Annunziata Nappo

Design and results of the pretest of the IDEFICS study

Background:During the preparatory phase of the baseline survey of the IDEFICS (Identification and prevention of dietary- and lifestyle-induced health effects in children and infants) study, standardised survey procedures including instruments, examinations, methods, biological sampling and software tools were developed and pretested for their feasibility, robustness and acceptability.Methods:A pretest was conducted of full survey procedures in 119 children aged 2–9 years in nine European survey centres (N per centre=4–27, mean 13.22). Novel techniques such as ultrasound measurements to assess subcutaneous fat and bone health, heart rate monitors combined with accelerometers and sensory taste perception tests were used.Results:Biological sampling, physical examinations, sensory taste perception tests, parental questionnaire and medical interview required only minor amendments, whereas physical fitness tests required major adaptations. Callipers for skinfold measurements were favoured over ultrasonography, as the latter showed only a low-to-modest agreement with calliper measurements (correlation coefficients of r=−0.22 and r=0.67 for all children). The combination of accelerometers with heart rate monitors was feasible in school children only. Implementation of the computer-based 24-h dietary recall required a complex and intensive developmental stage. It was combined with the assessment of school meals, which was changed after the pretest from portion weighing to the more feasible observation of the consumed portion size per child. The inclusion of heel ultrasonometry as an indicator of bone stiffness was the most important amendment after the pretest.Discussion:Feasibility and acceptability of all procedures had to be balanced against their scientific value. Extensive pretesting, training and subsequent refinement of the methods were necessary to assess the feasibility of all instruments and procedures in routine fieldwork and to exchange or modify procedures that would otherwise give invalid or misleading results.

High‐sensitivity C‐reactive Protein is a Predictive Factor of Adiposity in Children: Results of the Identification and prevention of Dietary‐ and lifestyle‐induced health Effects in Children and InfantS (IDEFICS) Study

Background Whereas cross‐sectional studies have shown that obesity is associated with increased C‐reactive protein (CRP) levels in children, little is known about the impact of low‐grade inflammation on body mass changes during growth. Methods and Results We assessed cross‐sectionally and longitudinally the association of high‐sensitivity (hs)‐CRP levels with overweight/obesity and related cardiometabolic risk factors in the Identification and prevention of Dietary‐ and lifestyle‐induced health Effects in Children and InfantS (IDEFICS) cohort. 16 224 children from 8 European countries (2 to 9 years) were recruited during the baseline survey (T0). After the exclusion of 7187 children because of missing hs‐CRP measurements and 2421 because of drug use during the previous week, the analysis was performed on 6616 children (Boys=3347; Girls=3269; age=6.3±1.7 years). Of them, 4110 were reexamined 2 years later (T1). Anthropometric variables, blood pressure, hs‐CRP, blood lipids, glucose and insulin were measured. The population at T0 was divided into 3 categories, according to the baseline hs‐CRP levels. Higher hs‐CRP levels were associated with significantly higher prevalence of overweight/obesity, body mass index (BMI) z‐score and central adiposity indices (P values all <0.0001), and with higher blood pressure and lower HDL‐cholesterol levels. Over the 2‐year follow‐up, higher baseline hs‐CRP levels were associated with a significant increase in BMI z‐score (P<0.001) and significantly higher risk of incident overweight/obesity. Conclusions Higher hs‐CRP levels are associated to higher body mass and overweight/obesity risk in a large population of European children. Children with higher baseline levels of hs‐CRP had a greater increase in BMI z‐score and central adiposity over time and were at higher risk of developing overweight/obesity during growth.

Antioxidant and cytotoxic properties of lyophilized beer extracts on HL-60 cell line

An impressive number of studies have suggested that red wine can be considered the protective beverage of choice against chronic and degenerative pathologies. Only few and controversial data are available on a potential, similar role for beer, which represents a more cost-effective, safe, and widely available beverage. Starting from the evidence that many antioxidant compounds present in red wine are also present at similar or even higher concentrations in beers, we first screened 48 commercially available beers and selected one (Mrt-HP) with very high polyphenol concentration and antioxidant activity estimated by ferric reducing antioxidant power. We demonstrated that a lyophilized preparation of Mrt-HP beer was cytotoxic with respect to a beer with low polyphenolic content (Trt-LP) when assayed on HL-60 human leukemia cell line. We measured a 60% decrease in cell viability at a polyphenol concentration of 250 μM quercetin equivalents. We also demonstrated that Mrt-HP cytotoxicity was not an artifact due to cell growth conditions because addition of Mrt-HP extracts to cell medium generated peroxide levels indistinguishable from controls. By means of cytofluorimetric analysis of pre-G1 population and caspase 3 activation, we demonstrated that Mrt-HP extracts activated apoptosis in HL-60 cell line. Finally, we found that the concentration of quercetin, resveratrol, and gallic acid in Mrt-HP was 10, 4.6, and 4.6-fold higher, respectively, than in Trt-LP, suggesting that the presence of these molecules might be responsible for the observed cytotoxicity. These data, together with the low in vivo beer toxicity reported in the literature, suggest a possible chemopreventive role for this beverage that requires further studies in animal models.

Physical activity, adiposity and urbanization level in children: results for the Italian cohort of the IDEFICS study

OBJECTIVES While there is extensive evidence about the influence of environmental factors on adult obesity, fewer studies have assessed how the environment influences body fat in children. This cross-sectional study investigated the distribution of adiposity indices according to urbanization level and patterns of physical activity among children in the Italian cohort of the IDEFICS study. METHODS The sample included 1673 preschool and school-aged children (mean age 6.1 years, standard deviation 1.7) living in rural (n = 579), suburban (n = 442) and urban (n = 652) areas. Anthropometric measures were taken and questionnaires were used to assess childrens lifestyles, including patterns of physical activity. RESULTS Children who lived in rural areas spent significantly more time in outdoor activities but participated in less structured physical activity compared with children living in suburban and urban areas. Adiposity estimated by the sum of skinfold thickness increased linearly from rural to urban areas, with results for suburban areas showing intermediate values. CONCLUSIONS The data show that geographical environmental factors influence patterns of physical activity and body fat in children. In particular, the results suggest an association between the time spent in unstructured outdoor activities and the degree of adiposity in schoolchildren. These results may have implications for public health, including efforts to increase freely available playgrounds as an effective measure to counteract the obesity epidemic in children.

Gender-related differences in the relationships between blood pressure, age, and body size in prepubertal children.

BACKGROUND The blood pressure (BP) increase with age is well documented in adults and children. However, in the pediatric age group, body size is the most important determinant of age-related BP increases. The aim of the present analysis was to investigate the relationships between age, gender, and body size and BP in children. METHODS To this purpose, data were analyzed from 4,514 prepubertal children, aged 6-11 years (71% of the eligible sample; boys = 2,283, girls = 2,231) from the ARCA Project, a screening of childhood obesity carried out in southern Italy. Girls who reported the occurrence of menarche were excluded from the analysis. The sample constituted roughly 20% of all the children attending the primary schools in the area. Weight, height, waist circumference, and BP were measured according to standardized procedures. RESULTS As expected, both systolic and diastolic BP significantly increased (P < 0.001) with age in boys and girls. However, after adjustment for waist circumference (as index of adiposity) and height (as index of body size), BP significantly increased with age only in girls (systolic BP: F = 4.380, P = 0.002; diastolic BP: F = 3.093, P = 0.01) but not in boys (systolic BP: F = 0.711, P = 0.55; diastolic BP: F = 2.180, P = 0.07). The association, however, was no longer apparent after the exclusion of children aged >10 years. CONCLUSIONS In prepubertal girls in the age range 6-11 years, but not in boys, age is significantly associated with BP independently of body size and adiposity.

Analysis of the association of leptin and adiponectin concentrations with metabolic syndrome in children: Results from the IDEFICS study

BACKGROUND AND AIMS Adipokines may play a role in the pathogenesis of the metabolic syndrome (MetS) in children. We aimed to evaluate the association of leptin, adiponectin, and its ratio (L/A ratio) with the metabolic syndrome (MetS) in a subsample of the IDEFICS (Identification and prevention of Dietary- and lifestyle-induced health EFfects In Children and infantS) cohort. METHODS AND RESULTS Leptin, adiponectin and MetS parameters were measured in a subsample of 1253 children (3-9.9 years) participating to the IDEFICS study, grouped as: Non-OW (underweight/normal weight) and OW/Ob (overweight/obese). MetS was defined using the sex- and age-specific cut-offs based on the distribution of MetS components in the IDEFICS cohort. The prevalence of the MetS among OW/Ob was 24.8% and 27.1% in boys and girls respectively, whereas ≤2% among Non-OW. OW/Ob had significantly higher leptin and L/A ratio as compared to Non-OW. Significantly higher leptin was found in OW/Ob with MetS as compared with OW/Ob without MetS. Significantly lower adiponectin was observed only in OW/Ob girls as compared to Non-OW. A 1SD increase in leptin and L/A ratio z-scores or a 1SD decrease in adiponectin z-score were significantly associated with higher risk of MetS. After adjustment for BMI or body fat mass (BFM) the association remained significant only for leptin. CONCLUSION We showed that in European children, higher leptin concentration is associated with MetS, even after adjusting for BMI or BFM, confirming an early role of leptin in MetS, while the association of adiponectin with MetS seems be mediated by body fat in this age range.

C-reactive protein reference percentiles among pre-adolescent children in Europe based on the IDEFICS study population

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.

PROTECTIVE EFFECT OF γ-IRRADIATION AGAINST HYPOCHLOROUS ACID- INDUCED HAEMOLYSIS IN HUMAN ERYTHROCYTES

Radiations may trigger protective response within a threshold of doses applied. Exposures above an upper threshold are generally detrimental, while exposures below a lower threshold may or may not increase risks for health. We recently reported that a cellular protective response occurs in interventional cardiologists to counteract the oxidative damage caused by radiation. Here, we demonstrated in an in vitro model represented by whole blood of healthy donors γ-irradiated with 220–440 mGy, that haemolysis of erythrocytes induced by hypochlorous acid was reduced by 40%. The protection triggered by γ-radiations made erythrocytes more resistant to oxidative insult caused by hypochlorous acid which was induced 3 h after irradiation and involved biochemical changes in the synthesis and turnover of glutathione. Overall, the biochemical remodelling induced by exposure to γ-radiations might contribute to generate new guidelines in professionally exposed workers.

Process Evaluation of the IDEFICS Intervention

Process evaluation is an essential part of intervention evaluation that is often overlooked or reported limitedly. This chapter explains in detail how the process evaluation within the IDEFICS study was developed and how the different measures were built. For each intervention module, different measures were developed to integrate multiple perspectives on the implementation. All measures were based on the framework of Linnan and Steckler (2002) to ensure a theory- and evidence-based approach. As process measures used in research are seldom reported in manuscripts, most of the measures were newly developed with the IDEFICS study itself. The measures itself are added to the chapter to share these instruments with other researchers so that they can build on our measures to develop their own.

140 O - Molecular analysis of APC mutations in familial adenomatous polyposis

Recently we identified three mutations in the APC gene, the insertion of one base, the deletion of 4 bp and the deletion of 2 bp in exons 5, 14 and 15, respectively, in FAP patients from the Neapolitan area. In this study we report two novel germ-line mutations, identified by means of PCR-SSCP analysis, protein truncation test (PTT) and direct sequencing: the deletion of an A residue at nucleotide 2638, and the insertion of an A residue after nucleotide 2803, both in exon 15. These mutations can be analysed by restriction enzyme digestion. Our data are in agreement and enlarge the notion that the majority of APC mutations occur in exon 15 and result in an early stop codon, thus giving rise to a truncated protein. Furthermore, in the course of screening of the APC gene, we have identified a new polymorphism in exon 15, fragment L, that can be used in cosegregation studies for the presymptomatic diagnosis of FAP and for family studies. This work was supported by grants from: M.U.R.S.T.; C.N.R. (P.F. Biotechnology and Bioinstrumentations, and P.F. Genetic Engineering), (Rome) and from AIRC (Milan).