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Objectives: The aim of this study is to present age- and sex-specific reference values of insulin, glucose, glycosylated haemoglobin (HbA1c) and the homeostasis model assessment to quantify insulin resistance (HOMA-IR) for pre-pubertal children. Methods: The reference population consists of 7074 normal weight 3- to 10.9-year-old pre-pubertal children from eight European countries who participated in at least one wave of the IDEFICS (‘identification and prevention of dietary- and lifestyle-induced health effects in children and infants’) surveys (2007–2010) and for whom standardised laboratory measurements were obtained. Percentile curves of insulin (measured by an electrochemiluminescence immunoassay), glucose, HbA1c and HOMA-IR were calculated as a function of age stratified by sex using the general additive model for location scale and shape (GAMLSS) method. Results: Levels of insulin, fasting glucose and HOMA-IR continuously show an increasing trend with age, whereas HbA1c shows an upward trend only beyond the age of 8 years. Insulin and HOMA-IR values are higher in girls of all age groups, whereas glucose values are slightly higher in boys. Median serum levels of insulin range from 17.4 and 13.2 pmol l −1 in 3–<3.5-year-old girls and boys, respectively, to 53.5 and 43.0 pmol l −1 in 10.5–<11-year-old girls and boys. Median values of glucose are 4.3 and 4.5 mmol l −1 in the youngest age group and 49.3 and 50.6 mmol l −1 in the oldest girls and boys. For HOMA-IR, median values range from 0.5 and 0.4 in 3–<3.5-year-old girls and boys to 1.7 and 1.4 in 10.5–<11-year-old girls and boys, respectively. Conclusions: Our study provides the first standardised reference values for an international European children’s population and provides the, up to now, largest data set of healthy pre-pubertal children to model reference percentiles for markers of insulin resistance. Our cohort shows higher values of Hb1Ac as compared with a single Swedish study while our percentiles for the other glucose metabolic markers are in good accordance with previous studies.

Objectives: The development of effective strategies to prevent childhood obesity and its comorbidities requires new, reliable early biomarkers. Here, we aimed to identify in peripheral blood cells potential transcript-based biomarkers of unhealthy metabolic profile associated to overweight/obesity in children. Methods: We performed a whole-genome microarray analysis in blood cells to identify genes differentially expressed between overweight and normal weight children to obtain novel transcript-based biomarkers predictive of metabolic complications. Results: The most significant enriched pathway of differentially expressed genes was related to oxidative phosphorylation, for which most of genes were downregulated in overweight versus normal weight children. Other genes were involved in carbohydrate metabolism/glucose homoeostasis or in lipid metabolism (for example, TCF7L2, ADRB3, LIPE, GIPR), revealing plausible mechanisms according to existing biological knowledge. A set of differentially expressed genes was identified to discriminate in overweight children those with high or low triglyceride levels. Conclusions: Functional microarray analysis has revealed a set of potential blood-cell transcript-based biomarkers that may be a useful approach for early identification of children with higher predisposition to obesity-related metabolic alterations.

Objectives: C-reactive protein (CRP) is involved in a wide range of diseases. It is a powerful marker for inflammatory processes used for diagnostic and monitoring purposes. We aimed to establish reference values as data on the distribution of serum CRP levels in young European children are scarce. Subjects: Reference values of high-sensitivity CRP concentrations were calculated for 9855 children aged 2.0–10.9 years, stratified by age and sex. The children were recruited during the population-based European IDEFICS study (Identification and prevention of Dietary- and lifestyle-induced health Effects in Children and infantS) with 18 745 participants recruited from 2007 to 2010. Results: In 44.1 % of the children, CRP values were below or equal the detection limit of 0.2 mg/l. Median CRP concentrations showed a slight negative age trend in boys and girls, whereas serum CRP values were slightly higher in girls than in boys across all age groups. Conclusions: Our population-based reference values of CRP may guide paediatric practice as elevated values may require further investigation or treatment. Therefore, the presented reference values represent a basis for clinical evaluation and for future research on risk assessment of diseases associated with increased CRP levels among children.

Background: Measuring dietary intake in children is notoriously difficult. Therefore, it is crucial to evaluate the performance of dietary intake assessment methods in children. Given the important contribution of milk consumption to calcium (Ca) and potassium (K) intakes, urinary calcium (UCa) and potassium (UK) excretions in spot urine samples could be used for estimating correlations with milk consumption frequencies. Objective: The aim of this study was to evaluate the assessment of milk consumption frequencies derived from the Food Frequency Questionnaire section of the Children's Eating Habits Questionnaire (CEHQ-FFQ) used in the IDEFICS (Identification and prevention of dietary- and lifestyle induced health effects in children and infants) study by comparing with UCa and UK excretions in spot urine samples. Design: This study was conducted as a setting-based community-oriented intervention study and results from the first cross-sectional survey have been included in the analysis. Subjects: A total of 10 309 children aged 2–10 years from eight European countries are included in this analysis. Methods: UCa and UK excretions were measured in morning spot urine samples. Calcium and potassium urine concentrations were standardised for urinary creatinine (Cr) excretion. Ratios of UCa/Cr and UK/Cr were used for multivariate regression analyses after logarithmic transformation to obtain normal distributions of data. Milk consumption frequencies were obtained from the CEHQ-FFQ. Multivariate regression analyses were used to investigate the effect of milk consumption frequencies on UCa and UK concentrations, adjusting for age, gender, study centre, soft drink consumption and frequency of main meals consumed at home. Results: A significant positive correlation was found between milk consumption frequencies and ratios of UK/Cr and a weaker but still significant positive correlation with ratios of UCa/Cr, when using crude or partial Spearman's correlations. Multivariate regression analyses showed that milk consumption frequencies were predictive of UCa/Cr and UK/Cr ratios, when adjusted for age, gender, study centre, soft drink consumption and frequency of main meals consumed at home. Mean ratios of UK/Cr for increasing milk consumption frequency tertiles showed a progressive increase in UK/Cr. Children consuming at least two milk servings per day had significantly higher mean UCa/Cr and UK/Cr ratios than children who did not. Large differences in correlations between milk consumption frequencies and ratios of UCa/Cr and UK/Cr were found between the different study centres. Conclusion: Higher milk consumption frequencies resulted in a progressive increase in UK/Cr and UCa/Cr ratios, reflecting the higher Ca and K intakes that coincide with increasing milk consumption, which constitutes a major K and Ca source in children's diet.

Large scale international multicentre studies require sophisticated quality management for the collection, processing and logistics of biological samples to ensure a maximum degree of standardisation across different environmental conditions and settings. This paper describes a quality management system for the collection of biological samples (QMS-BS) which was applied during IDEFICS, a large European multicentre study. The application was evaluated by several criteria like response rates for the different types of biological samples, measures of sample quality, compliance with the QMS-BS and efficiency of the document and sample control and of the quality assurance system. Response rates varied from 56.6% for venous blood collection to 90.1% for saliva collection. All sample types were associated with problems of sample quality (e.g. haemolysis of blood samples, lack of cooling for urine samples or desiccation of saliva samples). Overall compliance with the QMS-BS was good, with some exceptions mainly related to sample control. In conclusion the QMS-BS is a valuable tool for the management of biological sample collection in epidemiological multicentre studies.

Background: Measurement of cholesterol and triglyceride (TG) fractions in blood has become standard practice in the early detection of atherosclerotic disease pathways. Considerable attention is given nowadays to the presence of these risk factors in children and to start preventive campaigns early in life. In this context, it is imperative to have valid comparative frameworks for interpretation of lipid levels. The aim of this study is to present sex- and age-specific reference values on blood lipid levels in European children aged 2.0–10.9 years. Methods: Fasting blood was obtained via either venipuncture or capillary sampling. In 13 579 European non-obese children (50.3% boys), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), TG and TC/HDL-C ratio levels were measured with a point-of-care analyser (Cholestech). Sex- and age-specific reference values were computed with the GAMLSS method with the statistical software R. Results: Reference curves and 1st, 3rd, 10th, 25th, 50th, 75th, 90th, 97th and 99th percentile values are presented. HDL-C showed a positive trend with age, from 2 years onwards, but was relatively stable above the age of 7. For LDL-C and TC, linear but small age-related trends were seen. The TC/HDL-C values showed a gradual negative trend from the age of 2 up to 6 and were relatively stable afterwards. For TG, no age trend was found ( P =0.285). Boys had higher mean HDL-C values than girls (1.414 vs 1.368 mmol l −1 ), and lower TC, LDL-C, TC/HDL-C and TG values (3.981 vs 4.087 mmol l −1 ; 2.297 vs 2.435 mmol l −1 ; 2.84 vs 3.01mmol l −1 ; and 0.509 vs 0.542 mmol l −1 , respectively). Conclusions: These new and recent references could serve as a European orientation of blood lipid values in children in the context of standard medical practice and for the purpose of public health screening.

Objective: To evaluate the influence of a standardised sampling protocol and process quality across the different IDEFICS (Identification and prevention of dietary- and lifestyle-induced health effects in children and infants) centres on the results of the biochemical measurements. Design: Baseline survey within the community-based intervention study. Subjects: A total of 16 224 children, aged 2–8 years, enrolled in the IDEFICS baseline survey in 8 European countries. Venous or capillary blood samples were collected from 12 430 children, urine samples from 13 890 children and saliva samples from 14 019 children. Methods: A set of quality indicators was recorded for the biological blood, urine and saliva samples collected during the IDEFICS study. Results of blood and urine measurements were analysed and stratified by selected quality indicators. Results: Concentrations of biological markers in blood and urine measured during the IDEFICS baseline survey are associated with several quality indicators assessed in this study. Between-country variations of these biomarkers are described. It was confirmed that fasting has a big influence on the concentration of certain biomarkers. Biomarkers in morning urine samples may be erroneous if the study subjects void during the night or if samples are not taken from the very first morning urine. Conclusions: The analysed data underline that a standardised sampling protocol is of major importance, especially in multicentre studies, but non-compliance is ever present in spite of well-defined standard operation procedures. Deviations from the protocol should therefore always be documented to avoid error pertaining to the concentration of biological markers.