Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DNA methylation profiling”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 811 records · Page 45Linked to original sources
PubMed2003

Copper induces the expression of cholesterogenic genes in human macrophages.

Accumulation of lipids and cholesterol by macrophages and subsequent transformation into foam cells are key features in development of atherosclerosis. Serum copper concentrations have been shown to be associated with cardiovascular disease. However, the mechanism behind the proatherogenic effect of copper is not clear. We used DNA microarrays to define the changes in gene expression profile in response to copper exposure of human macrophages. Expression monitoring by DNA microarray revealed 91 genes that were regulated. Copper increased the expression of seven cholesterogenic genes (3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) synthase, IPP isomerase, squalene synthase, squalene epoxidase, methyl sterol oxidase, H105e3 mRNA and sterol-C5-desaturase) and low-density lipoprotein receptor (LDL-R), and decreased the expression of CD36 and lipid binding proteins. The expression of LDL-R and HMG CoA reductase was also investigated using real time PCR. The expression of both of these genes was increased after copper treatment of macrophages (P<0.01 and P<0.01, respectively). We conclude that copper activates cholesterogenic genes in macrophages, which may provide a mechanism for the association between copper and atherosclerosis. The effect of copper on cholesterogenic genes may also have implications for liver steatosis in early stages of Wilson's disease.

Cholesterol↗
PubMed1994

A deficiency for nucleotide excision repair strongly potentiates the mutagenic effectiveness of methyl bromide in Drosophila.

A DNA repair assay measuring hypermutability response in the absence of nucleotide excision repair (NER) was employed to study the impact of a deficiency in NER on the induction of forward mutations (X-chromosomal recessive lethals) by methyl bromide (MeBr) in Drosophila melanogaster. Postmeiotic male germ-cell stages reacted with MeBr were introduced in either NER competent oocytes (exr+) or in cells from the NER- strain mus-201. The high average M(exr-)/M(exr+) hypermutability ratio of 8.3 determined for MeBr is similar to the M(exr-)/M(exr+) indices found for other monofunctional alkylating agents with high Swain-Scott s values, such as methyl methanesulphonate and dimethyl sulphate. It is concluded that the genotoxic profile of methyl bromide is in keeping with those of high s-value alkylating agents but it seems incompatible with a methylating agent producing substantial amounts of O6 methylguanine.

Alkylating Agents↗
PubMed2001

Analysis of chromatin-immunopurified MeCP2-associated fragments.

As molecular biologists, we are continuing to unravel the interactions by which DNA binding proteins mediate the expression of genes. The chromatin immunoprecipitation (ChIP) technique provides us with an exquisite tool to investigate the interplay between chromatin structure and its role in regulating transcription, replication, and recombination in vivo. We describe a robust assay used to identify the molecular determinants associated with chromatin. In this article we illustrate the ChIP technique and use the transcriptionally silent-hypermethylated multidrug resistance (MDR1) gene as the platform for methyl-CpG binding protein 2 (MeCP2) localization on chromatin. Driven by the hypothesis that repression is strongly dependent on the methylation profile of the endogenous promoter, we demonstrate that MDR1 is targeted by MeCP2. Methylated MDR1 chromatin is highly enriched with MeCP2 and is in striking contrast to localization observed in cells in which MDR1 is transcriptionally active. In a distinct model system we discuss experimental methods used to immunopurify the MeCP2 repressor complex on chromatin and quantify protein-DNA association by competitive PCR approach.

Cell Line↗
PubMed2005

Role of the alkali labile sites, reactive oxygen species and antioxidants in DNA damage induced by methylated trivalent metabolites of inorganic arsenic.

In the last decade arsenic metabolism has become an important matter of discussion. Methylation of inorganic arsenic (iAs) to monomethylarsonic acid (MMA(V)) and dimethylarsinic acid (DMA(V)) is considered to decrease arsenic toxicity. However, in addition to these pentavalent metabolites, the trivalent metabolites monomethylarsonous (MMA(III)) and dimethylarsinous acid (DMA(III)) have been identified recently as intermediates in the metabolic pathway of arsenic in cultured human cells. To examine the role of oxidative damage in the generation of DNA strand breaks by methylated trivalent arsenic metabolites, we treated human lymphocytes with both metabolites at non-cytotoxic concentrations. We further tested whether these effects are sensitive to modulation by the antioxidants ascorbate (Vitamin C) and selenomethionine (Se-Met). Both trivalent metabolites produced oxidative stress related DNA damage, consisting of single strand breaks and alkali-labile sites, with MMA(III) being more potent at low concentrations than DMA(III). Neither MMA(III) nor DMA(III) induced DNA-double strand breaks. The oxidative stress response profiles of the metabolites were parallel as determined by lipid peroxidation induction. MMA(III) induced peroxidation from the lowest concentration tested, while effects of DMA(III) were apparent only at concentrations above 10 muM. The antioxidant Se-Met exhibited a more pronounced inhibition of trivalent arsenic metabolite-induced oxidative-DNA damage than did vitamin C. The present findings suggest that DNA damage by methylated trivalent metabolites at non-cytotoxic concentrations may be mediated by a mix of reactive oxygen and nitrogen oxidized species.

Adult↗
PubMed1991

Methylation status of CpG-rich islands on active and inactive mouse X chromosomes.

Single copy probes derived from CpG-rich island clones from Eag I and Not I linking libraries and nine rare-cutter restriction endonucleases were used to investigate the methylation status of CpG-rich islands on the inactive and active X chromosomes (Chr) of the mouse. Thirteen of the 14 probes used detected CpG-rich islands in genomic DNA. The majority of island CpGs detected by rare-cutter restriction endonucleases were methylated on the inactive X Chr and unmethylated on the active X Chr, but some heterogeneity within the cell population used to make genomic DNA was detected. The CpG-rich islands detected by two putative pseudoautosomal probes remained unmethylated on both the active and inactive X Chrs. Otherwise, distance from the X Chr inactivation center did not affect the methylation profile of CpG-rich islands. We conclude that methylation of CpG-rich islands is a general feature of X Chr inactivation.

Animals↗
PubMed1986

Conformational analysis of the octamer d(G-G-C-C-G-G-C-C) in aqueous solution. A one-dimensional and two-dimensional proton NMR study at 300 MHz and 500 MHz.

Proton NMR studies in H2O and 2H2O were carried out on the self-complementary DNA octamer d(G-G-C-C-G-G-C-C) and compared with similar studies on the dimethylated analogue d(G-G-m5C-m5C-G-G-C-C) [Sanderson, M. R., Mellema, J.-R., van der Marel, G. A., Wille, G., van Boom, J. H. & Altona, C. (1983) Nucleic Acids Res. 11, 3333-3346]. Base, H1', H2' and H2" resonances were assigned by means of two-dimensional nuclear Overhauser spectroscopy (NOESY) and correlated spectroscopy (COSY) experiments. Chemical shift vs temperature profiles were used to analyze the temperature-dependent conformational behaviour and to extract thermodynamic parameters for the duplex-to-coil transition. Analysis of proton-proton couplings were used to discriminate between J1'2' and J1'2" and between the H2' and H2" resonances as well as to obtain conformational parameters of the sugar rings. From the NOESY and COSY experiments, the temperature profiles and the analysis of the coupling data it is concluded that: (a) the compound adopts a B-DNA-type helix in solution; (b) the sugar rings in the intact duplex display limited conformational freedom; (c) methylation of the cytosine bases at the C5 position has no measurable effect on the conformational behaviour of the octamer, nor on the conformation of the sugar rings; however, methylation does increase the stability of the duplex in aqueous solution under conditions of low salt concentration.

Carbohydrate Conformation↗
PubMed2006

Knockdown of ALR (MLL2) reveals ALR target genes and leads to alterations in cell adhesion and growth.

ALR (MLL2) is a member of the human MLL family, which belongs to a larger SET1 family of histone methyltransferases. We found that ALR is present within a stable multiprotein complex containing a cohort of proteins shared with other SET1 family complexes and several unique components, such as PTIP and the jumonji family member UTX. Like other complexes formed by SET1 family members, the ALR complex exhibited strong H3K4 methyltransferase activity, conferred by the ALR SET domain. By generating ALR knockdown cell lines and comparing their expression profiles to that of control cells, we identified a set of genes whose expression is activated by ALR. Some of these genes were identified by chromatin immunoprecipitation as direct ALR targets. The ALR complex was found to associate in an ALR-dependent fashion with promoters and transcription initiation sites of target genes and to induce H3K4 trimethylation. The most characteristic features of the ALR knockdown cells were changes in the dynamics and mode of cell spreading/polarization, reduced migration capacity, impaired anchorage-dependent and -independent growth, and decreased tumorigenicity in mice. Taken together, our results suggest that ALR is a transcriptional activator that induces the transcription of target genes by covalent histone modification. ALR appears to be involved in the regulation of adhesion-related cytoskeletal events, which might affect cell growth and survival.

Animals↗
PubMed2006

Diverse epigenetic profile of novel human embryonic stem cell lines.

Human embryonic stem cells (hESCs) are a promising model for studying mechanisms of regulation of early development and differentiation. OCT4, NANOG, OCT4-related genes and some others were recently described to be important in pluripotency maintenance. Lesser is known about molecular mechanisms involved in their regulation. Apart from genetic regulation of gene expression epigenetic events, particularly methylation, play an important role in early development. Using RT-PCR we studied the expression of pluripotency-related genes OCT4, NANOG, DPPA3 and DPPA5 during hESCs differentiation to embryoid bodies. Analysis of methylation profiles of promoter or putative regulatory regions of the indicated genes demonstrated that expression of the pluripotency-maintaining genes correlated with their methylation status, whereas methylation of DPPA3 and DPPA5 varied between cell lines. We propose that DNA methylation underlies the developmental stage-specific mechanisms of pluripotency-related genes expression and reactivation and may have an impact on differentiation potential of hESC lines.

5' Flanking Region↗
PubMed2001

Cytosine-phosphoguanine methylation of estrogen receptors in endometrial cancer.

We hypothesize that estrogen receptors (ERs) are differentially expressed in endometrial cancer. To test this hypothesis, we investigated the expression profile of ERalpha (ERalpha-A, ERalpha-B, ERalpha-C) and ERbeta genes and CpG methylation status in endometrial cancer cell lines and tissues using reverse transcription-PCR and methylation-specific PCR and direct DNA sequencing. The results demonstrated that ERalpha-A, ERalpha-B, and ERbeta were normally expressed whereas ERalpha-C gene was inactivated in all endometrial cancer cell lines. We further investigated the mechanisms of ERalpha-C gene inactivation through CpG methylation pathways. The treatment with demethylating agent (5'-aza-2'-deoxycytidine) restored ERalpha-C gene expression in all endometrial cancer cell lines. We further confirmed these findings with methylation-specific PCR and direct DNA sequencing and found that only ERalpha-C was methylated on all five different CpG sites in all cell lines. We further analyzed 88 cancerous and 46 normal endometrial tissues. The results demonstrated that only ERalpha-C was inactivated and methylated in 94% of cancer tissues. In 32 pairs of cancerous and normal endometrial tissues from the same patient, ERalpha-C was methylated in 29 of 32 cancer tissues but unmethylated in all normal endometrial tissues. This is the first report that demonstrates selective ERalpha-C gene inactivation through CpG methylation pathway in uterine endometrial cancer.

CpG Islands↗
PubMed1989

Age-associated changes in DNA methylation and mRNA level of the c-myc gene in spleen and liver of mice.

Age-associated changes in DNA structure and mRNA level were studied for the c-myc gene in spleen and liver of mice at 2, 14 and 26 months of age. Neither amplification nor rearrangement of the gene was detected in either tissue at any age. However, a significant alteration was observed in the methylation profile. The profile of the gene and its vicinity was determined using various methylation-sensitive restriction enzymes. In both tissues, the gene had an unmethylated domain ranging from -2 kb upstream of the 5' end of the first exon to the 3' end of the first intron. It was flanked by partially methylated regions, where age-dependent changes as well as tissue specificity were observed. In the spleen, age-related hypomethylation was observed at both the 3' and 5' sides of the domain. In contrast, hypermethylation was found in the liver only at the 3' side. The steady-state level of c-myc mRNA showed a drastic decrease in liver from youth to middle age, while splenic mRNA changed little. The correlation between the changes of mRNA and DNA methylation is discussed.

Aging↗
PubMed2002

SGS1 is a multicopy suppressor of srs2: functional overlap between DNA helicases.

Sgs1 is a member of the RecQ family of DNA helicases, which have been implicated in genomic stability, cancer and ageing. Srs2 is another DNA helicase that shares several phenotypic features with Sgs1 and double sgs1srs2 mutants have a severe synthetic growth phenotype. This suggests that there may be functional overlap between these two DNA helicases. Consistent with this idea, we found the srs2Delta mutant to have a similar genotoxin sensitivity profile and replicative lifespan to the sgs1Delta mutant. In order to directly test if Sgs1 and Srs2 are functionally interchangeable, the ability of high-copy SGS1 and SRS2 plasmids to complement the srs2Delta and sgs1Delta mutants was assessed. We report here that SGS1 is a multicopy suppressor of the methyl methanesulphonate (MMS) and hydroxyurea sensitivity of the srs2Delta mutant, whereas SRS2 overexpression had no complementing ability in the sgs1Delta mutant. Domains of Sgs1 directly required for processing MMS-induced DNA damage, most notably the helicase domain, are also required for complementation of the srs2Delta mutant. Although SGS1 overexpression was unable to rescue the shortened mean replicative lifespan of the srs2Delta mutant, maximum lifespan was significantly increased by multicopy SGS1. We conclude that Sgs1 is able to partially compensate for the loss of Srs2.

Cell Division↗
PubMed2004

Promoter hypermethylation of cancer-related genes: a strong independent prognostic factor in acute lymphoblastic leukemia.

Promoter hypermethylation plays an important role in the inactivation of cancer-related genes. This abnormality occurs early in leukemogenesis and seems to be associated with poor prognosis in acute lymphoblastic leukemia (ALL). To determine the extent of hypermethylation in ALL, we analyzed the methylation status of the CDH1, p73, p16, p15, p57, NES-1, DKK-3, CDH13, p14, TMS-1, APAF-1, DAPK, PARKIN, LATS-1, and PTEN genes in 251 consecutive ALL patients. A total of 77.3% of samples had at least 1 gene methylated, whereas 35.9% of cases had 4 or more genes methylated. Clinical features and complete remission rate did not differ among patients without methylated genes, patients with 1 to 3 methylated genes (methylated group A), or patients with more than 3 methylated genes (methylated group B). Estimated disease-free survival (DFS) and overall survival (OS) at 11 years were 75.5% and 66.1%, respectively, for the nonmethylated group; 37.2% and 45.5% for methylated group A; and 9.4% and 7.8% for methylated group B (P < .0001 and P = .0004, respectively). Multivariate analysis demonstrated that the methylation profile was an independent prognostic factor in predicting DFS (P < .0001) and OS (P = .003). Our results suggest that the methylation profile may be a potential new biomarker of risk prediction in ALL.

Adolescent↗
PubMed2004

Kaiso is a genome-wide repressor of transcription that is essential for amphibian development.

DNA methylation in animals is thought to repress transcription via methyl-CpG specific binding proteins, which recruit enzymatic machinery promoting the formation of inactive chromatin at targeted loci. Loss of DNA methylation can result in the activation of normally silent genes during mouse and amphibian development. Paradoxically, global changes in gene expression have not been observed in mice that are null for the methyl-CpG specific repressors MeCP2, MBD1 or MBD2. Here, we demonstrate that xKaiso, a novel methyl-CpG specific repressor protein, is required to maintain transcription silencing during early Xenopus laevis development. In the absence of xKaiso function, premature zygotic gene expression occurs before the mid-blastula transition (MBT). Subsequent phenotypes (developmental arrest and apoptosis) strongly resemble those observed for hypomethylated embryos. Injection of wild-type human kaiso mRNA can rescue the phenotype and associated gene expression changes of xKaiso-depleted embryos. Our results, including gene expression profiling, are consistent with an essential role for xKaiso as a global repressor of methylated genes during early vertebrate development.

Animals↗
PubMed2001

Isolate PM1 populations are dominant and novel methyl tert-butyl ether-degrading bacterial in compost biofilter enrichments.

The gasoline additive MTBE, methyl tert-butyl ether, is a widespread and persistent groundwater contaminant. MTBE undergoes rapid mineralization as the sole carbon and energy source of bacterial strain PM1, isolated from an enrichment culture of compost biofilter material. In this report, we describe the results of microbial community DNA profiling to assess the relative dominance of isolate PM1 and other bacterial strains cultured from the compost enrichment. Three polymerase chain reaction (PCR)-based profiling approaches were evaluated: denaturing gradient gel electrophoresis (DGGE) analysis of 230 bp 16S rDNA fragments; thermal gradient gel electrophoresis (TGGE) analysis of 575 bp 16S rDNA fragments; and non-denaturing polyacrylamide gel electrophoresis of 300-1,500 bp fragments containing 16S/23S ribosomal intergenic transcribed spacer (ITS) regions. Whereas all three DNA profiling approaches indicated that PM1-like bands predominated in mixtures from MTBE-grown enrichments, ITS profiling provided the most abundant and specific sequence data to confirm strain PM1's presence in the enrichment. Moreover, ITS profiling did not produce non-specific PCR products that were observed with T/DGGE. A further advantage of ITS community profiling over other methods requiring restriction digestion (e.g. terminal restriction fragment length polymorphisms) was that it did not require an additional digestion step or the use of automated sequencing equipment. ITS bands, excised from similar locations in profiles of the enrichment and PM1 pure culture, were 99.9% identical across 750 16S rDNA positions and 100% identical across 691 spacer positions. BLAST comparisons of nearly full-length 16S rDNA sequences showed 96% similarity between isolate PM1 and representatives of at least four different genera in the Leptothrix subgroup of the beta-Proteobacteria (Aquabacterium, Leptothrix, Rubrivivax and Ideonella). Maximum likelihood and parsimony analyses of 1,249 nucleotide positions supported isolate PM1's position in a separate lineage within the Leptothrix subgroup.

Bacteria↗
PubMed2006

DNA methylation patterns in adenomas from FAP, multiple adenoma and sporadic colorectal carcinoma patients.

Colorectal adenomas have traditionally been regarded as homogeneous. The aim of our study was to identify molecular features that may differentiate sporadic adenomas from familial adenomas such as Familial Adenomatous Polyposis (FAP) and Multiple Adenoma patients. DNA methylation was tested at Methylated IN Tumor (MINT) loci (1,2,12,31) and the CpG promoter region of genes MLH1, HPP1, MGMT, p14ARF and p16INK4a in FAP-associated adenomas (33) from 5 patients with a known APC mutation (Group 1, FAP), adenomas (29) from 4 Multiple Adenoma patients (Group 2 Multiple), adenomas (14) from 3 patients with sporadic colorectal cancers showing high microsatellite instability (Group 3, MSI-H) and adenomas (16) from 7 patients, with sporadic colorectal cancers showing microsatellite stable or low level instability (Group 4, MSS/MSI-L). Aberrant Crypt Foci (ACFs), Hyperplastic Polyps (HPs) and cancers were also examined for methylation status as well as K-ras mutation. Multiple Adenoma patients were examined for germline polymorphisms in the base excision repair gene, MYH. The familial syndrome, FAP -associated adenomas showed a significantly low frequency of MINT methylation (15.5%,) compared to sporadic MSS/MSI-L-associated adenomas (35.5%). Group 3 (MSI-H) adenomas were different in that many showed serration and a high level of methylation (57.1%). Group 2, Multiple Adenoma cases, resembled sporadic MSS/MSI-L-associated adenomas. However the promoter regions of key genes, MGMT, p14ARF and p16INK4a were methylated to a greater extent than MINTs in both sporadic and familial adenomas. Genetic profiling of adenomas supports the concept that adenomas belonging to familial syndromes pursue a different pathway to tumorigenesis than their sporadic counterpar/ts from their earliest formation.

Adenoma↗
PubMedDate not supplied

[Hypermethylation as a potential prognostic factor and a clue to a better understanding of the molecular pathogenesis of medulloblastoma--results of a genomewide methylation scan].

BACKGROUND: The molecular mechanisms controlling initiation and progression of medulloblastomas are largely unclear. Changes in DNA methylation of promoter regions have been shown to disturb the expression of growth regulatory genes. PATIENTS AND METHODS: We evaluated DNA methylation patterns in 17 medulloblastomas, 5 stPNETs and 5 medulloblastoma cell lines using Restriction Landmark Genomic Scanning (RLGS), a method displaying up to 2.000 potential gene loci in a single gene. To test whether previously characterized tumor suppressor genes are affected by hypermethylation we performed MS-PCR for p15INK4B, p16INK4A, VHL, TP53 and E-cadherin. RESULTS: The analysis of RLGS profiles from tumors revealed an abundance of hypermethylation in primary tumors and cell lines. Extrapolated to the human genome with its approximately 36,000 genes a total of 420 loci become hypermethylated in the tumor genomes. The previously characterized medulloblastoma breakpoint cluster in 17p11.2 appears to be a hotspot for aberrant methylation. Cox regression analysis of survival data identified seven CpG islands for which hypermethylation is suggestive of a poor prognosis. MS-PCR analysis of known genes demonstrated hypermethylation of p16INK4A in a limited number of tumors. The pattern of DNA hypermethylation was similar in medulloblastomas and stPNETs. However, some CpG islands were shown to be specific for a tumor type, while others were shared targets. CONCLUSIONS: Hypermethylation is a common abnormality in primary medulloblastomas and supratentorial PNETs. Several hundreds of CpG islands are potential targets for methylation in medulloblastomas including the breakpoint cluster in 17p11.2. The methylation status of certain gene sequences appears to be associated with the clinical outcome. Promoter hypermethylation has an outstanding potential as a marker for the identification of novel tumor suppressors as well as diagnostic and therapeutic targets in medulloblastomas.

Adolescent↗
PubMed1995

A role for mismatch repair in production of chromosome aberrations by methylating agents in human cells.

We have shown previously that certain alkylation products, or alkylation derived lesions, which induce chromosome aberrations (abs) persist for at least two cell cycles in Chinese hamster ovary cells. The increase in abs in the second cycle after treatment contrasts with the classical observation of reduction in ab yield with successive mitoses following ionizing radiation. Here we present evidence that processing of lesions by mismatch repair is a mechanism for ab induction by methylating agents. Our previous studies implicated O6-methylguanine (O6MeG) as an important lesion in induction of abs, particularly in the second cell cycle after treatment. In the absence of repair of O6MeG by alkylguanine DNA alkyltransferase (AGT), new abs were induced in the second cycle after treatment with e.g. methylnitronitrosoguanidine (MNNG) and methylnitrosourea (MNU). Thus, we hypothesized that abs were produced not by O6MeG or its repair in the first S phase, but by subsequent processing of the lesions. We suggested that after replication proceeded past the O6MeG lesion in the first S phase, inserting an incorrect base on the newly synthesized strand, recognition and repair by mismatch repair in the second S phase led to a chromosome ab. Here we used MT1 cells, a human lymphoblastoid cell line that has a defect in strand-specific mismatch repair. MT1 cells are alkylation tolerant and have a mutator phenotype, compared with their parent line, TK6; both MT1 and TK6 cells lack AGT so do not remove the methyl group from O6MeG. While the initial levels of abs at the first metaphase were similar in MT1 and TK6 cells, ab levels in MT1 cells were greatly reduced in the second and third cell cycles following treatment with MNNG, dimethylnitrosamine and MNU, in contrast with the parent TK6 cells, which had more abs in the second cell cycle than in the first. This supports the hypothesis that repair of mismatched base pairs involving O6MeG is one mechanism for induction of chromosome abs. In contrast to the difference in response to methylating agents between TK6 cells and mismatch repair-deficient MT1 cells, the profile of ab induction by an ethylating agent, ethylnitronitrosourea, was similar in MT1 cells to those for TK6 cells and CHO cells.

Alkylating Agents↗
PubMed2001

Homocysteine metabolism in children with Down syndrome: in vitro modulation.

The gene for cystathionine beta-synthase (CBS) is located on chromosome 21 and is overexpressed in children with Down syndrome (DS), or trisomy 21. The dual purpose of the present study was to evaluate the impact of overexpression of the CBS gene on homocysteine metabolism in children with DS and to determine whether the supplementation of trisomy 21 lymphoblasts in vitro with selected nutrients would shift the genetically induced metabolic imbalance. Plasma samples were obtained from 42 children with karyotypically confirmed full trisomy 21 and from 36 normal siblings (mean age 7.4 years). Metabolites involved in homocysteine metabolism were measured and compared to those of normal siblings used as controls. Lymphocyte DNA methylation status was determined as a functional endpoint. The results indicated that plasma levels of homocysteine, methionine, S-adenosylhomocysteine, and S-adenosylmethionine were all significantly decreased in children with DS and that their lymphocyte DNA was hypermethylated relative to that in normal siblings. Plasma levels of cystathionine and cysteine were significantly increased, consistent with an increase in CBS activity. Plasma glutathione levels were significantly reduced in the children with DS and may reflect an increase in oxidative stress due to the overexpression of the superoxide dismutase gene, also located on chromosome 21. The addition of methionine, folinic acid, methyl-B(12), thymidine, or dimethylglycine to the cultured trisomy 21 lymphoblastoid cells improved the metabolic profile in vitro. The increased activity of CBS in children with DS significantly alters homocysteine metabolism such that the folate-dependent resynthesis of methionine is compromised. The decreased availability of homocysteine promotes the well-established "folate trap," creating a functional folate deficiency that may contribute to the metabolic pathology of this complex genetic disorder.

Adenosine↗