HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Fillon-Emery, N. Articles by Nicolas, J. P. Search for Related Content PubMed PubMed Citation Articles by Fillon-Emery, N. Articles by Nicolas, J. P. Agricola Articles by Fillon-Emery, N. Articles by Nicolas, J. P.

Homocysteine concentrations in adults with trisomy 21: effect of B vitamins and genetic polymorphisms^1,2,3,4

¹ From the Faculté de Médecine, Laboratory of Medical Biochemistry, Vandoeuvre-Lès-Nancy, France (NF-E, FB, and JPN); the ISAB, Laboratory of Nutritional Genomics, Beauvais, France (AC); the Jérôme Lejeune Medical Center, Paris (CM, HB, and M-OR); INSERM U525 and CMP, Nancy, France (BH and DL); and the Division of Medical Genetics, Department of Medicine, McGill University, Montréal, Quebec (DSR)

² Supported by a grant from the Fondation Jerôme Lejeune and the Fondation pour la Recherche Médicale.

³ Address reprint requests to JP Nicolas, Laboratory of Medical Biochemistry, Faculty of Medicine, BP 184, 54505 Vandoeuvre cedex, France. E-mail: jean-pierre.nicolas20{at}wanadoo.fr

⁴ Address correspondence to Abalo Chango, ISAB –Agrohealth, Laboratory of Nutritional Genomics, BP 30313, Rue Pierre Waguet, 60026-Beauvais, France. E-mail: abalo.chango{at}isab.fr.

See the corresponding letter to the Editor on page XXX.

	ABSTRACT

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Background:The effects of supplementation with B vitamins and of common polymorphisms in genes involved in homocysteine metabolism on plasma total homocysteine (tHcy) concentrations in trisomy 21 are unknown.

Objectives:We aimed to determine the effects of orally administered folic acid and of folic acid combined with vitamin B-12, vitamin B-6, or both on tHcy in adults with trisomy 21. The study was also intended to analyze the possible influence of gene polymorphisms.

Design:One hundred sixty adults with trisomy 21 and 160 healthy, unrelated subjects aged 26 ± 4 y were included. Plasma tHcy, red blood cell folate, serum folate, and vitamin B-12 were measured. Genotyping for the common methylenetetrahydrofolate reductase (MTHFR) 677CT, MTHFR 1298AC, cystathionine ß-synthase 844Ins68, methionine synthase 2756AC, methionine synthase reductase 66AG, and reduced folate carrier 80GA polymorphisms was carried out.

Results:The mean tHcy concentration (9.8 ± 0.7 µmol/L) of cases who did not use vitamins was not significantly different from that of controls (9.4 ± 0.3 µmol/L). Plasma tHcy concentrations (7.6 ± 0.3 mmol/L) in cases who used folic acid were significantly lower than in cases who did not. Folic acid combined with vitamin B-12 did not significantly change tHcy concentrations compared with those in cases who used only folic acid. Folic acid combined with vitamins B-6 and B-12 significantly lowered tHcy (6.5 ± 0.5 µmol/L). The difference in tHcy according to MTHFR genotype was not significant. However, tHcy concentrations were slightly higher in TT homozygotes among the controls but not among the cases.

Conclusion:This study provides information on the relation between several polymorphisms in genes involved in homocysteine and folate metabolism in adults with trisomy 21.

Key Words: Down syndrome • trisomy 21 • folate • homocysteine • polymorphism • CBS • MTR • MTRR • MTHFR • RFC • vitamin B-12 • vitamin B-6

	INTRODUCTION

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Down syndrome (MIM 190685), or trisomy 21, is a complex metabolic and genetic disorder that stems from the failure of chromosome 21 to segregate normally during meiosis (1). More than 80 clinical features have been observed in individuals with trisomy 21, including cognitive impairment, congenital heart disease, and childhood leukemia (2–4). These features are complex and involve various combinations of genes and environmental factors. These include allelic heterogeneity for chromosome 21 genes, epistatic interactions (chromosome 21 genes with genes on 21 or on other chromosomes), and environmental events (5). Many of these clinical features refer to observations in children whose folate requirement may be higher than that of the adults in the present study.

The cystathionine ß-synthase gene (CBS) on chromosome 21q22.3 codes for a pyridoxal 5-phosphate–dependent enzyme that condenses homocysteine and serine to form cystathionine (6, 7). CBS is over-expressed in individuals with trisomy 21 (8). Another gene on chromosome 21q22.3 (RFC1 or SLC19A1) codes for the reduced folate carrier, which is responsible for 5-methyltetrahydrofolate internalization within cells. Homocysteine is situated at a critical regulatory branch point in the sulfur metabolism produced by cellular demethylation of dietary methionine. It can be remethylated to methionine by the transfer of the methyl group of methyltetrahydrofolate (9), which requires methionine synthase (5-methyltetrahydrofolate–homocysteine S-methyltransferase) with both vitamin B-12 and S-adenosylmethionine as cofactors, or can be converted to cysteine in the transsulfuration pathway, which requires CBS and vitamin B-6 as cofactors. The release of homocysteine into the extracellular medium represents the third route of cellular homocysteine disposal.

Over the past decade, several contributions have shown that genetic polymorphisms might influence plasma total homocysteine (tHcy) concentrations, either directly or by affecting plasma folate concentrations. Polymorphisms of the 5,10-methylenetetrahydrofolate reductase gene (MTHFR) have been studied extensively in various populations. Suboptimal doses of folate have been shown to increase plasma homocysteine concentrations in the presence of the 677CT polymorphism (10). Many other polymorphisms in genes encoding proteins involved in homocysteine and folate have been reported. A polymorphism in the gene encoding methionine synthase (MTR) results in an adenine-to-guanine substitution at nucleotide 2756 in cDNA (11), and a polymorphism in the gene encoding methionine synthase reductase (MTRR) results in an adenine-to-guanine substitution at nucleotide 66. Both polymorphisms may be associated with higher tHcy concentrations (12, 13). For the CBS gene, the most common variation is a 68–base pair (bp) insertion (844Ins68) polymorphism in the coding region of exon 8 (14). More recently, we published a polymorphism in the human RFC1 cDNA and hypothesized that it influences plasma tHcy concentrations (15). This polymorphism is a guanine-to-adenine exchange at nucleotide 80. The RFC1 gene is involved in cellular folate uptake. The effect of the 80GA polymorphism has only been studied in a limited way (16-18).

The aim of the present study was to evaluate the effect of vitamin supplementation on tHcy concentrations in patients with trisomy 21. We attempted to determine the consequences of orally administered folic acid or folic acid combined with vitamin B-12, vitamin B-6, or both. An additional goal of the study was to analyze the possible influence of the MTHFR 677CT, MTHFR 1298AC, CBS 844Ins68, MTR 2756AG, MTRR 66AG, and RFC 80GA allele variants on homocysteine concentrations.

	SUBJECTS AND METHODS

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Subjects
The participants were 160 French individuals (87 men and 73 women) aged 26 ± 4 y ( ± SD; range: 15–46 y) with full trisomy 21 confirmed by karyotype, and 160 healthy, unrelated control subjects matched for sex and age. Controls were enrolled provided that they had no history of cardiovascular disease, cancer, epilepsy, diabetes, or impaired renal function. None of the healthy subjects enrolled used alcohol, anticonvulsants, or antifolate and none took vitamin or mineral supplements. A questionnaire was completed by each participant to indicate whether he or she had taken a vitamin or mineral supplement before the blood sample being given. For persons with trisomy 21, the study included both participants who were taking and those who were not taking vitamin B supplements (folic acid, vitamin B-6, or vitamin B-12). Of the 160 individuals, 25 used no form of vitamin supplements. Other participants used drug-based vitamin B, such as Speciafoldine (contains 5 mg folic acid per pill; Aventis, Paris), MAGNEB₆ (5 mg vitamin B-6 chlorhydrate per vial; Sanofi-Synthelabo, Paris), and vitamin B-12 Aguettant (100 µg cyanocobalamin; Aguettant, Lyon, France). Sixty-five persons used folic acid (Speciafoldine) only, 23 persons used folic acid combined with vitamin B-12 (vitamin B-12 Aguettant), 25 persons used folic acid combined with vitamin B-6 (MAGNEB₆), and 22 persons used all 3 vitamins. The study protocol was approved by the Ethical Committee for Studies on Humans of the CHU de Nancy, and written informed consent was obtained from the participants' parents or the participants themselves.

Blood sampling and biochemical determination
Fasting blood samples were collected from the participants by venipuncture. After collection, whole blood was kept at 4 °C until centrifuged at 400 x g for 15 min. The time elapsing between blood collection and plasma separation was <1 h. Samples were stored at –80 °C until analyzed. The tHcy concentration (the total amount of protein- and non-protein-bound homocysteine) was measured by HPLC with fluorescence detection (19), involving derivatization with 7-fluorobenzo-2-axo-1,3-diazole-4-sulfonate (SBD-F) according to Ubbink et al (20, 21). Briefly, after reduction with tri-n-butylphosphine, deproteinization with trichloroacetic acid, centrifugation, and derivatization with SBD-F, the compounds are isocratically separated by C₁₈ reversed-phase HPLC and quantified by fluorescence detection. The mobile phase was 0.1 mol/L phosphate buffer:acetonitrile (pH 1.5; 94:6 by vol). The excitation and emission wavelengths were 384 and 516 nm, respectively. The retention time was 5 min for Hcy and 11 min for the internal standard (N-acetylcysteine). Red blood cell (RBC) folate, serum folate, and serum vitamin B-12 were measured by immunoassays (Abbott, Rungis, France). The Simul TRAC-SNB Kit (ICN Pharmaceuticals, Orangeburg, NY) uses purified intrinsic factor. In the procedure, endogenous serum binders for both folate and vitamin B-12 are destroyed after incubation with a dithiothreitol tracer for 15 min followed by a 10-min extraction reaction at an alkaline pH. This eliminates the need to heat the sample at 100 °C. Vitamin B-12 and folate concentrations are measured simultaneously in a single tube. The vitamin B-12 and folate tracers, binders, and standards are supplied in combined form.

Genomic DNA isolation and analysis by polymerase chain reaction
Human genomic DNA was isolated from peripheral blood samples with NucleoSpin Blood Quick Pure (Machery-Nagel, Cergy Pontoise, France). The MTHFR 677CT variant was identified by using polymerase chain reaction (PCR) followed by restriction enzyme digestion of the amplified product (22–24). Briefly, PCR primers for amplification of the MTHFR mutation generate a 198-bp fragment. The CT substitution at bp 677 creates a HinfI recognition sequence. If the mutation is present, then HinfI digests the 198-bp fragment into 175- and 23-bp fragments that can then be analyzed by electrophoresis.

The common MTR polymorphism that causes an A-to-G transition at bp 2756, converting an aspartic acid (D-919) into glycine, was identified according to the method of Van Der Put et al (25). Amplified PCR products were digested with the restriction enzyme HaeIII to yield a 265-bp fragment for the wild type 265, 180- and 80-bp fragments for the heterozygote, and 180- and 85-bp fragments for the homozygous recessive genotype.

The MTHFR 1298AC polymorphism was measured by using the technique described by Van Der Put et al (26). An amplified PCR product of 163 bp was digested with the restriction enzyme MboII to yield 56-, 31-, 30-, 28-, and 18-bp fragments for the wild type and 84-, 31-, 30-, and 18-bp fragments for the homozygous recessive genotype.

The MTRR A-to-G transition at bp 66 converting an isoleucine into a methionine residue was identified according to a modified method of Wilson et al (27) and was tested by creating an artificially generated NdeI restriction site. After NdeI digestion, a PCR fragment of 301 bp remains uncut in the presence of the G allele but is cut into 279- and 22-bp fragments in the presence of the A allele.

The common CBS polymorphism that causes an insertion of 68 bp at the 844 position was identified according to the method of Tsai et al (14). An amplified PCR product of 174 bp was obtained for the wild type, and 242- and 174-bp fragments were obtained for the heterozygote. The 242-bp fragment was digested by BsrI into 200- and 42-bp fragments. The RFC 80GA polymorphism was identified as described elsewhere (15).

Statistical analysis
Statistical analyses were performed by using STATVIEW-5 (SAS Institute Inc, Cary, NC) on a personal computer. Differences in allele frequencies and genotype distribution among the different groups were assessed by chi-square analysis. Results are presented as means ± SEMs. A Mann-Whitney test was used to analyze the effect of vitamin supplementation on blood parameters. A Kruskal-Wallis test was used to measure the effect of age on homocysteine values in different groups. A Bonferroni test was done to check that positive results were not due to the multiple-comparison procedure. Both significant values for the Mann-Whitney and Bonferroni tests are given in the tables concerned. P values <0.05 were considered statistically significant. A Friedman test was done for each potential interaction: age by treatment, case-control by genotype distribution, and genotype by group.

	RESULTS

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

 
The mean fasting plasma tHcy concentration of the trisomy 21 cases who did not take supplemental vitamins (ie, nonusers: 9.8 ± 0.7 µmol/L) was not significantly different from that of the controls (9.4 ± 0.3 µmol/L). There were also no significant differences between nonusers and controls in RBC folate, serum folate, or serum vitamin B-12 (Table 1). With folate supplementation alone, the mean tHcy concentration in the cases (7.6 ± 0.3 µmol/L) was significantly lower than that in the controls (P < 0.0003, Mann-Whitney test) and was significantly different from that in nonusers. When folic acid was combined with vitamin B-12, plasma tHcy (7.1 ± 0.6 µmol/L) was not significantly different from that in folic acid users. Subjects who took folic acid combined with vitamin B-6 had lower plasma tHcy concentrations (8.0 ± 0.6 µmol/L) than did the nonusers, but not significantly so. The plasma tHcy concentration was also significantly lower in cases who took folic acid, vitamin B-12, and vitamin B-6 combined (6.5 ± 0.5 µmol/L) than in nonusers (P < 0.0001). With folic acid use only, RBC folate and serum folate concentrations were 3.7 and 10.7 times, respectively, the concentrations of nonusers. There was no significant difference in the serum vitamin B-12 concentration with folate use alone.

Table 2

Table 3

View this table: [in this window] [in a new window]   TABLE 3 MTHFR 677CT and 1298AC, MTR 2756AG, MTRR 66AG, RFC 80GA, and CBS 844Ins68 gene polymorphism distributions and allele frequencies in trisomy 21 cases and controls1

Table 4

Table 4

View this table: [in this window] [in a new window]   TABLE 4 Plasma total homocysteine concentrations according to MTHFR 677CT and 1298AC, MTR 2756AG, MTRR 66AG, RFC 80GA, and CBS 844Ins68 gene polymorphisms1

	DISCUSSION

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

In earlier studies involving other trisomy 21 patients, we (AC) and other investigators showed that the plasma tHcy concentration is lower in trisomy 21 (31, 32). Lower plasma tHcy concentrations are consistent with a functional folate deficiency resulting from CBS over-expression and higher enzyme activity. The plasma tHcy results in the present trisomy 21 population (the nonusers), however, failed to confirm these main findings between trisomy 21 patients and controls.

Concerning amino acid concentrations in persons with trisomy 21, Lejeune et al (33) showed that the concentrations of serum amino acids in persons with trisomy 21 differ from the concentrations in healthy controls. That study covered a mixed age range and included many adults. However, Heggarty et al (34) failed to find the decrease in aminothiols (cysteine, cystathionine, and methionine) and serine that is supposed to occur in persons with trisomy 21. It is not clear why the fasting plasma tHcy concentration in the nonusers in the present study was not significantly different from that of the controls. One possible explanation for our failure to confirm reduced tHcy concentrations in trisomy 21 may be that we measured tHcy in adults, who have a much lower folate requirement for growth. As shown in Table 4, the MTHFR 677CT polymorphism was not associated with any significant variation in plasma tHcy concentrations among cases or controls. In the absence of a control group of vitamin users, it is difficult to conclude whether the slight increase in tHcy seen in the controls but not in the cases homozygous for the T allele reflected different metabolic backgrounds of the cases and controls. Another issue to study is whether controls would have a similar 10-fold increase in folate concentrations with folic acid vitamin use. If the increase in serum folate in persons with Down syndrome is greater than that in healthy persons, it could indicate a functional folate deficiency (ie, folate is not retained inside the cells).

The genotype results for CBS confirm the earlier report of Ge et al (30) showing a significant high frequency of the CBS Ins68+/– genotype in trisomy 21. On the other hand, we found a lower frequency of the homozygous RFC1 80AAA genotype in cases than in controls (Table 3). In the present study, tHcy concentrations in cases who used folic acid + vitamin B-6 + vitamin B-12 were higher in RFC1 80AAA homozygotes than in heterozygotes (Table 4). However, because only 3 individuals had the 80AAA genotype, these findings must be verified in a larger sample. The real effect of this polymorphism in folate transport is not yet known. Patients with trisomy 21 have been reported to have abnormal sensitivity to methotrexate and have been found to have characteristic in vivo and in vitro methotrexate toxicity (35, 36). The RFC gene on chromosome 21 is involved in maternal-to-fetal transplacental folate transport. A dosage effect in trisomy 21 and a possible functional effect of the polymorphism may contribute toward an imbalance of one-carbon-derived metabolites.

In conclusion, the present study may contribute information concerning the link between the metabolic consequences of additional chromosomal material and polymorphisms in genes related to vitamin metabolism. The study was performed in adults and did not take into account any specific clinical features of down syndrome. Our results suggest that there are no significant differences in fasting blood tHcy concentrations between healthy controls and adult trisomy 21 patients that justify B vitamin supplementation.

	ACKNOWLEDGMENTS

 
We thank all those participating in this study and the Centre Médical Jérome Lejeune and the Centre de Medecine Preventive of Nancy. We also thank Bérangère Marie for her contribution to the statistical analysis.

NF-E contributed to collecting samples, analyzing the laboratory data, and writing the manuscript. AC was co-principal investigator and was responsible for analyzing the laboratory data and writing the manuscript. CM was the clinician investigator responsible for collecting and managing the patients and samples during the study. FB contributed to analyzing the laboratory data. HB was a protocol designer responsible for collecting and managing the patients and samples during the study. BH was a protocol designer responsible for collecting and managing the control samples during the study. DSR contributed to data interpretation and the writing of the manuscript. M-OR was co-principal investigator and is the head of the Jerome Lejeune Medical Center. DL was the senior researcher, a protocol designer, and responsible for evaluating the statistics. JPN was co-principal investigator and is the head of the medical biochemistry research group. None of the authors had any conflicts of interest.

See the corresponding letter to the Editor on page XXX.

	REFERENCES

TOP ABSTRACT INTRODUCTION SUBJECTS AND METHODS RESULTS DISCUSSION REFERENCES

 

1. Epstein CJ. Down syndrome. In: Rosenberg RN, Prusiner SB, DiMauro S, Barchi RL, eds. The molecular and genetic basis of neurological disease. Boston: Butterworth-Heinemann, 1997:51–79.
2. Korenberg JR, Chen XN, Schipper R, et al. Down syndrome phenotypes: the consequences of chromosomal imbalance. Proc Natl Acad Sci U S A 1994;91:4997–5001.[Abstract/Free Full Text]
3. Epstein CJ. Epilogue: toward the twenty-first century with Down syndrome—a personal view of how far we have come and of how far we can reasonably expect to go. Prog Clin Biol Res 1995;393:241–6.[Medline]
4. Reeves RH. Down's syndrome. A complicated genetic insult. Lancet 2001;358(suppl):S23 (letter).[Medline]
5. Epstein CJ, Korenberg JR, Anneren G, et al. Protocols to establish genotype-phenotype correlations in Down syndrome. Am J Hum Genet 1991;49:207–35.[Medline]
6. Munke M, Kraus JP, Ohura T, Francke U. The gene for cystathionine beta-synthase (CBS) maps to the subtelomeric region on human chromosome 21q and to proximal mouse chromosome 17. Am J Hum Genet 1988;42:550–9.[Medline]
7. Mudd SH, Levy HL. Plasma homocyst(e)ine or homocysteine? N Engl J Med 1995;333:325 (letter).[Free Full Text]
8. Chadefaux-Vekemans B, Coude M, Muller F, et al. Methylenetetrahydrofolate reductase polymorphism in the etiology of Down syndrome. Pediatr Res 2002;51:766–7.[Medline]
9. Finkelstein JD, Martin JJ, Harris BJ. Methionine metabolism in mammals. The methionine-sparing effect of cystine. J Biol Chem 1988;263:11750–4.[Abstract/Free Full Text]
10. Malinow MR, Duell PB, Hess DL, et al. Reduction of plasma homocyst(e)ine levels by breakfast cereal fortified with folic acid in patients with coronary heart disease. N Engl J Med 1998;338:1009–15.[Abstract/Free Full Text]
11. Leclerc D, Campeau E, Goyette P, et al. Human methionine synthase: cDNA cloning and identification of mutations in patients of the cblG complementation group of folate/cobalamin disorders. Hum Mol Genet 1996;5:1867–74.[Abstract/Free Full Text]
12. Harmon DL, Shields DC, Woodside JV, et al. Methionine synthase D919G polymorphism is a significant but modest determinant of circulating homocysteine concentrations. Genet Epidemiol 1999;17:298–309.[Medline]
13. Bosco P, Gueant-Rodriguez RM, Anello G, et al. Methionine synthase (MTR) 2756 (A > G) polymorphism, double heterozygosity methionine synthase 2756 AG/methionine synthase reductase (MTRR) 66 AG, and elevated homocysteinemia are three risk factors for having a child with Down syndrome. Am J Med Genet 2003;121A:219–24.
14. Tsai MY, Garg U, Key NS, Hanson NQ, Suh A, Schwichtenberg K. Molecular and biochemical approaches in the identification of heterozygotes for homocystinuria. Atherosclerosis 1996;122:69–77.[Medline]
15. Chango A, Emery-Fillon N, de Courcy GP, et al. A polymorphism (80G->A) in the reduced folate carrier gene and its associations with folate status and homocysteinemia. Mol Genet Metab 2000;70:310–5.[Medline]
16. Winkelmayer WC, Eberle C, Sunder-Plassmann G, Fodinger M. Effects of the glutamate carboxypeptidase II (GCP2 1561CT) and reduced folate carrier (RFC1 80G>A) allelic variants on folate and total homocysteine levels in kidney transplant patients. Kidney Int 2003;63:2280–5.[Medline]
17. Morin I, Devlin AM, Leclerc D, et al. Evaluation of genetic variants in the reduced folate carrier and in glutamate carboxypeptidase II for spina bifida risk. Mol Genet Metab 2003;79:197–200.[Medline]
18. De Marco P, Calevo MG, Moroni A, et al. Reduced folate carrier polymorphism (80A–>G) and neural tube defects. Eur J Hum Genet 2003;11:245–52.[Medline]
19. Barbe F, Abdelmouttaleb I, Chango A, et al. Detection of moderate hyperhomocysteinemia: comparison of the Abbott fluorescence polarization immunoassay with the Bio-Rad and SBD-F high-performance liquid chromatographic assays. Amino Acids 2001;20:435–40.[Medline]
20. Ubbink JB, Vermaak WJ, Bissbort SH. High-performance liquid chromatographic assay of human lymphocyte kynureninase activity levels. J Chromatogr 1991;566:369–75.[Medline]
21. Ubbink JB, Hayward Vermaak WJ, Bissbort S. Rapid high-performance liquid chromatographic assay for total homocysteine levels in human serum. J Chromatogr 1991;565:441–6.[Medline]
22. Frosst P, Blom HJ, Milos R, et al. A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase. Nat Genet 1995;10:111–3.[Medline]
23. Goyette P, Frosst P, Rosenblatt DS, Rozen R. Seven novel mutations in the methylenetetrahydrofolate reductase gene and genotype/phenotype correlations in severe methylenetetrahydrofolate reductase deficiency. Am J Hum Genet 1995;56:1052–9.[Medline]
24. Van der Put NM, Steegers-Theunissen RP, Frosst P, et al. Mutated methylenetetrahydrofolate reductase as a risk factor for spina bifida. Lancet 1995;346:1070–1.[Medline]
25. Van der Put NM, Eskes TK, Blom HJ. Is the common 677C–>T mutation in the methylenetetrahydrofolate reductase gene a risk factor for neural tube defects? A meta-analysis. QJM 1997;90:111–5.[Abstract/Free Full Text]
26. Van der Put NM, Gabreels F, Stevens EM, et al. A second common mutation in the methylenetetrahydrofolate reductase gene: an additional risk factor for neural-tube defects? Am J Hum Genet 1998;62:1044–51.[Medline]
27. Wilson A, Platt R, Wu Q, et al. A common variant in methionine synthase reductase combined with low cobalamin (vitamin B12) increases risk for spina bifida. Mol Genet Metab 1999;67:317–23.[Medline]
28. de la Monte SM. Molecular abnormalities of the brain in Down syndrome: relevance to Alzheimer's neurodegeneration. J Neural Transm Suppl 1999;57:1–19.[Medline]
29. Wang HX, Wahlin A, Basun H, Fastbom J, Winblad B, Fratiglioni L. Vitamin B(12) and folate in relation to the development of Alzheimer's disease. Neurology 2001;56:1188–94.[Free Full Text]
30. Ge Y, Jensen T, James SJ, et al. High frequency of the 844ins68 cystathionine-beta-synthase gene variant in Down syndrome children with acute myeloid leukemia. Leukemia 2002;16:2339–41.[Medline]
31. Chadefaux B, Ceballos I, Hamet M, et al. Is absence of atheroma in Down syndrome due to decreased homocysteine levels? Lancet 1988;2:741 (letter).
32. Pogribna M, Melnyk S, Pogribny I, Chango A, Yi P, James SJ. Homocysteine metabolism in children with Down syndrome: in vitro modulation. Am J Hum Genet 2001;69:88–95.[Medline]
33. Lejeune J, Rethore MO, de Blois MC, et al. Amino acids and trisomy 21. Ann Genet 1992;35:8–13.[Medline]
34. Heggarty HJ, Ball R, Smith M, Henderson MJ. Amino acid profile in Down's syndrome. Arch Dis Child 1996;74:347–9.[Abstract]
35. Peeters MA, Rethore MO, Lejeune J. In vivo folic acid supplementation partially corrects in vitro methotrexate toxicity in patients with Down syndrome. Br J Haematol 1995;89:678–80.[Medline]
36. Peeters MA, Salabelle A, Attal N, et al. Excessive glutamine sensitivity in Alzheimer's disease and Down syndrome lymphocytes. J Neurol Sci 1995;133:31–41.[Medline]

This article has been cited by other articles:

				T. Tamura and M. F. Picciano Folate and human reproduction Am. J. Clinical Nutrition, May 1, 2006; 83(5): 993 - 1016. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Fillon-Emery, N. Articles by Nicolas, J. P. Search for Related Content PubMed PubMed Citation Articles by Fillon-Emery, N. Articles by Nicolas, J. P. Agricola Articles by Fillon-Emery, N. Articles by Nicolas, J. P.