Alcohol and delirium tremens: effects of average number of drinks per day and beverage type.

OBJECTIVE: Associations of amount of alcohol intake and beverage type with the risk of delirium tremens (DT) have not been studied. This longitudinal study investigated if the average number of drinks per day and beverage type predict DT. METHODS: A cohort of 3 582 alcohol-dependent men and women aged 19-82 without previous DT were interviewed about alcohol intake and beverage type at baseline in 1994-2005 and followed through record linkage in Danish nationwide registers to identify incident DT. Data were analyzed by means of Cox regression models. RESULTS: An average number of drinks per day of 20-30 or >30 was associated with hazard ratios (HRs) of 1.38 (95% CI 1.03-1.84) and 1.64 (95% CI 1.19-2.27) relative to the reference category (1-9 drinks). Independently of amount consumed and covariates (age, gender, civil status and work status), beverage type (spirits vs. mixed alcohol) was associated with a HR of 1.63 (95% CI 1.08-2.46). Male gender was robustly associated with increased risk (HR = 1.62 (95% CI 1.25-2.08). CONCLUSIONS: In alcohol-dependent men and women, daily alcohol intake above a threshold of 20 beverages or 240 g alcohol and a preference for spirits increase the risk of developing DT.

Parental alcohol use disorder with and without other mental disorders and offspring alcohol use disorder.

OBJECTIVE: To examine the association between parental alcohol use disorder (AUD) with and without other mental disorders and offspring AUD. METHODS: Using data from Danish nationwide registers, we identified 15 477 offspring with parental AUD and 154 392 reference individuals from the general population. Parental AUD was defined as registration for AUD treatment. Parental mental disorders were identified in medical registers and comprised psychotic, mood, anxiety, personality, drug use, and other non-alcohol-related mental disorders. AUD in offspring was identified from medical, pharmacy, treatment and cause of death registers. Hazard ratios (HRs) of AUD were estimated using Cox regression models. RESULTS: AUD in one or both parents was associated with higher risks of AUD in offspring compared with reference individuals. Paternal AUD plus other mental disorder (HR = 2.27, 95% confidence interval (CI): 2.10-2.46) and paternal AUD alone (HR = 2.21, 95% CI: 2.07-2.36) were associated with higher offspring AUD risk. Similarly, maternal AUD plus other mental disorder (HR = 3.02, 95% CI: 2.66-3.43) and maternal AUD alone (HR = 2.57, 95% CI: 2.20-3.01) were associated with higher offspring AUD risk. CONCLUSIONS: Offspring with parental AUD are at increased risk of AUD irrespective of exposure to other parental mental disorders.

Optimisation of asymmetric flow field-flow fractionation for the characterisation of nanoparticles in coated polydisperse TiO2 with applications in food and feed.

Titanium dioxide (TiO2) has various applications in consumer products and is also used as an additive in food and feeding stuffs. For the characterisation of this product, including the determination of nanoparticles, there is a strong need for the availability of corresponding methods of analysis. This paper presents an optimisation process for the characterisation of polydisperse-coated TiO2 nanoparticles. As a first step, probe ultrasonication was optimised using a central composite design in which the amplitude and time were the selected variables to disperse, i.e., to break up agglomerates and/or aggregates of the material. The results showed that high amplitudes (60%) favoured a better dispersion and time was fixed in mid-values (5 min). In a next step, key factors of asymmetric flow field-flow fraction (AF4), namely cross-flow (CF), detector flow (DF), exponential decay of the cross-flow (CFexp) and focus time (Ft), were studied through experimental design. Firstly, a full-factorial design was employed to establish the statistically significant factors (p < 0.05). Then, the information obtained from the full-factorial design was utilised by applying a central composite design to obtain the following optimum conditions of the system: CF, 1.6 ml min-1; DF, 0.4 ml min-1; Ft, 5 min; and CFexp, 0.6. Once the optimum conditions were obtained, the stability of the dispersed sample was measured for 24 h by analysing 10 replicates with AF4 in order to assess the performance of the optimised dispersion protocol. Finally, the recovery of the optimised method, particle shape and particle size distribution were estimated.

5-HT2B receptor antagonists attenuate myofibroblast differentiation and subsequent fibrotic responses in vitro and in vivo.

Pulmonary fibrosis is characterized by excessive accumulation of connective tissue, along with activated extracellular matrix (ECM)-producing cells, myofibroblasts. The pathological mechanisms are not well known, however serotonin (5-HT) and 5-HT class 2 (5-HT2) receptors have been associated with fibrosis. The aim of the present study was to investigate the role of 5-HT2B receptors in fibrosis, using small molecular 5-HT2B receptor antagonists EXT5 and EXT9, with slightly different receptor affinity. Myofibroblast differentiation [production of alpha-smooth muscle actin (α-SMA)] and ECM synthesis were quantified in vitro, and the effects of the receptor antagonists were evaluated. Pulmonary fibrosis was also modeled in mice by subcutaneous bleomycin administrations (under light isoflurane anesthesia), and the effects of receptor antagonists on tissue density, collagen-producing cells, myofibroblasts and decorin expression were investigated. In addition, cytokine expression was analyzed in serum. Lung fibroblasts displayed an increased α-SMA (P < 0.05) and total proteoglycan production (P < 0.01) when cultured with TGF-ß1 together with 5-HT, which were significantly reduced with both receptor antagonists. Following treatment with EXT5 or EXT9, tissue density, expression of decorin, number of collagen-producing cells, and myofibroblasts were significantly decreased in vivo compared to bleomycin-treated mice. Receptor antagonization also significantly reduced systemic levels of TNF-α and IL-1ß, indicating a role in systemic inflammation. In conclusion, 5-HT2B receptor antagonists have potential to prevent myofibroblast differentiation, in vitro and in vivo, with subsequent effect on matrix deposition. The attenuating effects of 5-HT2B receptor antagonists on fibrotic tissue remodeling suggest these receptors as novel targets for the treatment of pulmonary fibrosis.

Effect of a high-protein diet on maintenance of blood pressure levels achieved after initial weight loss: the DiOGenes randomized study.

Randomized trials have shown significant blood pressure (BP) reductions after increased protein compared with carbohydrate intake, but the effect on BP maintenance after initial weight loss is unclear. We examined the effect of a high-protein diet on the maintenance of reduced BP after weight loss in 420 overweight adults from the Diet, Obesity and Genes study. After an 8-week weight-loss period (>8% BW), subjects (42±6 years) were randomized to either a high-protein diet (23-28 en% protein) or a lower-protein control diet (10-15 en% protein) for 26 weeks. BMI after weight loss was 30.3±4.3 kg m(-2), BP was 118/73 mm Hg and 28 subjects (6.5%) used antihypertensive agents. Systolic BP during 26 weeks of weight maintenance dietary intervention increased in both treatment groups, but it was 2.2 mm Hg less (95% CI: -4.6 to 0.2 mm Hg, P=0.08) in the high-protein group than in the lower-protein control group. In 191 (pre)hypertensive subjects (baseline systolic BP⩾120 mm Hg), a larger difference was observed (-4.2 mm Hg (-7.7, -0.7), P=0.02). The effect was attenuated after adjustment for initial BP (-3.4 mm Hg (-6.9, -0.03), P=0.048), and after additional adjustment for weight change (-2.7 mm Hg (-6.1, 0.4), P=0.11). Adjustment for 24-h urinary excretion of sodium and potassium did not change the results. Diastolic BP yielded similar results. These findings suggest that a BP reduction after weight loss is better maintained when the intake of protein is increased at the expense of carbohydrates. This effect is partly mediated by body weight.

Long-term weight changes in obese young adult men and subsequent all-cause mortality.

BACKGROUND: Although the expectation is that weight gain increases mortality and weight loss among those overweight reduces mortality, results on weight gain and mortality in young adults are conflicting, and weight loss is less explored. We investigated the association between long-term weight change and all-cause mortality in a broad range of body mass index (BMI) in young men. METHODS: Among 362200 Danish draftees, examined between 1943 and 1977, all obese (BMI 31.0 kg m(-2); n=1930), and a random 1% sample of the others (n=3601) were identified at a mean age of 20 years (range: 18-25 years). All the obese and half the controls were re-examined between 4 and 40 years later (mean age 35 years). Weight changes were defined as: weight loss <-0.1 kg m(-2) per year, weight stability within ±0.1 kg m(-2) per year and weight gain >0.1 kg m(-2) per year. Hazard ratios (HR) and 95% confidence intervals (CI) were estimated using Cox regression. RESULTS: Among the 908 obese and 1073 controls followed for 30 years after re-examination 220 and 232 died. HR of the weight stable obese was 2.32 (CI: 1.56-3.44) compared with the weight stable controls. In the obese cohort there was no association between weight loss, adjusted for initial BMI, and mortality (HR: 0.99; CI: 0.68-1.45) compared with weight stable obese. Too few controls lost weight to allow assessment of weight loss. Weight gain was associated with increased mortality in the obese (HR: 1.50; CI: 1.07-2.10) and controls (HR: 1.54; CI: 1.14-2.09) compared with weight stable obese and controls, respectively. Neither the time between the two examinations, life-style factors nor exclusion of diseased individuals influenced the results. CONCLUSIONS: Although there were increased mortality of the weight-stable obese compared with controls, there was no association between weight loss and mortality in the obese. Weight gain increased mortality regardless of the initial weight.

Validation of a near infrared microscopy method for the detection of animal products in feedingstuffs: results of a collaborative study.

The performance characteristics of a near infrared microscopy (NIRM) method, when applied to the detection of animal products in feedingstuffs, were determined via a collaborative study. The method delivers qualitative results in terms of the presence or absence of animal particles in feed and differentiates animal from vegetable feed ingredients on the basis of the evaluation of near infrared spectra obtained from individual particles present in the sample. The specificity ranged from 86% to 100%. The limit of detection obtained on the analysis of the sediment fraction, prepared as for the European official method, was 0.1% processed animal proteins (PAPs) in feed, since all laboratories correctly identified the positive samples. This limit has to be increased up to 2% for the analysis of samples which are not sedimented. The required sensitivity for the official control is therefore achieved in the analysis of the sediment fraction of the samples where the method can be applied for the detection of the presence of animal meal. Criteria for the classification of samples, when fewer than five spectra are found, as being of animal origin needs to be set up in order to harmonise the approach taken by the laboratories when applying NIRM for the detection of the presence of animal meal in feed.

Weight and weight gain during early infancy predict childhood obesity: a case-cohort study.

BACKGROUND: Infant weight and weight gain are positively associated with later obesity, but whether there is a particular critical time during infancy remains uncertain. OBJECTIVE: The aim was to investigate when and how weight and weight gain during infancy become associated with childhood obesity. METHODS: In a cohort representing 28 340 children born from 1959-67 and measured in Copenhagen schools, 962 obese children (2007 World Health Organization criteria), were compared with a 5% randomly selected sub-cohort of 1417 children. Information on weight at birth, 2 weeks, 1, 2, 3, 4, 6 and 9 months was retrieved from health visitors' records. Odds ratios and 95% confidence intervals (CI) for childhood obesity by tertiles of weight at each age and by change in tertiles of weight between two consecutive measurements were estimated using multivariate logistic regression with adjustment for indicators of socioeconomic status, preterm birth, and breastfeeding. RESULTS: Compared with children in the middle weight-tertile, children with a weight in the upper tertile had a 1.36-fold (CI, 1.10-1.69) to 1.72-fold (CI, 1.36-2.18) higher risk of childhood obesity from birth through 9 months, whereas children in the lower weight-tertile had almost half the risk of obesity from 2 through 9 months. The risk of childhood obesity associated with change in weight-tertile in each interval was stable at ∼1.5-fold per weight-tertile increase throughout infancy. CONCLUSIONS: Infant weight and weight gain are associated with obesity in childhood already during the first months of life. Determinants of weight gain shortly after birth may be a suitable target for prevention of obesity.

Circulating ACE is a predictor of weight loss maintenance not only in overweight and obese women, but also in men.

BACKGROUND: Circulating angiotensin-converting enzyme (ACE) was identified as a predictor of weight loss maintenance in overweight/obese women of the Diogenes project. OBJECTIVE: To investigate whether ACE acted also as a predictor in men of the Diogenes study and to compare it with that in women. DESIGN: Subjects, who lost ≥ 8% of body weight induced by low-caloric diet in an 8-week weight loss period, were assigned to weight loss maintenance with dietary intervention for 6 months. SUBJECTS: 125 overweight/obese healthy men from eight European countries who completed whole intervention. MEASUREMENTS: Concentrations and activity of serum ACE at baseline and after the 8-week weight loss, in addition to anthropometric and physiological parameters. RESULTS: Serum ACE concentration decreased by 11.3 ± 10.6% during the weight loss period in men. A greater reduction is associated with less body weight regain during the maintenance period (r=0.227, P=0.012). ACE change was able to predict a weight regain ≤ 20% after 6 months, with an odds ratio of 1.59 (95% confidence interval (CI): 1.09-2.33, P=0.016) for every 10% reduction, which was independent of body mass index and weight loss. The prediction power was weaker in men than in women, but without a significant sex difference (P=0.137). In pooled subjects (N=218), the odds ratio was 1.96 (95% CI: 1.46-2.64, P<0.001). CONCLUSIONS: A greater reduction of ACE during weight loss is favorable for weight maintenance in both men and women. This can offer useful information for personalized advice to improve weight loss maintenance. It also confirms the role of ACE in the metabolic pathways of weight regulation.

Lifelong doubling of mortality in men entering adult life as obese.

BACKGROUND: The association between obesity in adults and excess morbidity and mortality is well established, but the impact of being obese in early adulthood on health throughout adult life needs elucidation. We investigated the all-cause mortality until 80 years of age in men starting adult life as obese. METHODS: Among 362,200 Danish young men, examined for military service between 1943 and 1977, all obese (defined as body mass index (BMI ≥ 31.0 kg m(-2)), and, as controls, a random 1% sample of the remaining population were identified. A total of 1862 obese, corresponding to all men above the 99.5 percentile in this population, and 3476 controls were included, at a median age of 19 years (range: 18-25 years of age). They were followed until 2007 and Cox regression models were used to estimate the mortality in the obese relative to the controls. In addition, two reference groups were used: normal weight men (BMI: 18.5-24.9 kg m(-2)) and the men with the lowest mortality in this cohort (BMI: 22.0-24.9 kg m(-2)). RESULTS: During the 65 years of follow-up, 1191 men died. At all ages from 18 to 80 years, the mortality in the obese was twice that of the controls (hazard ratio (HR): 2.10; 95% confidence interval (CI): 1.84-2.39). The median survival proportion (0.5) was reached about 8 years earlier in the obese than in either of the reference groups. Relative to the normal weight and men with the lowest mortality HRs of 2.14 (95% CI: 1.86-2.45) and 2.38 (95% CI: 2.00-2.85), respectively, were estimated for the obese. Neither year of birth nor education significantly influenced the excess mortality. CONCLUSION: Men entering adult life as obese experience a lifelong doubling of mortality, a finding that strongly supports the continued need to avoid beginning adult life as obese.

Inter-laboratory comparison of an analytical method for the determination of the feed additive semduramicin in poultry feed at authorised level by liquid chromatography single quadrupole mass spectrometry.

A collaborative study was carried out according to internationally recognised guidelines in order to establish the performance characteristics of an LC/MS method for the determination of the feed additive semduramicin (SEM) in poultry feed at the level (20-25 mg kg(-1)) authorised within the European Union. Fifteen laboratories participated in the validation study, and all reported results. The content of SEM in the tested materials, provided as blind duplicates, ranged from 11.5 mg kg(-1), which corresponds to half the mean authorised level, to 45.0 mg kg(-1), which corresponds to twice the mean authorised level. All the materials were analysed by the participating laboratories using two different quantification approaches: standard addition and external standard calibration. The relative standard deviation of reproducibility (RSD(R)) for both quantification approaches varied from 8% to 18%, corresponding to HORRAT values ranging from 0.8 to 1.5, which were therefore in all cases below the critical value of 2.0. Consequently, the proposed analytical method and both quantification approaches can be considered to be fully validated and transferable to the control laboratories and applied for the determination of SEM in poultry compound feed at authorised level within the frame of official control. Further steps in the administrative procedure aiming to adopt the method as part of an ISO/CEN standard are currently ongoing.

Predictors of weight loss maintenance and attrition during a 6-month dietary intervention period: results from the DiOGenes study.

There is a need to develop tools to predict individual weight loss maintenance and attrition prognosis. We aimed to identify predictors of weight loss maintenance outcome and attrition in subjects from eight European countries in the DiOGenes project. A total of 932 overweight/obese subjects (body mass index: 27-42 kg m(-2) ) were enrolled in an 8-week low-calorie diet (LCD). The 776 subjects (83%) who achieved at least 8% reduction in their initial body weight were randomized into five dietary arms varying in protein content and glycemic index for a 6-month weight maintenance period. Baseline characteristics, weight loss at weeks 1, 3 and 8 of LCD were assessed as predictors of weight loss maintenance and attrition using multivariate regression and correlation models. The multivariate model showed that the 6-month weight loss maintenance was predicted by: 7.889 - 0.343 × weight loss at week 3 + 1.505 × weight loss at week 8 + 2.422 × gender (0 = male and 1 = female gender) (R(2) = 51%, P = 0.0001). A greater weight loss at week 8 was associated with a lower attrition during the subsequent 6-month dietary intervention period (OR = 0.92, 95% CI: 0.88-0.97, P = 0.001). Furthermore, the men showed an increased likelihood for attrition during the dietary intervention period (OR = 1.54, 95% CI: 1.07-2.20, P = 0.02). A greater weight loss during 8 weeks of LCD and female gender predict better 6-month weight maintenance of weight loss, whereas the baseline characteristics did not predict outcome. Attrition could be strongly predicted by gender and weight loss during LCD.

Apoptosis induced by the potential chemotherapeutic drug N1, N11-Diethylnorspermine in a neuroblastoma cell line.

Neuroblastoma is a highly malignant neoplasm found in young children. Although children with high-risk neuroblastoma respond to chemotherapy, relapses are common. On account of poor treatment outcome, new treatment strategies are constantly sought for neuroblastoma. Polyamine analogues are potentially novel substances for treatment of neuroblastoma. In this study, we have treated two neuroblastoma cell lines, SH-SY5Y and LA-N-1, with the spermine analogue N1, N11-Diethylnorspermine (DENSPM). SH-SY5Y was the most sensitive cell line, in which DENSPM treatment resulted in an inhibition of cell proliferation and an induction of cell death. The cell death induced by DENSPM treatment was apoptotic, as evidenced by cleavage of procaspase 3 and induction of caspase-3 activity. In contrast, DENSPM treatment only resulted in a slight inhibition of cell proliferation in LA-N-1 cells. There were several possible causes for the lower sensitivity to DENSPM treatment in the latter cell line when compared with SH-SY5Y cells. DENSPM-induced polyamine depletion was more extensive in SH-SY5Y cells than in LA-N-1 cells. This was partly because of a higher induction of the polyamine catabolic enzyme spermidine/spermine N1-acetyltransferase in the cell line SH-SY5Y. The DENSPM-induced polyamine depletion was also caused by the inhibition of ornithine decarboxylase. LA-N-1 cells contained a higher level of the prosurvival protein survivin, which was further increased after DENSPM treatment. In contrast, DENSPM treatment resulted in a decreased survivin level in SH-SY5Y cells.

Initial weight loss on an 800-kcal diet as a predictor of weight loss success after 8 weeks: the Diogenes study.

OBJECTIVE: The purpose of this study was to investigate whether pre-treatment subject characteristics and weight change during the first weeks of a low-calorie diet (LCD) can predict weight loss outcomes at the end of a controlled 8-week weight loss period in overweight and obese adults. SUBJECTS: A total of 932 overweight and obese subjects of both genders were included at eight European centers, and underwent an 8-week LCD period. RESULTS: The weight loss at week 8 was positively correlated with initial body weight (Spearman's rho=0.62), height (rho=0.43), body mass index (rho=0.43), waist (rho=0.48) and hip circumference (rho=0.33), sagittal diameter (rho=0.45), fat mass (rho=0.35) and fat-free mass (rho=0.52), and gender (rho=-0.36) (all P<0.01). In the multivariate regression model, adjusted for center, only initial body weight, early weight loss (week 1) and weight loss at week 3 were significant predictors of weight loss outcome at week 8: weight loss (kg) at week 8=0.09+0.046 x baseline body weight (kg)-0.311 x weight loss (kg) at week 1+1.284 x weight loss (kg) at week 3 (R(2)=68%, P<0.0001). A weight loss of > or =2.6 kg at week 1 during the LCD period was identified as the optimal cut-off predictor for at least 10 kg weight loss at week 8. CONCLUSIONS: This study suggests that initial body weight, early weight loss (week 1) and weight loss at week 3 are predictors of final weight loss during an 8-week LCD, and may be used as early biomarkers of subsequent responses to an LCD diet.

Key parameters for the development of a NIR microscopic method for the quantification of processed by-products of animal origin in compound feedingstuffs.

The aim of this work is to show new advances in the analytical methods developed in the frame of the ban of processed animal by-products in compound feed that is currently applied within the European Union. With this aim, studies to develop a quantitative near infrared microscopy (NIRM) approach have been undertaken in order to fulfil future requirements of European legislation like the introduction of tolerance levels that would require for official control purposes the availability of specific quantitative methods. The capabilities of the NIRM method have been improved; no sample preparation is required and the acquisition parameters are optimised. Both the gross and the fine fractions of the samples are considered; the reflexion mode was used to analyse the gross raw fraction and the transmission mode was chosen to analyse the fine raw fraction. Parameters for reflexion analyses were already fixed in our previous studies while those of transmission mode have been determined in the present study. Because particles are too small, it is difficult to mark them; spectra were collected using the mapping technique. Quantitative analyses have been carried out for different percentages of adulteration (0.5, 1, 2 and 5%). Results were depending on the particle size distribution of the feed and of the fish meal which led to experimental values of adulteration varying between 0.13-0.92%, 0.93-3.7%, 2.42-5.83% and 1.95-9.39% for theoretical percentages of adulteration equal to 0.5, 1, 2 and 5%, respectively. The established protocol with the key parameters proposed has to be considered for the development of an accurate method of quantification.

eNOS phosphorylation and translocation are altered in male but not female mice by increased activation of the Gαq protein.

Little is known about sex-dependent physiological and pathophysiological differences in cardiac endothelial nitric oxide synthase (eNOS) expression and activation. Therefore, we investigated cardiac morphology and eNOS protein expression, including its translocation-dependent activation and phosphorylation, in cardiac tissue of male and female wild-type mice and transgenic heart-failure mice having a cardiac-specific, 5-fold overexpression of the Galphaq protein. In addition, we measured calcineurin protein expression. Heart-to-body weight ratio was increased in Galphaq mice. Female wild-type mice showed higher eNOS protein expression and activation (translocation and phosphorylation) than did wild-type males. In cardiac tissue of Galphaq mice, these sex-dependent differences remained or were enhanced. Protein expression of the catalytic subunit calcineurin A, which has been shown to dephosphorylate eNOS, was higher in wild-type males than in wild-type females. These differences were increased in the Galphaq mice model. We conclude that sex differences exist in cardiac eNOS protein expression and phosphorylation. Increased activation of the Galphaq protein appears to alter eNOS protein expression and phosphorylation only in males.

Validation of an analytical method for the determination of glyceroltriheptanoate (GTH) in processed animal by-products: results of a collaborative study.

The performance characteristics of a method for the determination of the marker substance glycerol triheptanoate (GTH) in processed animal by-products (ABPs) based on gas chromatography (GC) coupled to mass spectrometry (MS) were determined via a collaborative study. Within the European Union, GTH needs to be added to the portion of processed ABPs that must not enter the feed and food chain (Categories 1 and 2) at a minimum concentration of 250 mg kg(-1) related to the fat fraction of the test samples analysed. The test materials included in the validation study consisted of three meat and bone meal (MBM) and three fat samples that contained GTH at different concentrations ranging from 61 to 455 mg kg(-1). The relative standard deviation of repeatability (RSD(r)) varied from 3.4 to 7.8% and the relative standard deviation of reproducibility (RSD(R)) varied from 9.0 to 16.5%, corresponding to HORRAT values that were, in all cases, equal or below the critical value of 2.0. The estimated trueness expressed in terms of average concentration compared to the target concentrations of GTH in all test materials varied from 95 to 107%, confirming acceptable values for the trueness of the method. Based on the acceptable values for precision and trueness, the method is fit for the intended purpose and can be used for official control purposes to determine GTH in processed animal by-products from Category 1 and Category 2.

Common variants near MC4R in relation to body fat, body fat distribution, metabolic traits and energy expenditure.

OBJECTIVE: Common variants near melanocortin receptor 4 (MC4R) have been related to fatness and type 2 diabetes. We examined the associations of rs17782313 and rs17700633 in relation to body fat, body fat distribution, metabolic traits, weight development and energy expenditure. METHODS: Obese young men (n = 753, BMI > or = 31.0 kg m(-2)) and a randomly selected group (n = 874) identified from a population of 174 800 men were re-examined in three surveys at mean ages 35, 46 and 49 years (S-35, S-46 and S-49). Measurements were available at upto eight times from birth to adulthood. Logistic regression analysis was used to assess odds ratio (OR) for the presence of the carrier allele for a given difference in phenotypic values. RESULTS: Rs17782313 minor C-allele was associated with overall, abdominal and peripheral fatness (range of OR = 1.06-1.14 per z-score units) at all three surveys, although only consistently significant at S-35 and S-46. Rs17700633 minor A-allele was also associated with the fatness measures, but significantly so only at S-49 for overall and abdominal fatness (range of OR = 1.03-1.15 per z-score units), and peripheral fatness (OR = 1.15-1.20 per z-score units). There were only few significant associations with metabolic traits. The rs17782313 C-allele and the rs17700633 A-allele were both associated with lower high-density lipoprotein cholesterol (range of OR = 0.64-0.84 per mol l(-1)), significantly at S-46. The rs17700633 A-allele was significantly associated with insulin (OR = 1.25 per 50 pmol l(-1)), leptin (OR = 1.42 per 10 ng microl(-1)) and insulin sensitivity (OR = 0.81 per model unit). The rs17782313 C-allele and the rs17700633 A-allele were both associated with BMI in childhood and adolescence (range of OR = 1.04-1.17 per z-score units), significant for the rs17782313 C-allele at the age of 13-19 years and for rs17700633 A-allele at age 7, 10, 13 and 19 years. No significant associations were found for energy expenditure. CONCLUSION: Near MC4R variants appear to contribute to body fat, body fat distribution, some metabolic traits, weight development during childhood, but not to energy expenditure.

The Diet, Obesity and Genes (Diogenes) Dietary Study in eight European countries - a comprehensive design for long-term intervention.

Diogenes is a Pan-European, randomized, controlled dietary intervention study investigating the effects of dietary protein and glycaemic index on weight (re)gain, metabolic and cardiovascular risk factors in obese and overweight families in eight European centres. The article is methodological in character, and the presentation of 'results' will be limited to baseline characteristics of the study populations included. A total of 891 families with at least one overweight/obese parent underwent screening. The parents started an initial 8-week low-calorie diet and families with minimum one parent attaining a weight loss of > or = 8%, were randomized to one of five energy ad libitum, low-fat (25-30 E%) diets for 6 or 12 months: low protein/low glycaemic index, low protein/high glycaemic index, high protein/low glycaemic index, high protein/high glycaemic index or control (national dietary guidelines). At two centres the families were provided dietary instruction plus free foods for 6 months followed by 6-month dietary instruction only. At the remaining six centres the families received dietary instruction only for 6 months. The median weight loss during the low-calorie diet was 10.3 kg (inter-quartile range: 8.7-12.8 kg, n = 775). A total of 773 adults and 784 children were randomized to the 6-month weight (re)gain prevention phase. Despite major cultural and dietary regional differences in Europe, interventions addressing effects of dietary factors are feasible with a reasonable attrition.

Use of glyceroltriheptanoate as marker for processed animal by-products: development and validation of an analytical method.

A recently published European Regulation requires that the artificial marker, glycerol triheptanoate (GTH), be added to processed animal by-product (ABPs) prohibited from entering the food chain. The objective of this new requirement is to allow full traceability and ensure that these materials are disposed of in a proper way. Here, we report the development and single-laboratory validation of an analytical method for the determination of GTH in meat and bone meal plus animal fat. The method comprises three steps: (1) extraction of GTH from the samples with petroleum ether when analysing meat and bone meal or dissolving the sample in n-hexane when analysing fat; (2) clean-up of the extract using commercially available SPE cartridges; (3) determination of GTH by GC/MS or GC with flame ionisation detection (FID). The results of the validation study demonstrated that the relative standard for intermediate precision varied between 2.5 and 8.2%, depending on GTH concentration and the detector utilised. In all cases, the relative recovery rate was above 96%. The limit of quantification was 16 mg kg(-1) (GTH/fat content of the sample) with MS as detector and 20 mg kg(-1) with FID. Moreover, the method has been successfully applied in a second laboratory, indicating its transferability. Considering the minimum GTH concentration in ABPs of 250 mg kg(-1), the method is considered suitable for the intended purpose and can be utilised by EU Member States laboratories for official control and monitoring.