Sara Pauwels

Dietary and supplemental maternal methyl-group donor intake and cord blood DNA methylation

ABSTRACT Maternal nutrition is critically involved in the development and health of the fetus. We evaluated maternal methyl-group donor intake through diet (methionine, betaine, choline, folate) and supplementation (folic acid) before and during pregnancy in relation to global DNA methylation and hydroxymethylation and gene specific (IGF2 DMR, DNMT1, LEP, RXRA) cord blood methylation. A total of 115 mother-infant pairs were enrolled in the MAternal Nutrition and Offsprings Epigenome (MANOE) study. The intake of methyl-group donors was assessed using a food-frequency questionnaire. LC-MS/MS and pyrosequencing were used to measure global and gene specific methylation, respectively. Dietary intake of methyl-groups before and during pregnancy was associated with changes in LEP, DNMT1, and RXRA cord blood methylation. Statistically significant higher cord blood LEP methylation was observed when mothers started folic acid supplementation more than 6 months before conception compared with 3–6 months before conception (34.6 ± 6.3% vs. 30.1 ± 3.6%, P = 0.011, LEP CpG1) or no folic acid used before conception (16.2 ± 4.4% vs. 13.9 ± 3%, P = 0.036 for LEP CpG3 and 24.5 ± 3.5% vs. 22.2 ± 3.5%, P = 0.045 for LEP mean CpG). Taking folic acid supplements during the entire pregnancy resulted in statistically significantly higher cord blood RXRA methylation as compared with stopping supplementation in the second trimester (12.3 ± 1.9% vs. 11.1 ± 2%, P = 0.008 for RXRA mean CpG). To conclude, long-term folic acid use before and during pregnancy was associated with higher LEP and RXRA cord blood methylation, respectively. To date, pregnant women are advised to take a folic acid supplement of 400 µg/day from 4 weeks before until 12 weeks of pregnancy. Our results suggest significant epigenetic modifications when taking a folic acid supplement beyond the current advice.

Maternal Methyl-Group Donor Intake and Global DNA (Hydroxy)Methylation before and during Pregnancy

It is still unclear to which extent methyl-group intake during pregnancy can affect maternal global DNA (hydroxyl)methylation. Pregnancy methylation profiling and its link with methyl-group intake in a healthy population could enhance our understanding of the development of pregnancy related disorders. One hundred forty-eight women were enrolled in the MANOE (MAternal Nutrition and Offspring’s Epigenome) study. Thiry-four women were enrolled before pregnancy and 116 during the first trimester of pregnancy. Global DNA (hydroxy)methylation in blood using LC-MS/MS and dietary methyl-group intake (methionine, folate, betaine, and choline) using a food-frequency questionnaire were estimated pre-pregnancy, during each trimester, and at delivery. Global DNA (hydroxy)methylation levels were highest pre-pregnancy and at weeks 18–22 of pregnancy. We observed a positive relation between folic acid and global DNA methylation (p = 0.04) and hydroxymethylation (p = 0.04). A high intake of methionine pre-pregnancy and in the first trimester showed lower (hydroxy)methylation percentage in weeks 11–13 and weeks 18–22, respectively. Choline and betaine intake in the first weeks was negatively associated with hydroxymethylation. Women with a high intake of these three methyl groups in the second and third trimester showed higher hyrdoxymethylation/methylation levels in the third trimester. To conclude, a time trend in DNA (hydroxy)methylation was found and women with higher methyl-group intake showed higher methylation in the third trimester, and not in earlier phases of pregnancy.

Reproducibility and validity of an FFQ to assess usual intake of methyl-group donors.

OBJECTIVE To develop and validate a novel FFQ to assess the daily intake of four methyl-group donors (methionine, choline, betaine and folate). DESIGN The relative validity of the FFQ was assessed by comparison with 7 d estimated diet records (n 80) and its reproducibility was evaluated by repeated administrations 6 weeks apart (n 92). Paired Student t tests were used to compare group means and de-attenuated intra-class correlations to investigate the ability of the FFQ to rank individuals according to their methyl-group donor intake. De-attenuated intra-class correlation coefficients were calculated between the test and reference method for methionine, choline, betaine, folate and the sum of methyl-group donors. The weighted kappa (κ w) was calculated as a measure of tertile agreement. SETTING Flanders, Belgium. SUBJECTS The FFQ was validated among Flemish women of reproductive age (18-35 years). RESULTS The questionnaire had an acceptable ranking ability (r=0·32-0·68; κ w=0·10-0·35), but overestimated the daily intake of folate (280·6 μg v. 212·0 μg) and betaine (179·1 mg v. 147·0 mg) compared with the 7 d estimated diet record. Cross-classification analysis indicated that 20 % (choline) of the participants were grossly misclassified in the validation study. The correlation between repeated administrations was good (r=0·62-0·83) with a maximal misclassification of 7 % for betaine (κ w=0·44-0·66). CONCLUSIONS These results indicate that this newly developed FFQ is a reliable instrument with acceptable validity for ranking individuals according to methyl-group donor intake (except for a poor agreement for choline (κ w=0·10) and a fair ranking ability for betaine (r=0·32)) in Flemish women of reproductive age.

Validation of a food-frequency questionnaire assessment of methyl-group donors using estimated diet records and plasma biomarkers: the method of triads

Abstract In this study, the intake of methyl-group donors (methionine, folate, betaine, and choline) among Flemish women of reproductive age (n = 30) assessed by a 7-d estimated diet record (7 d EDR) and food-frequency questionnaire (FFQ) was compared with plasma S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), and SAH:SAM ratio. Pearson correlation coefficients were calculated between each of the dietary methods and the validity coefficient was calculated using the method of triads. Correlations were higher between intake assessed by the FFQ and biomarkers than between 7 d EDR and biomarkers. The validity coefficients of the FFQ, when using SAH as a biomarker, were high (0.86 for methionine to 0.94 for folate), when the SAH:SAM ratio was used as a biomarker the validity coefficients ranged from 0.63 to 1.00. These data indicate that the FFQ is a reliable tool to estimate the intake of the methyl-group donors in women of reproductive age.

The effect of paternal methyl-group donor intake on offspring DNA methylation and birth weight

Most nutritional studies on the development of children focus on mother-infant interactions. Maternal nutrition is critically involved in the growth and development of the fetus, but what about the father? The aim is to investigate the effects of paternal methyl-group donor intake (methionine, folate, betaine, choline) on paternal and offspring global DNA (hydroxy)methylation, offspring IGF2 DMR DNA methylation, and birth weight. Questionnaires, 7-day estimated dietary records, whole blood samples, and anthropometric measurements from 74 fathers were obtained. A total of 51 cord blood samples were collected and birth weight was obtained. DNA methylation status was measured using liquid chromatography-tandem mass spectrometry (global DNA (hydroxy)methylation) and pyrosequencing (IGF2 DMR methylation). Paternal betaine intake was positively associated with paternal global DNA hydroxymethylation (0.028% per 100 mg betaine increase, 95% CI: 0.003, 0.053, P=0.03) and cord blood global DNA methylation (0.679% per 100 mg betaine increase, 95% CI: 0.057, 1.302, P=0.03). Paternal methionine intake was positively associated with CpG1 (0.336% per 100 mg methionine increase, 95% CI: 0.103, 0.569, P=0.006), and mean CpG (0.201% per 100 mg methionine increase, 95% CI: 0.001, 0.402, P=0.049) methylation of the IGF2 DMR in cord blood. Further, a negative association between birth weight/birth weight-for-gestational age z-score and paternal betaine/methionine intake was found. In addition, a positive association between choline and birth weight/birth weight-for-gestational age z-score was also observed. Our data indicate a potential impact of paternal methyl-group donor intake on paternal global DNA hydroxymethylation, offspring global and IGF2 DMR DNA methylation, and prenatal growth.

The Maternal Nutrition and Offspring's Epigenome (MANOE) Study: a prospective, monocentric, observational study

Epigenetic modifications have the ability to change the susceptibility to metabolic diseases like obesity. DNA methylation can change during a life course due to environmental exposures like diet.

532 Shedding a light on grey literature searches for occupational health topics: a belgian case study on chemicals exposure

Introduction For occupational health topics with hardly any published white literature available, grey literature can be a generous information source. This abstract describes the search and use of grey literature in preparation of the PROBE (Hazardous chemical Products Register for Occupational use in Belgium) study, aiming to map both occupational exposure to chemicals in Belgian workers and the need for knowledge about such exposure. Methods A cascade of methods was applied. First, relevant associations, organisations, agencies and bodies were identified through interviews with field experts and general internet search engines. Then, specific domains within Google Advanced Search were applied to geographically limit the results to Europe and Belgium. As quality filters, the domain limits org, edu and gov were applied. A second approach consisted in specific grey literature gateways. Finally, references in retrieved documents were explored for additional information sources. Results This multifaceted approach generated a comprehensive overview of evidence based data. The compiled information can be categorised as databases with exposure data and chemical risk assessments, data from similar research in other countries, methodological insights in chemicals selection and exposure surveillance techniques, interim reports of ongoing research, reports, white papers, and legislation. The pathway of grey literature databases was abandoned, as its literature was outdated. The retrieved information provided us with the necessary acumen in the selection of relevant chemicals and appropriate assessment strategies to strengthen the proposed study protocol. Discussion A grey literature search is a challenging and lengthy process as the information is dispersed, hard to access and fragmented. The standard review methods for white literature do not apply to grey literature searches. The complex architecture of grey literature requires an innovative, creative and iterative approach. Nevertheless we succeeded in tapping valuable information from this source. Further initiative is needed to improve grey information availability and retrieval.

915 Maternal occupation is associated with maternal global dna (hydroxy) methylation in the second trimester of pregnancy

Introduction Environmental factors, such as nutrition and occupational exposure can influence epigenetic marks like DNA methylation, which play a role in the development of chronic diseases. Methods Data of the MAternal Nutrition and Offspring’s Epigenome (MANOE) study was used to assess the effect of maternal occupation on maternal and infant DNA (hydroxy)methylation levels. Mothers were categorised in job categories according to the International Standard Classification of Occupations (ISCO). Maternal global DNA (hydroxy)methylation levels during each trimester of pregnancy and at delivery (n=122) was measured in whole blood via LC-MS/MS. Data were analysed with a one-Way ANOVA. Results We found statistically significant differences in maternal global DNA methylation (p=0.008) and global DNA hydroxymethylation (p=0.004) at 20 weeks of pregnancy. Post hoc tests revealed that global DNA methylation and global DNA hydroxymethylation level was significantly lower when the mother had an intellectual/scientific/artistic profession (6.36% and 0.13%) as opposed to being a manager (7.77%, p=0.007% and 0.22%, p=0.002) or administrative staff (7.71%, p=0.003% and 0.2%, p=0.005). No significant differences between different working groups were found for global DNA (hydroxy)methylation in the first and third trimester of pregnancy and at delivery. Conclusion The mother’s occupation was associated with maternal global DNA (hydroxy)methylation levels only in the second trimester of pregnancy. The change in maternal global DNA (hydroxy)methylation in the second trimester of pregnancy could be due to hormonal changes during pregnancy, a shift in the one-carbon metabolism in the middle of pregnancy, but based on these results we also have to take into account maternal occupational exposure.

520 Probe: hazardous chemical products register for occupational use in belgium

During their job, workers are exposed to a wide variety of working conditions including chemical substances that are potentially detrimental to employees’ health. Today, Belgian data on occupational exposure to dangerous chemicals are collected by Occupational Health Services (OHS) merely for the purpose of assuring the appropriate health screening. This makes these data of little use for epidemiological research and exposure surveillance on one hand and for policy development by competent authorities on the other hand. The PROBE (Hazardous chemical Products Register for Occupational use in Belgium) study is set up to investigate the exposure of Belgian workers to dangerous chemical products, including type, duration and frequency of exposure. PROBE consists of a systematic collection and analysis of occupational chemical exposure data. A trained, motivated, and representative sample of occupational physicians from both internal and external OHS will be invited to participate. The data will be collected on a regular basis over a period of 5 months. Besides demographics, exposure measurements and health related data will be collected. First, a pilot will be kicked off in a limited sample of occupational physicians, testing the feasibility of the program. The final goal of the project is to register in a comprehensive but easy way the exposure to dangerous chemicals in order to improve preventive measures, to ensure workers’ health and to develop a national surveillance policy.