Stability of the human sperm DNA methylome to folic acid fortification and short-term supplementation.

STUDY QUESTION: Do short-term and long-term exposures to low-dose folic acid supplementation alter DNA methylation in sperm? SUMMARY ANSWER: 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. WHAT IS KNOWN ALREADY: 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. STUDY DESIGN, SIZE, DURATION: 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. PARTICIPANTS/MATERIALS, SETTING, METHODS: 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. MAIN RESULTS AND THE ROLE OF CHANCE: 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. LARGE SCALE DATA: Illumina HumanMethylation450 BeadChip data from this study have been submitted to the NCBI Gene Expression Omnibus under the accession number GSE89781. LIMITATIONS, REASONS FOR CAUTION: 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. WIDER IMPLICATIONS OF THE FINDINGS: 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. STUDY FUNDING/COMPETING INTERESTS: 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.

Low seminal plasma folate concentrations are associated with low sperm density and count in male smokers and nonsmokers.

OBJECTIVE: To measure folate levels in seminal plasma from smokers and nonsmokers and to evaluate relationships between seminal plasma folate levels and both folate nutriture and semen quality measures. DESIGN: Observational study. SETTING: United States Department of Agriculture, Western Human Nutrition Research Center, Presidio of San Francisco, San Francisco, California. PATIENT(S): Healthy male smokers (n=24) and nonsmokers (n=24). MAIN OUTCOME MEASURE(S): Blood levels of plasma folate and homocysteine, seminal plasma total, non-methyl- and 5-methyltetrahydrofolate concentrations, and total sperm count and density. RESULTS: Total seminal plasma folate concentrations were on average 1.5 times higher than blood plasma folate concentrations in all men. Seminal plasma folates contained 5-methyltetrahyrdofolate (74% of total) and non-methyltetrahydrofolates (26% of total); all samples had less than four glutamyl residues. Total and 5-methyltetrahydrofolate concentrations correlated significantly with blood plasma folate and homocysteine concentrations. Seminal plasma non-methyltetrahydrofolate levels correlated significantly with sperm density and total sperm count. Seminal plasma of smokers contained a proportionally lower concentration of non-methyltetrahydrofolates compared with nonsmokers. CONCLUSION(S): Seminal plasma total folate and 5-methyltetrahydrofolate concentrations reflect folate nutriture. The non-methyltetrahydrofolate fraction of seminal plasma may be important for male reproductive function.

Ascorbate is depleted by smoking and repleted by moderate supplementation: a study in male smokers and nonsmokers with matched dietary antioxidant intakes.

BACKGROUND: Lack of reliable dietary data has hampered the ability to effectively distinguish between effects of smoking and diet on plasma antioxidant status. As confirmed by analyses of comprehensive food-frequency questionnaires, the total dietary intakes of fruit and vegetables and of dietary antioxidants were not significantly different between the study groups in the present study, thereby enabling isolation of the effect of smoking. OBJECTIVE: Our objective was to investigate the effect of smoking on plasma antioxidant status by measuring ascorbic acid, alpha-tocopherol, gamma-tocopherol, beta-carotene, and lycopene, and subsequently, to test the effect of a 3-mo dietary supplementation with a moderate-dose vitamin cocktail. DESIGN: In a double-blind, placebo-controlled design, the effect of a vitamin cocktail containing 272 mg vitamin C, 31 mg all-rac-alpha-tocopheryl acetate, and 400 microg folic acid on plasma antioxidants was determined in a population of smokers (n = 37) and nonsmokers (n = 38). The population was selected for a low intake of fruit and vegetables and recruited from the San Francisco Bay area. RESULTS: Only ascorbic acid was significantly depleted by smoking per se (P < 0.01). After the 3-mo supplementation period, ascorbic acid was efficiently repleted in smokers (P < 0.001). Plasma alpha-tocopherol and the ratio of alpha- to gamma-tocopherol increased significantly in both supplemented groups (P < 0.05). CONCLUSIONS: Our data suggest that previous reports of lower concentrations of plasma vitamin E and carotenoids in smokers than in nonsmokers may primarily have been caused by differences in dietary habits between study groups. Plasma ascorbic acid was depleted by smoking and repleted by moderate supplementation.

Meal-induced changes in plasma, erythrocyte, and urinary zinc concentrations in adult women.

We attempted to determine whether there is a limit to the transient, meal-induced decline in plasma zinc and whether there is a concomitant increase in erythrocyte and erythrocyte membrane concentrations. Premenopausal women participated in a 17-h fasting trial, a one-meal trial with breakfast at 0700, and a three-meal trial with meals at 0700, 0900, and 1100. During fasting, plasma zinc increased 9%; it decreased 11% and 19% in the one- and three-meal trials, respectively (P < 0.001). A limit to the decline in plasma zinc was reached after the second meal in the three-meal trial. Erythrocyte, erythrocyte membrane zinc, and serum calcium and phosphorus concentrations did not change significantly during the three trials. Serum glucose concentrations were weakly related to plasma zinc concentrations, suggesting that the postprandial decline in plasma zinc is associated with the metabolic changes caused by food intake.