title: Stability of the human sperm DNA methylome to folic acid fortification and short-term supplementation.
abstract: Do short-term and long-term exposures to low-dose folic acid supplementation alter DNA methylation in sperm?
  
  
  
  No alterations in sperm DNA methylation patterns were found following the administration of low-dose folic acid supplements of 400 μg/day for 90 days (short-term exposure) or when pre-fortification of food with folic acid and post-fortification sperm samples (long-term exposure) were compared.
  
  
  
  Excess dietary folate may be detrimental to health and DNA methylation profiles due to folate's role in one-carbon metabolism and the formation of S-adenosyl methionine, the universal methyl donor. DNA methylation patterns are established in developing male germ cells and have been suggested to be affected by high-dose (5 mg/day) folic acid supplementation.
  
  
  
  This is a control versus treatment study where genome-wide sperm DNA methylation patterns were examined prior to fortification of food (1996-1997) in men with no history of infertility at baseline and following 90-day exposure to placebo (n = 9) or supplement containing 400 μg folic acid/day (n = 10). Additionally, pre-fortification sperm DNA methylation profiles (n = 19) were compared with those of a group of post-fortification (post-2004) men (n = 8) who had been exposed for several years to dietary folic acid fortification.
  
  
  
  Blood and seminal plasma folate levels were measured in participants before and following the 90-day treatment with placebo or supplement. Sperm DNA methylation was assessed using the whole-genome and genome-wide techniques, MassArray epityper, restriction landmark genomic scanning, methyl-CpG immunoprecipitation and Illumina HumanMethylation450 Bead Array.
  
  
  
  Following treatment, supplemented individuals had significantly higher levels of blood and seminal plasma folates compared to placebo. Initial first-generation genome-wide analyses of sperm DNA methylation showed little evidence of changes when comparing pre- and post-treatment samples. With Illumina HumanMethylation450 BeadChip arrays, no significant changes were observed in individual probes following low-level supplementation; when compared with those of the post-fortification cohort, there were also few differences in methylation despite exposure to years of fortified foods.
  
  
  
  Illumina HumanMethylation450 BeadChip data from this study have been submitted to the NCBI Gene Expression Omnibus under the accession number GSE89781.
  
  
  
  This study was limited to the number of participants available in each cohort, in particular those who were not exposed to early (pre-1998) fortification of food with folic acid. While genome-wide DNA methylation was assessed with several techniques that targeted genic and CpG-rich regions, intergenic regions were less well interrogated.
  
  
  
  Overall, our findings provide evidence that short-term exposure to low-dose folic acid supplements of 400 μg/day, over a period of 3 months, a duration of time that might occur during infertility treatments, has no major impact on the sperm DNA methylome.
  
  
  
  This work was supported by a grant to J.M.T. from the Canadian Institutes of Health Research (CIHR: MOP-89944). The authors have no conflicts of interest to declare.
authors.affiliation: Departments of Pediatrics and of Human Genetics of McGill University, Montreal Children's Hospital and Research Institute of the McGill University Health Centre, Glen Site, 1001 Décarie Boulevard Block E- Room EM0.2236 Montreal, QC H4A 3J1, Canada.
authors.forename: D
authors.initials: D
authors.lastname: Chan
authors.affiliation: Department of Obstetrics and Gynaecology, Biochemistry and Molecular Medicine of Université de Montréal, Research Center of the Sainte-Justine University Hospital, Montreal, QC H3T 1C5, Canada.
authors.forename: S
authors.initials: S
authors.lastname: McGraw
authors.affiliation: Lady Davis Institute for Medical Research, Jewish General Hospital, Montreal, QC H3T 1E2, Canada.
authors.forename: K
authors.initials: K
authors.lastname: Klein
authors.affiliation: Department of Biological Sciences, Diablo Valley College, Pleasant Hill, CA 94523, USA.
authors.forename: L M
authors.initials: LM
authors.lastname: Wallock
authors.affiliation: Department of Epigenomics and Cancer Risk Factors, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Konermann
authors.affiliation: Department of Epigenomics and Cancer Risk Factors, German Cancer Research Center (DKFZ), Heidelberg 69120, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Plass
authors.affiliation: Royal Victoria Hospital of the McGill University Health Centre and Department of Surgery, McGill University, Montréal, QC H4A 3J1, Canada.
authors.forename: P
authors.initials: P
authors.lastname: Chan
authors.affiliation: Department of Pharmacology and Therapeutics, McGill University, Montréal, QC H3A 0G4, Canada.
authors.forename: B
authors.initials: B
authors.lastname: Robaire
authors.affiliation: USDA, Agricultural Research Service, Western Human Nutrition Research Center, Davis, CA 95616, USA.
authors.forename: R A
authors.initials: RA
authors.lastname: Jacob
authors.affiliation: Lady Davis Institute for Medical Research, Jewish General Hospital, Montreal, QC H3T 1E2, Canada.|Departments of Oncology, Epidemiology, Biostatistics and Occupational Health, and Human Genetics, McGill University, Montréal, QC H4A 3J1, Canada.
authors.forename: C M T
authors.initials: CM
authors.lastname: Greenwood
authors.affiliation: Departments of Pediatrics and of Human Genetics of McGill University, Montreal Children's Hospital and Research Institute of the McGill University Health Centre, Glen Site, 1001 Décarie Boulevard Block E- Room EM0.2236 Montreal, QC H4A 3J1, Canada jacquetta.trasler@mcgill.ca.|Department of Pharmacology and Therapeutics, McGill University, Montréal, QC H3A 0G4, Canada.
authors.forename: J M
authors.initials: JM
authors.lastname: Trasler
mesh_terms: D000328:Adult; D019175:DNA Methylation / Q000187:drug effects*; D019587:Dietary Supplements; D004311:Double-Blind Method; D005492:Folic Acid / Q000008:administration & dosage* / Q000032:analysis; D006801:Humans; D008297:Male; D012661:Semen / Q000737:chemistry; D013094:Spermatozoa / Q000187:drug effects / Q000378:metabolism*; D055815:Young Adult
chemical_list: D005492:Folic Acid
keywords: DNA methylation; epigenetics; folic acid supplementation; food fortification; sperm