption could help to prevent development of type 2 diabetes.

Determination of 8-epi PGF(2alpha) concentrations as a biomarker of oxidative stress using triple-stage liquid chromatography/tandem mass spectrometry.

F2-isoprostanes are a family of prostaglandin F2-like compounds that are formed by free-radical-catalyzed peroxidation of arachidonic acid. Several F2-isoprostanes, but in particular 8-epi PGF(2alpha), are widely used as oxidative stress biomarkers. An analytical method based on liquid chromatography with negative electrospray ionization (ESI) coupled to tandem mass spectrometric detection (LC/MS/MS) was developed for the determination of 8-epi PGF(2alpha) concentrations in human plasma, whole blood, erythrocytes and urine. 8-epi PGF(2alpha)-d4, a stable isotope derivative of 8-epi PGF(2alpha), was used as an internal standard (IS). A 50 microL sample was focused on-column and separated on two 3 microm particle size SUPELCOSIL ABZ+Plus HPLC columns (15 cm x 4.6 mm and 7.5 cm x 4.6 mm) connected in series. An Applied Biosystems 4000 Q TRAP LC/MS/MS system with ESI was operated in multiple reaction monitoring (MRM) mode with the precursor-to-product ion transitions m/z 353.4 --> 193.1 (8-epi PGF(2alpha)), 357.4 --> 197.1 (8-epi PGF(2alpha)-d4), used for quantification. The assay was fully validated and found to have adequate accuracy, precision, linearity, sensitivity and selectivity. The mass limit of detection (mLOD) was 1 pg of analyte eluting from the column. The assay has been successfully applied to the analysis of human plasma, whole blood, erythrocytes and urine samples.