Syddansk Universitet Exploring a causal role of DNA methylation in the relationship between maternal vitamin B12 during pregnancy and child's IQ at age 8, cognitive performance and educational attainment A two-step Mendelian randomization study

An adequate intake of vitamin B12 during pregnancy plays an important role in offspring neurodevelopment, potentially via epigenetic processes. We used a two-step Mendelian randomization approach to assess whether DNA methylation plays a mediating and causal role in associations between maternal vitamin B12 status and offspring’s cognition. Firstly, we estimated the causal effect of maternal vitamin B12 levels on cord blood DNA methylation using the maternal FUT2 genotypes rs492602:A>G and rs1047781:A>T as proxies for circulating vitamin B12 levels in the Avon Longitudinal Study of Parents and Children (ALSPAC) and we tested the observed associations in a replication cohort. Secondly, we estimated the causal effect of DNA methylation on IQ using the offspring genotype at sites close to the methylated CpG site as a proxy for DNA methylation in ALSPAC and in a replication sample. The first step Mendelian randomization estimated that maternal vitamin B12 had a small causal effect on DNA methylation in offspring at three CpG sites, which was replicated for one of the sites. The second step Mendelian randomization found weak evidence of a causal effect of DNA methylation at two of these sites on childhood performance IQ which was replicated for one of the sites. The findings support a causal effect of maternal vitamin B12 levels on cord blood DNA methylation, and a causal effect of vitamin B12-responsive DNA methylation changes on children’s cognition. Some limitations were identified and future studies using a similar approach should aim to overcome such issues. Received: February 15, 2017. Revised: April 7, 2017. Accepted: April 24, 2017 VC The Author 2017. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. 3001 Human Molecular Genetics, 2017, Vol. 26, No. 15 3001–3013 doi: 10.1093/hmg/ddx164 Advance Access Publication Date: 27 April 2017 Association Studies Article

[1]  R. Bosch,et al.  Effects of maternal vitamin B12 supplementation on early infant neurocognitive outcomes: a randomized controlled clinical trial , 2017, Maternal & child nutrition.

[2]  J. Kleinman,et al.  Genetic and epigenetic analysis of schizophrenia in blood—a no-brainer? , 2016, Genome Medicine.

[3]  Robin M. Murray,et al.  An integrated genetic-epigenetic analysis of schizophrenia: evidence for co-localization of genetic associations and differential DNA methylation , 2016, Genome Biology.

[4]  N. Wray,et al.  Predicting gene targets from integrative analyses of summary data from GWAS and eQTL studies for 28 human complex traits , 2016, Genome Medicine.

[5]  J. Mill,et al.  Epigenetic Research in Neuropsychiatric Disorders: the “Tissue Issue” , 2016, Current Behavioral Neuroscience Reports.

[6]  G. Davey Smith,et al.  Challenges and novel approaches for investigating molecular mediation , 2016, Human molecular genetics.

[7]  Daniel R Weinberger,et al.  Association of DNA Methylation Differences With Schizophrenia in an Epigenome-Wide Association Study. , 2016, JAMA psychiatry.

[8]  Tom R. Gaunt,et al.  Systematic identification of genetic influences on methylation across the human life course , 2016, Genome Biology.

[9]  Shan V Andrews,et al.  DNA methylation of cord blood cell types: Applications for mixed cell birth studies , 2016, Epigenetics.

[10]  P. Visscher,et al.  Integration of summary data from GWAS and eQTL studies predicts complex trait gene targets , 2016, Nature Genetics.

[11]  A. Hofman,et al.  Prenatal folate, homocysteine and vitamin B12 levels and child brain volumes, cognitive development and psychological functioning: the Generation R Study , 2016, British Journal of Nutrition.

[12]  Allyson L. Lister,et al.  Data Resource Profile: Accessible Resource for Integrated Epigenomic Studies (ARIES). , 2015, International journal of epidemiology.

[13]  Lorna M. Lopez,et al.  Modulation of Genetic Associations with Serum Urate Levels by Body-Mass-Index in Humans , 2015, PloS one.

[14]  Juan Zhang,et al.  Cholesterol metabolism and homeostasis in the brain , 2015, Protein & Cell.

[15]  Ian J. Deary,et al.  Common genetic variants associated with cognitive performance identified using the proxy-phenotype method , 2014, Proceedings of the National Academy of Sciences.

[16]  G. Davey Smith,et al.  Mendelian randomization: genetic anchors for causal inference in epidemiological studies , 2014, Human molecular genetics.

[17]  Y. Jacquemyn,et al.  Vitamin B12 and folic acid status of term pregnant women and newborns in the Antwerp region, Belgium. , 2014, Clinical and experimental obstetrics & gynecology.

[18]  M. Szyf,et al.  Association of Childhood Chronic Physical Aggression with a DNA Methylation Signature in Adult Human T Cells , 2014, PloS one.

[19]  R. Freire,et al.  USP29 controls the stability of checkpoint adaptor Claspin by deubiquitination , 2014, Oncogene.

[20]  T. VanderWeele,et al.  Mendelian randomization in health research: Using appropriate genetic variants and avoiding biased estimates☆ , 2014, Economics and human biology.

[21]  P. Visscher,et al.  Childhood intelligence is heritable, highly polygenic and associated with FNBP1L , 2014, Molecular Psychiatry.

[22]  George Davey Smith,et al.  Maternal iron levels early in pregnancy are not associated with offspring IQ score at age 8, findings from a Mendelian randomization study , 2014, European Journal of Clinical Nutrition.

[23]  Peggy Hall,et al.  The NHGRI GWAS Catalog, a curated resource of SNP-trait associations , 2013, Nucleic Acids Res..

[24]  Alexandra M. Binder,et al.  The causal effect of red blood cell folate on genome-wide methylation in cord blood: a Mendelian randomization approach , 2013, BMC Bioinformatics.

[25]  Patrick T. Davies,et al.  Parental Depressive Symptoms and Adolescent Adjustment: Responses to Children's Distress and Representations of Attachment as Explanatory Mechanisms , 2013, Parenting, science and practice.

[26]  S. Ring,et al.  Prenatal alcohol exposure and offspring cognition and school performance. A ‘Mendelian randomization’ natural experiment , 2013, International journal of epidemiology.

[27]  S. Thompson,et al.  Use of allele scores as instrumental variables for Mendelian randomization , 2013, International journal of epidemiology.

[28]  B. Pierce,et al.  Efficient Design for Mendelian Randomization Studies: Subsample and 2-Sample Instrumental Variable Estimators , 2013, American journal of epidemiology.

[29]  Jonathan P. Beauchamp,et al.  GWAS of 126,559 Individuals Identifies Genetic Variants Associated with Educational Attainment , 2013, Science.

[30]  D. Gudbjartsson,et al.  Genetic Architecture of Vitamin B12 and Folate Levels Uncovered Applying Deeply Sequenced Large Datasets , 2013, PLoS genetics.

[31]  Ruth Pidsley,et al.  A data-driven approach to preprocessing Illumina 450K methylation array data , 2013, BMC Genomics.

[32]  R. Plomin,et al.  Methylomic analysis of monozygotic twins discordant for autism spectrum disorder and related behavioural traits , 2013, Molecular Psychiatry.

[33]  Per Magnus,et al.  Association between maternal use of folic acid supplements and risk of autism spectrum disorders in children. , 2013, JAMA.

[34]  D. Lawlor,et al.  Vitamin B-12 Status during Pregnancy and Child’s IQ at Age 8: A Mendelian Randomization Study in the Avon Longitudinal Study of Parents and Children , 2012, PloS one.

[35]  David P. Friedman,et al.  The Signature of Maternal Rearing in the Methylome in Rhesus Macaque Prefrontal Cortex and T Cells , 2012, The Journal of Neuroscience.

[36]  Kenny Q. Ye,et al.  An integrated map of genetic variation from 1,092 human genomes , 2012, Nature.

[37]  D. Lawlor,et al.  Maternal and offspring fasting glucose and type 2 diabetes-associated genetic variants and cognitive function at age 8: a Mendelian randomization study in the Avon Longitudinal Study of Parents and Children , 2012, BMC Medical Genetics.

[38]  M. Gillman,et al.  Maternal intake of methyl-donor nutrients and child cognition at 3 years of age. , 2012, Paediatric and perinatal epidemiology.

[39]  M. Szyf,et al.  Peripheral SLC6A4 DNA Methylation Is Associated with In Vivo Measures of Human Brain Serotonin Synthesis and Childhood Physical Aggression , 2012, PloS one.

[40]  R. Dobson,et al.  Functional annotation of the human brain methylome identifies tissue-specific epigenetic variation across brain and blood , 2012, Genome Biology.

[41]  J. Tost,et al.  Complete pipeline for Infinium(®) Human Methylation 450K BeadChip data processing using subset quantile normalization for accurate DNA methylation estimation. , 2012, Epigenomics.

[42]  J. Trent,et al.  Genome-wide association study identifies novel loci associated with serum level of vitamin B12 in Chinese men. , 2012, Human molecular genetics.

[43]  Richard T. Barfield,et al.  CpGassoc: an R function for analysis of DNA methylation microarray data , 2012, Bioinform..

[44]  D. Lawlor,et al.  Cohort Profile: The ‘Children of the 90s’—the index offspring of the Avon Longitudinal Study of Parents and Children , 2012, International journal of epidemiology.

[45]  D. Lawlor,et al.  Cohort Profile: The Avon Longitudinal Study of Parents and Children: ALSPAC mothers cohort , 2012, International journal of epidemiology.

[46]  C. Potter,et al.  Genetic and Non-Genetic Influences during Pregnancy on Infant Global and Site Specific DNA Methylation: Role for Folate Gene Variants and Vitamin B12 , 2012, PloS one.

[47]  Gusztav Belteki,et al.  Periconceptional maternal micronutrient supplementation is associated with widespread gender related changes in the epigenome: a study of a unique resource in the Gambia. , 2012, Human molecular genetics.

[48]  G. Davey Smith,et al.  Two-step epigenetic Mendelian randomization: a strategy for establishing the causal role of epigenetic processes in pathways to disease. , 2012, International journal of epidemiology.

[49]  E. Susser,et al.  Folic acid supplements in pregnancy and severe language delay in children. , 2011, JAMA.

[50]  Torben Hansen,et al.  Genome-Wide Population-Based Association Study of Extremely Overweight Young Adults – The GOYA Study , 2011, PloS one.

[51]  Ricardo Araya,et al.  Smoking Is Associated with, but Does Not Cause, Depressed Mood in Pregnancy – A Mendelian Randomization Study , 2011, PloS one.

[52]  Xiao Zhang,et al.  Comparison of Beta-value and M-value methods for quantifying methylation levels by microarray analysis , 2010, BMC Bioinformatics.

[53]  G. Wang,et al.  Relationship of folate, vitamin B12 and methylation of insulin-like growth factor-II in maternal and cord blood , 2010, European Journal of Clinical Nutrition.

[54]  Donald W. Bowden,et al.  Candidate genes for non-diabetic ESRD in African Americans: a genome-wide association study using pooled DNA , 2010, Human Genetics.

[55]  Fang Wang,et al.  Relation between hypomethylation of long interspersed nucleotide elements and risk of neural tube defects. , 2010, The American journal of clinical nutrition.

[56]  N. Timpson,et al.  Severe Obesity in Young Women and Reproductive Health: The Danish National Birth Cohort , 2009, PloS one.

[57]  S. Chanock,et al.  Genome-wide significant predictors of metabolites in the one-carbon metabolism pathway. , 2009, Human molecular genetics.

[58]  Toshiko Tanaka,et al.  Genome-wide association study of vitamin B6, vitamin B12, folate, and homocysteine blood concentrations. , 2009, American journal of human genetics.

[59]  J. Troendle,et al.  Maternal Vitamin B12 Status and Risk of Neural Tube Defects in a Population With High Neural Tube Defect Prevalence and No Folic Acid Fortification , 2009, Pediatrics.

[60]  D. Bhat,et al.  Vitamin B12 Status of Pregnant Indian Women and Cognitive Function in their 9-year-old Children , 2008, Food and nutrition bulletin.

[61]  S. Chanock,et al.  Common variants of FUT2 are associated with plasma vitamin B12 levels , 2008, Nature Genetics.

[62]  J. Heron,et al.  A United Kingdom Population-Based Study of Intellectual Capacities in Children With and Without Soiling, Daytime Wetting, and Bed-wetting , 2007, Pediatrics.

[63]  D. Lawlor,et al.  Re: Estimation of bias in nongenetic observational studies using "Mendelian triangulation" by Bautista et al. , 2007, Annals of epidemiology.

[64]  R. Obeid,et al.  Biochemical indexes of the B vitamins in cord serum are predicted by maternal B vitamin status. , 2005, The American journal of clinical nutrition.

[65]  M. Pearce,et al.  Gene–gene interaction in folate-related genes and risk of neural tube defects in a UK population , 2004, Journal of Medical Genetics.

[66]  K. Binks,et al.  Low erythrocyte folate status and polymorphic variation in folate-related genes are associated with risk of neural tube defect pregnancy. , 2004, Molecular genetics and metabolism.

[67]  S. Ebrahim,et al.  'Mendelian randomization': can genetic epidemiology contribute to understanding environmental determinants of disease? , 2003, International journal of epidemiology.

[68]  Peter B. Jones,et al.  Gene expression analysis in schizophrenia: Reproducible up-regulation of several members of the apolipoprotein L family located in a high-susceptibility locus for schizophrenia on chromosome 22 , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[69]  M. Ashburner,et al.  Gene Ontology: tool for the unification of biology , 2000, Nature Genetics.

[70]  A. Czeizel,et al.  Prevention of the first occurrence of neural-tube defects by periconceptional vitamin supplementation. , 1992, The New England journal of medicine.

[71]  M. Gnant,et al.  Prevention of neural tube defects: Results of the Medical Research Council Vitamin Study , 1991, The Lancet.

[72]  R. Berry,et al.  Periconceptional use of multivitamins and the occurrence of neural tube defects. , 1989, JAMA.

[73]  D. Kitay.  FOLIC ACID AND REPRODUCTION , 1979, Clinical obstetrics and gynecology.

[74]  E. Rush,et al.  Vitamin B12: one carbon metabolism, fetal growth and programming for chronic disease , 2014, European Journal of Clinical Nutrition.

[75]  David Wechsler,et al.  Manual for the Wechsler intelligence scale for children , 1974 .