PMCCPMCCPMCC

Search tips
Search criteria 

Advanced

 
Logo of nihpaAbout Author manuscriptsSubmit a manuscriptNIH Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
Am J Clin Nutr. Author manuscript; available in PMC Feb 1, 2009.
Published in final edited form as:
PMCID: PMC2628952
NIHMSID: NIHMS73940
Importance of methyl donors during reproduction
Steven H. Zeisel, MD, PhD
Steven H. Zeisel, Nutrition Research Institute, Department of Nutrition, School of Public Health and School of Medicine, University of North Carolina at Chapel Hill, CB# 7461, Chapel Hill, NC 27599;
Tel: (919) 843-4731, Fax: (919) 843-8555, E-mail: steven_zeisel/at/unc.edu
There is growing evidence that optimal dietary intake of folate and choline (both involved in 1-carbon transfer or methylation) is important for successful completion of fetal development. Significant portions of the population are eating diets low in one, or both of these nutrients. Folates are important for normal neural tube closure in early gestation and the efficacy of diet fortification with folic acid in reducing incidence of neural tube defects is a major success story for public health nutrition. Similarly, maternal dietary choline is important for normal neural tube closure in the fetus and, later in gestation, also is important for neurogenesis in the fetal hippocampus with effects on memory that persist in adult offspring; higher choline intake being associated with enhanced memory performance. Though both folates and choline have many potentially independent mechanisms whereby they could influence fetal development, these two nutrients also have a common mechanism for action - altered methylation and related epigenetic effects on gene expression.
Keywords: folate, choline, DNA methylation, single nucleotide polymorphism, diet requirement, brain development
Introduction
Dietary intake of folates and choline can be marginal during pregnancy and both nutrients have important effects on brain development. Though these nutrients participate in multiple different biochemical pathways (Figure 1), their metabolism intersects at an important step in 1-carbon metabolism. Perhaps this common pathway explains why both nutrients are required during critical periods of neurogenesis in the brain and spinal cord.
Figure 1
Figure 1
Choline and folate metabolic pathways intersect
Dietary folates, in the form of tetrahydrofolates (THF), are essential cofactors for several biochemical reactions that transfer one carbon units (1). 10-formylTHF (formed from formate and THF by the enzyme C1-THF synthase, the product of the MTHFD1 gene) is required for the biosynthesis of purines (1). 5,10-methyleneTHF, derived from serine and THF, is required for thymidylate biosynthesis. Also, 5,10-methyleneTHF can be reduced to 5-methylTHF (formed by methyleneTHF reductase, the product of the MTHFR gene) and this is needed for the biosynthesis of methionine from homocysteine, eventually influencing biosynthesis of S-adenosylmethionine (the most important methyl-group donor) (1). Thus, variation in dietary folate intake could influence fetal outcome by at least three distinct mechanisms - alteration of DNA biosynthesis, accumulation of toxic levels of homocysteine and perturbation of methylation reactions.
Dietary choline can be acetylated to form acetylcholine, a neurotransmitter (2), or phosphorylated and then used as a precursor for the biosynthesis of phosphatidylcholine and sphingomyelin in mammalian membranes (3-5). Choline is committed to become a methyl donor after it is oxidized to form betaine in the inner mitochondrial membrane, catalyzed by choline dehydrogenase (the product of the CHDH gene) (6). In an alternative pathway to that previously described using 5-methylTHF, the methyl-groups of betaine can be used for the synthesis of methionine from homocysteine, thereby influencing S-adenosylmethionine biosynthesis (7). Thus, variation in dietary choline intake could influence fetal outcome by four distinct mechanisms - perturbation of acetylcholine biosynthesis, changes in membrane synthesis, accumulation of toxic levels of homocysteine and perturbation of methylation reactions.
The dietary requirements for choline and folate are inter-related because the folate and choline metabolic pathways intersect at the point that homocysteine is converted to methionine (8). These two pathways act in parallel, and both lower homocysteine concentrations (9). In the first pathway, vitamins B12 and THF are required cofactors in a reaction catalyzed by methionine synthase (10). Deficiency of these nutrients (11, 12), or single nucleotide polymorphisms in the genes for the enzymes involved in this pathway (10, 12, 13), result in elevated plasma homocysteine concentrations.
The alternative, choline-dependent pathway for the methylation of homocysteine to form methionine is catalyzed by betaine homocysteine methyltransferase (the product of the BHMT gene) (14). Betaine, derived from dietary choline, is the methyl-group donor in this reaction and supplemental oral betaine can lower plasma homocysteine concentrations (15, 16). After betaine donates a methyl-group to homocysteine, the resulting methyl-groups in dimethylglycine can be scavenged using THF as a cofactor (17). Because folate and choline are metabolically related, perturbing metabolism of one results in compensatory changes in metabolism of the other (18-20).
Rats treated with the anti-folate methotrexate have diminished pools of choline metabolites in liver (19, 21). Conversely, rats ingesting a choline-deficient diet have diminished tissue concentrations of folate (20), methionine and S-adenosylmethionine (22) and have elevated plasma homocysteine concentrations (23). Humans who are depleted of choline develop elevated homocysteine concentrations in plasma after a methionine loading test (24). These interactions between choline and folate metabolism are such that it is difficult to separate all of their effects on reproductive outcome.
Dietary intake of folic acid can be marginal during pregnancy, resulting in decreased folate concentrations in serum and red cells to the point that some pregnant women can become clinically folate deficient (25, 26). Normally, the embryonic brain and spinal cord begins as a flat plate that must roll up and then join edges to form a tube. For some reason, this does not happen normally when folates are not available, resulting in the fetus having a neural tube birth defect (NTD). This congenital malformation of the brain and spinal cord results from failure of normal developmental processes in the fetus that must occur during a critical window in time (21-28 days postconception in humans). Mothers with lower erythrocyte folate concentrations are more likely to have a baby with a NTD (27), and folic acid, administered to women who had previously had a child with a neural tube defect, lowers risk of recurrence by 72% (28). There is also an effect of folic acid in women who have never had a baby with an NTD, as rates of this birth defect fell by 26% in the United States after enriched cereal grains sold in the United States were fortified with 140 μg of folic acid per 100 gm of grain (29). A similar fortification program prevented 47% of NTDs in Canada (30). Thus, it is apparent that folate availability is very important during the first few weeks of pregnancy. Little thought has been given to folate nutrition during later pregnancy, but there are significant negative effects of folate deficiency in later gestation on neurogenesis in some areas of the brain related to memory function (31).
These observations have greater significance because genes of folate metabolism are polymorphic, variants are relatively common, and some can increase dietary requirements for folate. Though humans share the same genes, there are many individual variations (single nucleotide polymorphisms; SNPs) in the codon sequences for these genes; In total, over ten million SNPs exist that occur in more than 1% of the population (32). Some common SNPs occur in >50% of the population. Most humans have at least 50,000 SNPs across their genes (33). Some fraction of these SNPs results either in alteration of regulation of gene expression or in changes in the gene product so that protein structure and function are altered, thereby altering metabolism and cell function.
As noted earlier, the product of the MTHFR gene commits folate 1-carbon units to the biosynthesis of methionine from homocysteine, A variant of 5,10-methyleneTHF reductase (MTHFR677 C->T) occurs in as many as 8-15% of the population (34, 35). This SNP results in an alanine-to-valine substitution that produces a thermally unstable enzyme with a 50% reduction of enzymatic activity in homozygous individuals.
There is elevated risk of NTDs for both maternal (50% increase) and fetal (80% increase) TT genotypes (36). Another common SNP of this gene (MTHFR 1298 A->C) also results in reduced enzymatic activity (37). Individuals having both MTHFR polymorphisms have greater risk of NTDs than those that have either polymorphism alone (38). As noted earlier, MTHFD1 encodes for the enzyme C1-THF synthase, catalyzing the synthesis of 10-formylTHF and 5,10-methyleneTHF, the cofactors for de novo purine and thymidylate biosynthesis. A genetic variant of MTHFD1, 653 R->Q is associated with increased maternal risk for NTDs (39). Thus, the interaction between dietary intake of folate and genetic predisposition clearly influence reproductive outcome.
Choline and reproductive outcome
Though choline is found in a variety of foods, including eggs and meats (40) (see www.nal.usda.gov/fnic/foodcomp/Data/Choline/Choline.html), there is significant variation (likely 3-4-fold) in dietary intake of choline among different people. Choline intake on ad libitum diets for males and females averages 8.4 mg/kg and. 6.7 mg/kg of choline per day, respectively (41) However, in several studies in the USA, investigators observed intakes that were less than half this amount in 25% of the women studied (42-44). Choline is derived not only from the diet, but as well from de novo synthesis of phosphatidylcholine catalyzed by phosphatidylethanolamine N-methyltransferase (the product of the PEMT gene) in the liver (45).
When deprived of dietary choline, most men and postmenopausal women develop fatty liver or muscle damage (24, 46). However, only a portion (44%) of premenopausal women develop such problems when choline deficient. The difference in requirement occurs because estrogen induces the PEMT gene and allows premenopausal women to make more of their needed choline endogenously (47). During pregnancy, estrogen concentration rises from approximately 1nM to 60nM at term (48, 49), suggesting that capacity for endogenous synthesis of choline should be highest during period when females need to support fetal development. This is fortunate, as pregnancy and lactation are times when demand for choline is especially high because of transport of choline from mother to fetus (50, 51) depletes maternal plasma choline in humans (52). Thus, despite enhanced capacity to synthesize choline, the demand for this nutrient is so high that stores are depleted. Pregnant rats had diminished total liver choline stores compared to non-mated controls and become as sensitive to choline-deficient diets as were male rats (53). Because milk contains a great deal of choline, lactation further increases maternal demand for choline resulting in further depletion of tissue stores (53, 54). These observations suggest that women depend on high rates of endogenous biosynthesis of choline induced by estrogen (47) and dietary intake of choline to sustain normal pregnancy.
Feeding rodents more choline during a few days in pregnancy increases the rate of brain neurogenesis in the fetus; it also decreases apoptosis (cell suicide) rates in these cells (55, 56). Low maternal choline intake during days 11-17 of gestation resulted in half as much neural progenitor cell proliferation and twice as much progenitor cell apoptosis in the fetal hippocampus (memory center) compared to fetuses from mothers fed choline adequate diets (56, 57). The offspring of choline deficient dams had diminished visuospatial and auditory memory for the rest of their lives (58). Conversely, more choline (about 4 times normal dietary levels) fed to pregnant dams enhanced visuospatial and auditory memory in their offspring by as much as 30% throughout life (58-64). Indeed, adult rodents normally lose memory function as they age, and offspring exposed to extra choline in utero did not show this “senility” (61, 63).
It seems that the progenitor cells of the neural tube are affected by choline in the same way as are the hippocampal progenitor cells are. In mice, choline is needed for normal neural tube closure in the fetus (65, 66) and in humans, women who eat a relatively low folate diet and are in the lowest quartile for dietary choline intake had four times the risk (compared with women in the highest quartile) of having a baby with a neural tube defect (42). This observation supports the suggestion that the basic research in rodents will be applicable to the human condition. Of course human and rat brains mature at different rates, with rat brain comparatively more mature at birth than is the human brain. In humans, the architecture of the hippocampus continues to develop after birth, and by 4 years of age it closely resembles adult structure (67). This area of brain is one of the few areas in which neurons continue to multiply slowly throughout life (68, 69).
The effects of dietary choline on fetal development have greater significance because genes of choline metabolism are polymorphic, variants are relatively common, and some can increase dietary requirements for choline. Among these functionally important SNPs, a number have been identified that explain differences in risk for developing organ dysfunction or damage when humans are fed diets low in choline (24, 46, 70). As discussed earlier, the gene PEMT encodes for a protein responsible for endogenous formation of choline in the liver (71) and it is induced by estrogen (47). In studies of organ dysfunction after choline deficiency in humans, a SNP in the promoter region of the PEMT gene (rs12325817) was associated with greatly increased susceptibility to choline deficiency in women but not in men (72). This SNP was very common, with 14% of a Chapel Hill NC population being homozygous for it, and three quarters of the population having one allele (72). Two SNPs in the coding region of the choline dehydrogenase gene (CHDH) are common. One (rs9001, 13% of of a Chapel Hill NC population has 1 allele) had a protective effect on susceptibility to choline deficiency, while a second variant (rs12676, 51% of a Chapel Hill NC population has 1 allele) was associated with increased susceptibility to choline deficiency (72).
Because choline and folate metabolism are intermingled, SNPs in one pathway can change the dietary requirement for the other nutrient. The methylene tetrahydrofolate dehydrogenase (MTHFD1) G1958A polymorphism affects the balance of flux between 5,10-methylene tetrahydrofolate and 10-formyl tetrahydrofolate and thereby reduces the availability of 5-methyl tetrahydrofolate for homocysteine remethylation (73). Premenopausal women who were carriers of this very common SNP (63% of the population has 1 allele) were more than 15 times as likely as non-carriers to develop signs of choline deficiency on a low-choline diet (73). It is of interest that the risk of having a child with a neural tube defect increases in mothers with this SNP (39).
DNA can be methylated at cytosine bases that are followed by a guanosine (CpG islands) (74), and S-adenosylmethionine, derived from methionine, choline and/or 5-methylTHF, is the source of the methyl-groups. Low dietary choline-folate intake not only depletes choline and folate metabolites, but also decreases S-adenosylmethionine concentrations (22, 75), with resulting hypomethylation of DNA (76, 77). DNA methylation influences gene transcription and genomic stability (78-80); increased methylation is usually associated with gene silencing or reduced gene expression (81) because methylated CpG islands attract capping proteins that hinder access to the gene for the transcription factors that normally induce gene expression (82). Once CpG islands in genes are methylated, the methylation is reproduced every time the gene is copied. Thus, effects of methylation persist, perhaps throughout life.
Changes in dietary availability of methyl-groups induces stable changes in gene methylation, altering gene expression and resulting phenotype (83, 84). For example, feeding pregnant Pseudoagouti Avy/a mouse dams a methyl-supplemented diet altered agouti gene expression in their offspring, as indicated by increased agouti/black mottling of their coats (83, 85). In a similar study, maternal dietary intake of methyl-groups influenced methylation of the gene Axin Fused which determined whether offspring had permanently kinked tails (86). Many of the changes in neurogenesis caused by altered availability of dietary choline or folate during pregnancy are likely to be mediated by altered DNA methylation. Decreased choline in diets of pregnant mice was associated with changes in DNA methylation in fetal brain that were specific to some CpG islands, and even to specific CpG sites, within genes that regulate cell cycling (87, 88); methylation of the CDKN3 gene promoter was decreased in fetal brain, resulting in over expression of this gene which inhibits cell proliferation (87). It is clear that the dietary manipulation of methyl donors (either deficiency or supplementation) can have a profound impact upon reproductive outcome through epigenetic mechanisms. For this reason, it is important that expert panels carefully consider recommendations for dietary intake of methyl donors during pregnancy.
Acknowledgments
This work was funded by a grant from the National Institutes of Health (DK55865, AG09525). Support for this work also was provided by grants from the NIH to the UNC Clinical Nutrition Research Unit (DK56350), the UNC General Clinical Research Center (RR00046) and the Center for Environmental Health and Susceptibility (ES10126).
Conflicts of Interest: The author has funding from Balchem, Mead Johnson Nutritionals, and from the Egg Nutrition Center. He serves on health advisory boards for Dupont, Solae and Metabolon. None of these funds or activities influenced the content of this manuscript.
1. Beaudin AE, Stover PJ. Folate-mediated one-carbon metabolism and neural tube defects: balancing genome synthesis and gene expression. Birth Defects Res C Embryo Today. 2007 Sep;81:183–203. [PubMed]
2. Blusztajn JK, Wurtman RJ. Choline and cholinergic neurons. Science. 1983;221:614–20. [PubMed]
3. Vance DE. Boehringer Mannheim Award lecture. Phosphatidylcholine metabolism: masochistic enzymology, metabolic regulation, and lipoprotein assembly. Biochem Cell Biol. 1990;68:1151–65. [PubMed]
4. Kent C. Regulation of phosphatidylcholine biosynthesis. Prog Lipid Res. 1990;29:87–105. [PubMed]
5. Hanada K, Horii M, Akamatsu Y. Functional reconstitution of sphingomyelin synthase in chinese hamster ovary cell membranes. Biochim Biophy Acta. 1991;1086:151–6. [PubMed]
6. Lin CS, Wu RD. Choline oxidation and choline dehydrogenase. J Prot Chem. 1986;5:193–200.
7. Niculescu MD, Zeisel SH. Diet, methyl donors and DNA methylation: interactions between dietary folate, methionine and choline. J Nutr. 2002 Aug;132:2333S–5S. [PubMed]
8. Finkelstein JD. Pathways and regulation of homocysteine metabolism in mammals. Semin Thromb Hemost. 2000;26:219–25. [PubMed]
9. Olthof MR, van Vliet T, Boelsma E, Verhoef P. Low dose betaine supplementation leads to immediate and long term lowering of plasma homocysteine in healthy men and women. J Nutr. 2003 Dec;133:4135–8. [PubMed]
10. Weisberg IS, Jacques PF, Selhub J, Bostom AG, Chen Z, Curtis Ellison R, Eckfeldt JH, Rozen R. The 1298A-->C polymorphism in methylenetetrahydrofolate reductase (MTHFR): in vitro expression and association with homocysteine. Atherosclerosis. 2001 Jun;156:409–15. [PubMed]
11. Shelnutt KP, Kauwell GP, Chapman CM, Gregory JF, 3rd, Maneval DR, Browdy AA, Theriaque DW, Bailey LB. Folate status response to controlled folate intake is affected by the methylenetetrahydrofolate reductase 677C-->T polymorphism in young women. J Nutr. 2003 Dec;133:4107–11. [PubMed]
12. Jacques PF, Bostom AG, Wilson PW, Rich S, Rosenberg IH, Selhub J. Determinants of plasma total homocysteine concentration in the Framingham Offspring cohort. Am J Clin Nutr. 2001 Mar;73:613–21. [PubMed]
13. Watkins D, Ru M, Hwang HY, Kim CD, Murray A, Philip NS, Kim W, Legakis H, Wai T, et al. Hyperhomocysteinemia due to methionine synthase deficiency, cblG: structure of the MTR gene, genotype diversity, and recognition of a common mutation, P1173L. Am J Hum Genet. 2002 Jul;71:143–53. [PubMed]
14. Sunden S, Renduchintala M, Park E, Miklasz S, Garrow T. Betaine-Homocysteine methyltransferase expression in porcine and human tissues and chromosomal localization of the human gene. Arch Biochem Biophys. 1997;345:171–4. [PubMed]
15. Steenge GR, Verhoef P, Katan MB. Betaine supplementation lowers plasma homocysteine in healthy men and women. J Nutr. 2003 May;133:1291–5. [PubMed]
16. Wendel U, Bremer H. Betaine in the treatment of homocystinuria due to 5,10-methylenetetrahydrofolate reductase deficiency. Eur J Pediatr. 1984;142:147–50. [PubMed]
17. Mudd SH, Ebert MH, Scriver CR. Labile methyl group balances in the human: the role of sarcosine. Metabolism. 1980;29:707–20. [PubMed]
18. Kim Y-I, Miller JW, da Costa K-A, Nadeau M, Smith D, Selhub J, Zeisel SH, Mason JB. Folate deficiency causes secondary depletion of choline and phosphocholine in liver. J Nutr. 1995;124:2197–203. [PubMed]
19. Selhub J, Seyoum E, Pomfret EA, Zeisel SH. Effects of choline deficiency and methotrexate treatment upon liver folate content and distribution. Cancer Res. 1991;51:16–21. [PubMed]
20. Varela Moreiras G, Selhub J, da Costa K, Zeisel SH. Effect of chronic choline deficiency in rats on liver folate content and distribution. J Nutr Biochem. 1992;3:519–22.
21. Pomfret EA, da Costa K, Zeisel SH. Effects of choline deficiency and methotrexate treatment upon rat liver. J Nutr Biochem. 1990;1:533–41. [PubMed]
22. Zeisel SH, Zola T, daCosta K, Pomfret EA. Effect of choline deficiency on S-adenosylmethionine and methionine concentrations in rat liver. Biochem J. 1989;259:725–9. [PubMed]
23. Varela-Moreiras G, Ragel C, Perez de Miguelsanz J. Choline deficiency and methotrexate treatment induces marked but reversible changes in hepatic folate concentrations, serum homocysteine and DNA methylation rates in rats. J Amer Coll Nutr. 1995;14:480–5. [PubMed]
24. da Costa KA, Gaffney CE, Fischer LM, Zeisel SH. Choline deficiency in mice and humans is associated with increased plasma homocysteine concentration after a methionine load. Am J Clin Nutr. 2005 Feb;81:440–4. [PMC free article] [PubMed]
25. Willoughby M, Jewell F. Folate status throughout pregnancy and in postpartum period. Brit Med J. 1968;4:356–60. [PMC free article] [PubMed]
26. Qvist I, Abdulla M, Jagerstad M, Svensson S. Iron, zinc and folate status during pregnancy and two months after delivery. Acta Obstet Gynecol Scand. 1986;65:15–22. [PubMed]
27. Smithells RW, Sheppard S, Schorah CJ. Vitamin dificiencies and neural tube defects. Arch Dis Child. 1976 Dec;51:944–50. [PMC free article] [PubMed]
28. MRC Vitamin Study Research Group Prevention of neural tube defects: results of the medical Research Council Vitamin Study. Lancet. 1991;338:131–7. [PubMed]
29. Spina bifida and anencephaly before and after folic acid mandate--United States, 1995-1996 and 1999-2000. MMWR Morb Mortal Wkly Rep. 2004 May 7;53:362–5. [PubMed]
30. Persad VL, Van den Hof MC, Dube JM, Zimmer P. Incidence of open neural tube defects in Nova Scotia after folic acid fortification. Can Med Assoc J. 2002 Aug 6;167:241–5. [PMC free article] [PubMed]
31. Craciunescu CN, Brown EC, Mar MH, Albright CD, Nadeau MR, Zeisel SH. Folic acid deficiency during late gestation decreases progenitor cell proliferation and increases apoptosis in fetal mouse brain. J Nutr. 2004 Jan;134:162–6. [PubMed]
32. McVean G, Spencer CC, Chaix R. Perspectives on human genetic variation from the HapMap Project. PLoS Genet. 2005 Oct;1:e54. [PMC free article] [PubMed]
33. Hinds DA, Stuve LL, Nilsen GB, Halperin E, Eskin E, Ballinger DG, Frazer KA, Cox DR. Whole-genome patterns of common DNA variation in three human populations. Science. 2005 Feb 18;307:1072–9. [PubMed]
34. Motulsky A. Nutritional ecogenetics: homocysteine-related arteriosclerotic vascular disease, neural tube defects, and folic acid. Am J Hum Genet. 1996;58:17–20. [PubMed]
35. van der Put N, Steegers-Theunissen R, Frosst P, et al. Mutated methylene tetrahydrofolate reductase as a risk factor for spinal bifida. Lancet. 1995;346:1070–1. [PubMed]
36. Blom HJ, Shaw GM, den Heijer M, Finnell RH. Neural tube defects and folate: case far from closed. Nature reviews. 2006 Sep;7:724–31. [PMC free article] [PubMed]
37. Weisberg I, Tran P, Christensen B, Sibani S, Rozen R. A second genetic polymorphism in methylenetetrahydrofolate reductase (MTHFR) associated with decreased enzyme activity. Molecular genetics and metabolism. 1998 Jul;64:169–72. [PubMed]
38. Relton CL, Wilding CS, Laffling AJ, Jonas PA, Burgess T, Binks K, Tawn EJ, Burn J. Low erythrocyte folate status and polymorphic variation in folate-related genes are associated with risk of neural tube defect pregnancy. Molecular genetics and metabolism. 2004 Apr;81:273–81. [PubMed]
39. Brody LC, Conley M, Cox C, Kirke PN, McKeever MP, Mills JL, Molloy AM, O’Leary VB, Parle-McDermott A, et al. A polymorphism, R653Q, in the trifunctional enzyme methylenetetrahydrofolate dehydrogenase/methenyltetrahydrofolate cyclohydrolase/formyltetrahydrofolate synthetase is a maternal genetic risk factor for neural tube defects: report of the Birth Defects Research Group. Am J Hum Genet. 2002 Nov;71:1207–15. [PubMed]
40. Zeisel SH, Mar MH, Howe JC, Holden JM. Concentrations of choline-containing compounds and betaine in common foods. J Nutr. 2003 May;133:1302–7. [PubMed]
41. Fischer LM, Scearce JA, Mar MH, Patel JR, Blanchard RT, Macintosh BA, Busby MG, Zeisel SH. Ad libitum choline intake in healthy individuals meets or exceeds the proposed adequate intake level. J Nutr. 2005 Apr;135:826–9. [PMC free article] [PubMed]
42. Shaw GM, Carmichael SL, Yang W, Selvin S, Schaffer DM. Periconceptional dietary intake of choline and betaine and neural tube defects in offspring. Am J Epidemiol. 2004 Jul 15;160:102–9. [PubMed]
43. Cho E, Willett WC, Colditz GA, Fuchs CS, Wu K, Chan AT, Zeisel SH, Giovannucci EL. Dietary choline and betaine and the risk of distal colorectal adenoma in women. J Natl Cancer Inst. 2007 Aug 15;99:1224–31. [PMC free article] [PubMed]
44. Bidulescu A, Chambless LE, Siega-Riz AM, Zeisel SH, Heiss G. Usual choline and betaine dietary intake and incident coronary heart disease: the Atherosclerosis Risk in Communities (ARIC) study. BMC Cardiovasc Disord. 2007;7:20. [PMC free article] [PubMed]
45. Zhu X, Mar MH, Song J, Zeisel SH. Deletion of the Pemt gene increases progenitor cell mitosis, DNA and protein methylation and decreases calretinin expression in embryonic day 17 mouse hippocampus. Brain Res Dev Brain Res. 2004 Apr 19;149:121–9. [PubMed]
46. da Costa KA, Badea M, Fischer LM, Zeisel SH. Elevated serum creatine phosphokinase in choline-deficient humans: mechanistic studies in C2C12 mouse myoblasts. Am J Clin Nutr. 2004 Jul;80:163–70. [PubMed]
47. Resseguie M, Song J, Niculescu MD, da Costa KA, Randall TA, Zeisel SH. Phosphatidylethanolamine N-methyltransferase (PEMT) gene expression is induced by estrogen in human and mouse primary hepatocytes. Faseb J. 2007 Aug;21:2622–32. [PMC free article] [PubMed]
48. Sarda IR, Gorwill RH. Hormonal studies in pregnancy. I. Total unconjugated estrogens in maternal peripheral vein, cord vein, and cord artery serum at delivery. Am J Obstet Gynecol. 1976 Feb 1;124:234–8. [PubMed]
49. Adeyemo O, Jeyakumar H. Plasma progesterone, estradiol-17 beta and testosterone in maternal and cord blood, and maternal human chorionic gonadotropin at parturition. Afr J Med Med Sci. 1993 Sep;22:55–60. [PubMed]
50. Sweiry JH, Yudilevich DL. Characterization of choline transport at maternal and fetal interfaces of the perfused guinea-pig placenta. J Physiol. 1985;366:251–66. [PubMed]
51. Sweiry JH, Page KR, Dacke CG, Abramovich DR, Yudilevich DL. Evidence of saturable uptake mechanisms at maternal and fetal sides of the perfused human placenta by rapid paired-tracer dilution: studies with calcium and choline. J Devel Physiol. 1986;8:435–45. [PubMed]
52. McMahon KE, Farrell PM. Measurement of free choline concentrations in maternal and neonatal blood by micropyrolysis gas chromatography. Clin Chim Acta. 1985;149:1–12. [PubMed]
53. Zeisel SH, Mar M-H, Zhou Z-W, da Costa K-A. Pregnancy and lactation are associated with diminished concentrations of choline and its metabolites in rat liver. J Nutr. 1995;125:3049–54. [PubMed]
54. Holmes-McNary M, Cheng WL, Mar MH, Fussell S, Zeisel SH. Choline and choline esters in human and rat milk and infant formulas. Am J Clin Nutr. 1996;64:572–6. [PubMed]
55. Craciunescu CN, Albright CD, Mar MH, Song J, Zeisel SH. Choline availability during embryonic development alters progenitor cell mitosis in developing mouse hippocampus. J Nutr. 2003 Nov;133:3614–8. [PMC free article] [PubMed]
56. Albright CD, Tsai AY, Friedrich CB, Mar MH, Zeisel SH. Choline availability alters embryonic development of the hippocampus and septum in the rat. Brain Res Dev Brain Res. 1999;113:13–20. [PubMed]
57. Albright CD, Friedrich CB, Brown EC, Mar MH, Zeisel SH. Maternal dietary choline availability alters mitosis, apoptosis and the localization of TOAD-64 protein in the developing fetal rat septum. Brain Res Dev Brain Res. 1999;115:123–9. [PubMed]
58. Meck WH, Williams CL. Choline supplementation during prenatal development reduces proactive interference in spatial memory. Brain Res Dev Brain Res. 1999;118:51–9. [PubMed]
59. Meck W, Williams C. Perinatal choline supplementation increases the threshold for chunking in spatial memory. Neuroreport. 1997;8:3053–9. [PubMed]
60. Meck W, Williams C. Characterization of the facilitative effects of perinatal choline supplementation on timing and temporal memory. Neuroreport. 1997;8:2831–5. [PubMed]
61. Meck W, Williams C. Simultaneous temporal processing is sensitive to prenatal choline availability in mature and aged rats. Neuroreport. 1997;8:3045–51. [PubMed]
62. Meck WH, Smith RA, Williams CL. Pre- and postnatal choline supplementation produces long-term facilitation of spatial memory. Dev Psychobiol. 1988;21:339–53. [PubMed]
63. Meck WH, Williams CL. Metabolic imprinting of choline by its availability during gestation: Implications for memory and attentional processing across the lifespan. Neurosci Biobehav Rev. 2003;27:385–99. [PubMed]
64. Williams CL, Meck WH, Heyer DD, Loy R. Hypertrophy of basal forebrain neurons and enhanced visuospatial memory in perinatally choline-supplemented rats. Brain Res. 1998;794:225–38. [PubMed]
65. Fisher MC, Zeisel SH, Mar MH, Sadler TW. Inhibitors of choline uptake and metabolism cause developmental abnormalities in neurulating mouse embryos. Teratology. 2001;64:114–22. [PubMed]
66. Fisher MC, Zeisel SH, Mar MH, Sadler TW. Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro. Faseb J. 2002 Apr;16:619–21. [PubMed]
67. Dani S, Hori A, Walter G, editors. Principals of neural aging. Elsevier; Amsterdam: 1997.
68. van Praag H, Kempermann G, Gage FH. Running increases cell proliferation and neurogenesis in the adult mouse dentate gyrus. Nat Neurosci. 1999;2:266–70. [PubMed]
69. Markakis EA, Gage FH. Adult-generated neurons in the dentate gyrus send axonal projections to field CA3 and are surrounded by synaptic vesicles. J Comp Neurol. 1999;406:449–60. [PubMed]
70. da Costa KA, Kozyreva OG, Song J, Galanko JA, Fischer LM, Zeisel SH. Common genetic polymorphisms affect the human requirement for the nutrient choline. Faseb J. 2006 Jul;20:1336–44. [PMC free article] [PubMed]
71. Vance DE, Walkey CJ, Cui Z. Phosphatidylethanolamine N-methyltransferase from liver. Biochim Biophys Acta. 1997;1348:142–50. [PubMed]
72. da Costa K, Kozyreva OG, Song J, Galanko JA, Fischer LM, Zeisel SH. Common genetic polymorphisms have major effects on the human requirement for the nutrient choline. Faseb J. 2006;20:1336–44. [PMC free article] [PubMed]
73. Kohlmeier M, da Costa KA, Fischer LM, Zeisel SH. Genetic variation of folate-mediated one-carbon transfer pathway predicts susceptibility to choline deficiency in humans. Proc Natl Acad Sci U S A. 2005 Nov 1;102:16025–30. [PubMed]
74. Holliday R, Grigg GW. DNA methylation and mutation. Mutat Res. 1993 Jan;285:61–7. [PubMed]
75. Shivapurkar N, Poirier LA. Tissue levels of S-adenosylmethionine and S-adenosylhomocysteine in rats fed methyl-deficient, amino acid-defined diets for one to five weeks. Carcinogenesis. 1983;4:1051–7. [PubMed]
76. Locker J, Reddy TV, Lombardi B. DNA methylation and hepatocarcinogenesis in rats fed a choline devoid diet. Carcinogenesis. 1986;7:1309–12. [PubMed]
77. Tsujiuchi T, Tsutsumi M, Sasaki Y, Takahama M, Konishi Y. Hypomethylation of CpG sites and c-myc gene overexpression in hepatocellular carcinomas, but not hyperplastic nodules, induced by a choline-deficient L-amino acid-defined diet in rats. Jpn J Cancer Res. 1999;90:909–13. [PubMed]
78. Jaenisch R. DNA methylation and imprinting: why bother? Trends Genet. 1997 Aug;13:323–9. [PubMed]
79. Jones PA, Gonzalgo ML. Altered DNA methylation and genome instability: a new pathway to cancer? Proc Natl Acad Sci U S A. 1997 Mar 18;94:2103–5. [PubMed]
80. Robertson KD, Wolffe AP. DNA methylation in health and disease. Nat Rev Genet. 2000 Oct;1:11–9. [PubMed]
81. Jeltsch A. Beyond Watson and Crick: DNA Methylation and Molecular Enzymology of DNA Methyltransferases. Chembiochem. 2002 May 3;3:382. [PubMed]
82. Fan G, Hutnick L. Methyl-CpG binding proteins in the nervous system. Cell Res. 2005 Apr;15:255–61. [PubMed]
83. Cooney CA, Dave AA, Wolff GL. Maternal methyl supplements in mice affect epigenetic variation and DNA methylation of offspring. J Nutr. 2002 Aug;132:2393S–400S. [PubMed]
84. Waterland RA, Jirtle RL. Transposable elements: targets for early nutritional effects on epigenetic gene regulation. Mol Cell Biol. 2003 Aug;23:5293–300. [PMC free article] [PubMed]
85. Wolff GL, Kodell RL, Moore SR, Cooney CA. Maternal epigenetics and methyl supplements affect agouti gene expression in Avy/a mice. Faseb J. 1998 Aug;12:949–57. [PubMed]
86. Waterland RA, Dolinoy DC, Lin JR, Smith CA, Shi X, Tahiliani KG. Maternal methyl supplements increase offspring DNA methylation at Axin fused. Genesis. 2006 Jul 25;44:401–6. [PubMed]
87. Niculescu MD, Craciunescu CN, Zeisel SH. Dietary choline deficiency alters global and gene-specific DNA methylation in the developing hippocampus of mouse fetal brains. Faseb J. 2006 Jan;20:43–9. [PMC free article] [PubMed]
88. Niculescu MD, Yamamuro Y, Zeisel SH. Choline availability modulates human neuroblastoma cell proliferation and alters the methylation of the promoter region of the cyclin-dependent kinase inhibitor 3 gene. J Neurochem. 2004 Jun;89:1252–9. [PMC free article] [PubMed]