We investigated the association between conotruncal heart defects (CTDs) and maternal and fetal single nucleotide polymorphisms (SNPs) in 60 genes in the folate, homocysteine and pathways. We also investigated whether periconceptional maternal folic acid supplementation modified associations between CTDs and SNPs.
Participants were enrolled in the National Birth Defects Prevention Study between 1997 and 2007. DNA samples from 616 case-parental triads affected by CTDs and 1,645 control-parental triads were genotyped using an Illumina® Golden Gate custom SNP panel. A hybrid design analysis, optimizing data from case and control trios, was used to identify maternal and fetal SNPs associated with CTDs.
Among 921 SNPs, 17 maternal and 17 fetal SNPs had a Bayesian false-discovery probability (BFDP) of <0.8. Ten of the 17 maternal SNPs and 2 of the 17 fetal SNPs were found within the glutamate-cysteine ligase, catalytic subunit (GCLC) gene. Fetal SNPs with the lowest BFDP (rs2612101, rs2847607, rs2847326, rs2847324) were found within the thymidylate synthetase (TYMS) gene. Additional analyses indicated that the risk of CTDs associated with candidate SNPs was modified by periconceptional folic acid supplementation. Nineteen maternal and 9 fetal SNPs had BFDP <0.8 for gene-by-environment (GxE) interactions with maternal folic acid supplementation.
These results support previous studies suggesting that maternal and fetal SNPs within folate, homocysteine and transsulfuration pathways are associated with CTD risk. Maternal use of supplements containing folic acid may modify the impact of SNPs on the developing heart.
conotruncal heart defects; single nucleotide polymorphisms; oxidative stress; genetics; folic acid; gene X environment interaction
Evidence exists for an association between use of vitamin supplements with folic acid in early pregnancy and reduced risk for offspring with conotruncal heart defects. A few observations have been made about nutrients related to one-carbon metabolism other than folate. Our prospective study attempted to extend information on nutrition and conotruncal heart defects by measuring analytes in mid-pregnancy sera.
This study included data from a repository of women’s mid-pregnancy serum specimens based on screened pregnancies in California from 2002–07. Each woman’s specimen was linked with delivery information to determine whether her fetus had a conotruncal heart defect or another structural malformation, or was nonmalformed. We identified 140 conotruncal cases and randomly selected 280 specimens as nonmalformed controls. Specimens were tested for a variety of analytes including: homocysteine, methylmalonic acid, folate, vitamin B12, pyridoxal phosphate, pyridoxal, pyridoxic acid, riboflavin, total choline, betaine, methionine, cysteine, cystathionine, arginine, asymmetric and symmetric dimethylarginine.
Results and Conclusions
We did not observe statistical evidence for substantial differences between cases and controls for any of the measured analytes. Analyses specifically targeting B-vitamins also did not reveal differences between cases and controls.
heart defects; nutrition; pregnancy; folic acid; B vitamins
Plasmodium vivax malaria causes significant morbidity and mortality worldwide, and only one drug is in clinical use that can kill the hypnozoites that cause P. vivax relapses. HIV and P. vivax malaria geographically overlap in many areas of the world, including South America and Asia. Despite the increasing body of knowledge regarding HIV protease inhibitors (HIV PIs) on P. falciparum malaria, there are no data regarding the effects of these treatments on P. vivax's hypnozoite form and clinical relapses of malaria. We have previously shown that the HIV protease inhibitor lopinavir-ritonavir (LPV-RTV) and the antibiotic trimethoprim sulfamethoxazole (TMP-SMX) inhibit Plasmodium actively dividing liver stages in rodent malarias and in vitro in P. falciparum, but effect against Plasmodium dormant hypnozoite forms remains untested. Separately, although other antifolates have been tested against hypnozoites, the antibiotic trimethoprim sulfamethoxazole, commonly used in HIV infection and exposure management, has not been evaluated for hypnozoite-killing activity. Since Plasmodium cynomolgi is an established animal model for the study of liver stages of malaria as a surrogate for P. vivax infection, we investigated the antimalarial activity of these drugs on Plasmodium cynomolgi relapsing malaria in rhesus macaques. Herein, we demonstrate that neither TMP-SMX nor LPV-RTV kills hypnozoite parasite liver stage forms at the doses tested. Because HIV and malaria geographically overlap, and more patients are being managed for HIV infection and exposure, understanding HIV drug impact on malaria infection is important.
Conotruncal heart defects (CTDs) are among the most severe birth defects worldwide. Studies of CTDs indicate both lifestyle behaviors and genetic variation contribute to the risk of CTDs. Based on a hybrid design using data from 616 case-parental and 1645 control-parental triads recruited for the National Birth Defects Prevention Study between 1997 and 2008, we investigated whether the occurrence of CTDs is associated with interactions between 921 maternal and/or fetal single nucleotide polymorphisms (SNPs) and maternal obesity and tobacco use. The maternal genotypes of the variants in the glutamate-cysteine ligase, catalytic subunit (GCLC) gene and the fetal genotypes of the variants in the glutathione S-transferase alpha 3 (GSTA3) gene were associated with an elevated risk of CTDs among obese mothers. The risk of delivering infants with CTDs among obese mothers carrying AC genotype for a variant in the GCLC gene (rs6458939) was 2.00 times the risk among those carrying CC genotype (95% confidence interval: 1.41, 2.38). The maternal genotypes of several variants in the glutathione-S-transferase (GST) family of genes and the fetal genotypes of the variants in the GCLC gene interacted with tobacco exposures to increase the risk of CTDs. Our study suggests that the genetic basis underlying susceptibility of the developing heart to the adverse effects of maternal obesity and tobacco use involve both maternal and embryonic genetic variants. These results may provide insights into the underlying pathophysiology of CTDs, and ultimately lead to novel prevention strategies.
Advanced maternal age and altered recombination are known risk factors for Down syndrome cases due to maternal non-disjunction of chromosome 21, whereas the impact of other environmental and genetic factors is unclear. The aim of this study was to investigate an association between low maternal socioeconomic status and chromosome 21 nondisjunction.
Data from 714 case and 977 control families were used to assess chromosome 21 meiosis I and meiosis II nondisjunction errors in the presence of three low socioeconomic status factors: (i) both parents had not completed high school, (ii) both maternal grandparents had not completed high school, and (iii) an annual household income of <$25,000. We applied logistic regression models and adjusted for covariates, including maternal age and race/ethnicity.
As compared with mothers of controls (n = 977), mothers with meiosis II chromosome 21 nondisjunction (n = 182) were more likely to have a history of one low socioeconomic status factor (odds ratio = 1.81; 95% confidence interval = 1.07–3.05) and ≥2 low socioeconomic status factors (odds ratio = 2.17; 95% confidence interval = 1.02–4.63). This association was driven primarily by having a low household income (odds ratio = 1.79; 95% confidence interval = 1.14–2.73). The same statistically significant association was not detected among maternal meiosis I errors (odds ratio = 1.31; 95% confidence interval = 0.81–2.10), in spite of having a larger sample size (n = 532).
We detected a significant association between low maternal socioeconomic status and meiosis II chromosome 21 non-disjunction. Further studies are warranted to explore which aspects of low maternal socioeconomic status, such as environmental exposures or poor nutrition, may account for these results.
chromosome nondisjunction; Down syndrome; risk factors; socioeconomic status; trisomy 21
We have previously shown that the HIV protease inhibitor lopinavir-ritonavir (LPV-RTV) and the antibiotic trimethoprim sulfamethoxazole (TMP-SMX) inhibit Plasmodium liver stages in rodent malarias and in vitro in P. falciparum. Since clinically relevant levels are better achieved in the non-human-primate model, and since Plasmodium knowlesi is an accepted animal model for the study of liver stages of malaria as a surrogate for P. falciparum infection, we investigated the antimalarial activity of these drugs on Plasmodium knowlesi liver stages in rhesus macaques. We demonstrate that TMP-SMX and TMP-SMX+LPV-RTV (in combination), but not LPV-RTV alone, inhibit liver stage parasite development. Because drugs that inhibit the clinically silent liver stages target parasites when they are present in lower numbers, these results may have implications for eradication efforts.
Background. Millions of individuals being treated for human immunodeficiency virus (HIV) live in malaria-endemic areas, but the effects of these treatments on malaria transmission are unknown. While drugs like HIV protease inhibitors (PIs) and trimethoprim-sulfamethoxazole (TMP-SMX) have known activity against parasites during liver or asexual blood stages, their effects on transmission stages require further study.
Methods. The HIV PIs lopinavir and saquinavir, the nonnucleoside reverse-transcriptase inhibitor nevirapine, and the antibiotic TMP-SMX were assessed for activity against Plasmodium falciparum transmission stages. The alamarBlue assay was used to determine the effects of drugs on gametocyte viability, and exflagellation was assessed to determine the effects of drugs on gametocyte maturation. The effects of drug on transmission were assessed by calculating the mosquito oocyst count as a marker for infectivity, using standard membrane feeding assays.
Results. Lopinavir and saquinavir have gametocytocidal and transmission blocking activities at or approaching clinically relevant treatment levels, while nevirapine does not. TMP-SMX is not gametocytocidal, but at prophylactic levels it blocks transmission.
Conclusions. Specific HIV treatments have gametocyte killing and transmission-blocking effects. Clinical studies are warranted to evaluate these findings and their potential impact on eradication efforts.
HIV; malaria; antiretrovirals; TMP-SMX; gametocytes; transmission
Studying empirically derived dietary patterns is useful in understanding dietary practice. We classified women by their dietary patterns using latent class analysis of 66 foods and studied the association of these patterns with neural tube defects (NTDs) and congenital heart defects (CHDs) in the US National Birth Defects Prevention Study (1997–2005). Logistic regression models used data from 1,047 with an NTD, 6,641 with a CHD, and 6,123 controls that were adjusted for maternal characteristics and tested the effect modification of multivitamin supplement use. Four latent dietary patterns were identified: prudent, Western, low-calorie Western, and Mexican. Among participants who did not use supplements, those in the Mexican, Western, and low-calorie Western classes were significantly more likely (odds ratios of 1.6, 1.5, and 1.4, respectively) to have offspring born with NTDs than were those in the prudent class after adjustment of for dietary folic acid intake. In contrast, among supplement users, there was no difference in the incidence of NTDs between classes. Associations between dietary class and CHD subgroups were not modified by supplement use except for tetralogy of Fallot; among supplement users, those in the Western class were twice as likely (95% confidence interval: 1.4, 2.8) as the prudent class to have offspring with tetralogy of Fallot. Women who adhered to a Western diet were 1.2 (95% confidence interval: 1.03, 1.35) times more likely to have an infant with septal heart defect than were women who adhered to a prudent diet. A prudent dietary pattern, even with folate fortification, may decrease the risk of NTDs and some heart defects.
congenital anomalies; diet; eating patterns; folic acid; latent class analysis
Both a lack of maternal folic acid supplementation and the presence of genetic variants that reduce enzyme activity in folate pathway genes have been linked to meiotic nondisjunction of chromosome 21; however, the findings in this area of research have been inconsistent. To better understand these inconsistencies, we asked whether maternal use of a folic acid-containing supplement before conception reduces risk for chromosome 21 nondisjunction. Using questionnaire data from the National Down Syndrome Project, a population-based case-control study, we compared the use of folic acid-containing supplements among mothers of infants with full trisomy 21 due to maternal nondisjunction (n=702) and mothers of infants born with no major birth defects (n=983). Using logistic regression, adjusting for maternal age, race/ethnicity, and infant age at maternal interview, we found no evidence of an association between lack of folic acid supplementation and maternal nondisjunction among all case mothers (OR=1.16; 95% CI: 0.90–1.48). In analyses stratified by meiotic stage and maternal age (<35 years or ≥ 35 years), we found an association among older mothers experiencing meiosis II nondisjunction errors (OR=2.00; 95% CI: 1.08–3.71). These data suggest that lack of folic acid supplementation may be associated specifically with MII errors in the aging oocyte. If confirmed, these results could account for inconsistencies among previous studies, as each study sample may vary by maternal age structure and proportion of meiotic errors.
Down syndrome; Trisomy 21; Aneuploidy; Nondisjunction; Chromosome segregation; Folic acid; Meiosis
In the apicomplexan protozoans motility and cell invasion are mediated by the TRAP/MIC2 family of transmembrane proteins, members of which link extracellular adhesion to the intracellular actomyosin motor complex. Here we characterize a new member of the TRAP/MIC2 family, named TRAP-Like Protein (TLP), that is highly conserved within the Plasmodium genus. Similar to the Plasmodium sporozoite protein, TRAP, and the ookinete protein, CTRP, TLP possesses an extracellular domain architecture that is comprised of von Willebrand factor A (vWA) and thrombospondin type 1 (TSP1) domains, plus a short cytoplasmic domain. Comparison of the vWA domain of TLP genes from multiple Plasmodium falciparum isolates showed relative low sequence diversity, suggesting that the protein is not under selective pressures of the host immune system. Analysis of transcript levels by quantitative reverse transcription polymerase chain reaction (RT-PCR) showed that TLP is predominantly expressed in salivary gland sporozoites of P. falciparum and P. berghei. Targeted disruption of P. berghei TLP resulted in a decreased capacity for cell traversal by sporozoites, and reduced infectivity of sporozoites in vivo, whereas in vitro sporozoite motility and hepatocyte invasion were unaffected. These results indicate a role of TLP in cell traversal by sporozoites.
Background. Although nonnucleoside reverse transcriptase inhibitors (NNRTIs) are usually part of first-line treatment regimens for human immunodeficiency virus (HIV), their activity on Plasmodium liver stages remains unexplored. Additionally, trimethoprim-sulfamethoxazole (TMP-SMX), used for opportunistic infection prophylaxis in HIV-exposed infants and HIV-infected patients, reduces clinical episodes of malaria; however, TMP-SMX effect on Plasmodium liver stages requires further study.
Methods. We characterized NNRTI and TMP-SMX effects on Plasmodium liver stages in vivo using Plasmodium yoelii. On the basis of these results, we conducted in vitro studies assessing TMP-SMX effects on the rodent parasites P. yoelii and Plasmodium berghei and on the human malaria parasite Plasmodium falciparum.
Results. Our data showed NNRTI treatment modestly reduced P. yoelii liver stage parasite burden and minimally extended prepatent period. TMP-SMX administration significantly reduced liver stage parasite burden, preventing development of patent parasitemia in vivo. TMP-SMX inhibited development of rodent and P. falciparum liver stage parasites in vitro.
Conclusions. NNRTIs modestly affect liver stage Plasmodium parasites, whereas TMP-SMX prevents patent parasitemia. Because drugs that inhibit liver stages target parasites when they are present in lower numbers, these results may have implications for eradication efforts. Understanding HIV drug effects on Plasmodium liver stages will aid in optimizing treatment regimens for HIV-exposed and HIV-infected infected patients in malaria-endemic areas.
Maternal nutritional status has been evaluated to clarify its role in development of neural tube defects (NTDs). Maternal folate intake during pregnancy has been closely evaluated for its association with NTDs.
The study objective was to examine associations between NTDs and other dietary periconceptional micronutrient intake, particularly nutrients involved in one-carbon metabolism or antioxidant activity.
Using data from the National Birth Defects Prevention Study, 1997–2005, logistic regression models were used to estimate the relative risk of NTDs based on maternal micronutrient intake.
Results were stratified according to folic acid supplement use, race/ethnicity, and maternal body mass index. Analyses included 954 cases (300 with anencephaly, 654 with spina bifida) and 6268 controls. Higher intakes of folate, thiamin, betaine, iron, and vitamin A were associated with decreased risk of anencephaly among some ethnic and clinical groups. In some groups, higher intakes of thiamin, riboflavin, vitamin B6, vitamin C, vitamin E, niacin, and retinol were associated with decreased risk of spina bifida.
In addition to folic acid, other micronutrients, including thiamin, betaine, riboflavin, vitamin B6, vitamin C, vitamin E, niacin, iron, retinol, and vitamin A, may decrease the risk of NTD occurrence.
Dietary periconceptional micronutrients; maternal nutrition; National Birth Defects Prevention Study; neural tube defects; one-carbon metabolism
The development of non-syndromic congenital heart defects (CHDs) involves a complex interplay of genetics, metabolism, and lifestyle. Previous studies have implicated maternal single nucleotide polymorphisms (SNPs) and altered metabolism in folate-related pathways as CHD risk factors.
We sought to discover associations between maternal SNPs and metabolites involved in the homocysteine, folate, and transsulfuration pathways, and determine if these associations differ between CHD cases and controls.
Genetic, metabolic, demographic, and lifestyle information was available for 335 mothers with CHD-affected pregnancies and 263 mothers with unaffected pregnancies. Analysis was conducted on 1160 SNPs, 13 plasma metabolites, and 2 metabolite ratios. A two-stage multiple linear regression was fitted to each combination of SNP and metabolite/ratio.
We identified 4 SNPs in the methionine adenosyltransferase II alpha (MAT2A) gene that were associated with methionine levels. Three SNPs in tRNA aspartic acid methyltransferase 1 (TRDMT1) gene were associated with total plasma folate levels. Glutamylcysteine (GluCys) levels were associated with multiple SNPs within the glutathione peroxidase 6 (GPX6) and O-6-methylguanine-DNA methyltransferase (MGMT) genes. The regression model revealed interactions between genotype and case-control status in the association of total plasma folate, total glutathione (GSH), and free GSH, to SNPs within the MGMT, 5,10-methenyltetrahydrofolate synthetase (MTHFS), and catalase (CAT) genes, respectively.
Our study provides further evidence that genetic variation within folate-related pathways accounts for inter-individual variability in key metabolites. We identified specific SNP-metabolite relationships that differed in mothers with CHD-affected pregnancies, compared to controls. Our results underscore the importance of multifactorial studies to define maternal CHD risk.
Maternal folic acid supplementation has been associated with a reduced risk for neural tube defects, and may be associated with a reduced risk for congenital heart defects, and other birth defects. Individuals with Down syndrome are at high risk for congenital heart defects and have been shown to have abnormal folate metabolism.
As part of the population-based case-control National Down Syndrome Project, 1011 mothers of infants with Down syndrome reported their use of folic acid-containing supplements. These data were used to determine whether lack of periconceptional maternal folic acid supplementation is associated with congenital heart defects in Down syndrome. We used logistic regression to test the relationship between maternal folic acid supplementation and the frequency of specific heart defects correcting for maternal race/ethnicity, proband sex, maternal use of alcohol and cigarettes, and maternal age at conception.
Lack of maternal folic acid supplementation was more frequent among infants with Down syndrome and atrioventricular septal defects (OR=1.69; 95% CI, 1.08–2.63; P=0.011) or atrial septal defects (OR=1.69; 95% CI, 1.11–2.58; P=0.007) than among infants with Down syndrome and no heart defect. Preliminary evidence suggests that the patterns of association differ by race/ethnicity and sex of the proband. There was no statistically significant association with ventricular septal defects (OR=1.26; 95% CI, 0.85–1.87; P=0.124).
Our results suggest that lack of maternal folic acid supplementation is associated with septal defects in infants with Down syndrome.
Atrial septal defect; Atrioventricular septal defect; Congenital heart defect; Down syndrome; Folic acid
Multiple investigators have established the feasibility of using buccal brush samples to genotype single nucleotide polymorphisms (SNPs) with high-density genome-wide microarrays, but there is currently no consensus on the accuracy of copy number variants (CNVs) inferred from these data. Regardless of the source of DNA, it is more difficult to detect CNVs than to genotype SNPs using these microarrays, and it therefore remains an open question whether buccal brush samples provide enough high-quality DNA for this purpose.
To demonstrate the quality of CNV calls generated from DNA extracted from buccal samples, compared to calls generated from blood samples, we evaluated the concordance of calls from individuals who provided both sample types. The Illumina Human660W-Quad BeadChip was used to determine SNPs and CNVs of 39 Arkansas participants in the National Birth Defects Prevention Study (NBDPS), including 16 mother-infant dyads, who provided both whole blood and buccal brush DNA samples.
We observed a 99.9% concordance rate of SNP calls in the 39 blood–buccal pairs. From the same dataset, we performed a similar analysis of CNVs. Each of the 78 samples was independently segmented into regions of like copy number using the Optimal Segmentation algorithm of Golden Helix SNP & Variation Suite 7.
Across 640,663 loci on 22 autosomal chromosomes, segment-mean log R ratios had an average correlation of 0.899 between blood-buccal pairs of samples from the same individual, while the average correlation between all possible blood-buccal pairs of samples from unrelated individuals was 0.318. An independent analysis using the QuantiSNP algorithm produced average correlations of 0.943 between blood-buccal pairs from the same individual versus 0.332 between samples from unrelated individuals.
Segment-mean log R ratios had an average correlation of 0.539 between mother-offspring dyads of buccal samples, which was not statistically significantly different than the average correlation of 0.526 between mother-offspring dyads of blood samples (p=0.302).
We observed performance from the subject-collected mail-in buccal brush samples comparable to that of blood. These results show that such DNA samples can be used for genome-wide scans of both SNPs and CNVs, and that high rates of CNV concordance were achieved whether using a change-point-based algorithm or one based on a hidden Markov model (HMM).
SNPs, Single nucleotide polymorphisms; CNVs, Copy number variants; NBDPS, National Birth Defects Prevention Study; Buccal brush
Inadequate folate status due to either genetic variation or nutritional deficiencies has been associated with an increased risk of congenital malformations including orofacial clefting, limb, cardiac and neural tube defects. Few epidemiologic studies have examined the association between limb reduction defects (LRDs) and folate-related genetic polymorphisms other than MTHFR 677C→T. We conducted a case–parental analysis of 148 families who participated in the National Birth Defects Prevention Study (NBDPS) to examine the association between nonsyndromic transverse and longitudinal LRDs with five single nucleotide polymorphisms (SNPs) in genes encoding for enzymes in folate and methionine pathways. Log-linear Poisson regression, adapted for analysis of case–parental data, assuming an additive genetic model was used to estimate genetic relative risks and 95% confidence intervals for the association between LRDs and each SNP. Among women who did not take multivitamin supplements, the MTHFR 677T variant acts via the offspring’s genome to increase the risk of LRDs. No association between LRDs and any fetal SNP was found among women who used multivitamin supplements. These results suggest the possibility that initiating folic acid supplementation prior to pregnancy may reduce the risk of having a LRD-affected pregnancy, especially in women whose offspring inherit one or two copies of the MTHFR 677T variant.
Folic acid; homocysteine; MTHFR; polymorphisms; case-parental analysis
Congenital heart defects (CHDs) are among the most prevalent and serious of birth defects. Multiple maternal factors are thought to contribute to CHD development including folate intake. Maternal DNA methylation, which is dependent on folate metabolism, may impact the risk of CHDs.
Our study was designed to determine whether maternal long interspersed nucleotide elements-1 (LINE-1) DNA hypomethylation is associated with increased occurrence of non-syndromic CHDs and whether maternal folate-dependent metabolites are correlated with DNA methylation status.
Using a case-control study design, we measured global DNA methylation status among mothers whose pregnancies were affected by non-syndromic CHDs (n=180) and mothers of unaffected pregnancies (n=187). Methylation of LINE-1 was used as a surrogate marker of global DNA methylation status. The association between DNA methylation and CHD risk was determined while adjusting for selected lifestyle factors.
LINE-1 DNA methylation was significantly lower in cases compared with controls (p=0.049). After covariate adjustments, a significant difference between cases and controls remained (p=0.010). Among women with LINE-1 methylation in the lowest decile of DNA methylation, the estimated risk of having a CHD-affected pregnancy was almost twice that of women in all other deciles (OR=1.91; 95% CI: 1.03, 3.58).
Our findings indicate that maternal LINE-1 DNA hypomethylation is associated with an increased risk of CHDs. Future studies investigating the association between maternal DNA methylation patterns and CHDs should be pursued.
Congenital heart defects; DNA methylation; LINE-1; folate; maternal biomarkers
Malaria was estimated to cause 800,000 deaths and 225 million cases worldwide in 2010. Worryingly, the first-line treatment currently relies on a single drug class called artemisinins, and there are signs that the parasite is becoming resistant to these drugs. The good news is that new technology has given us new approaches to drug discovery. New drugs generated this way are probably 10-15 years away from the clinic. Other antimalarials that may offer hope include those rehabilitated after not being used for some time, those that act as inhibitors of resistance mechanisms, those that limit infection while allowing protective immunity to develop, and those which are drugs borrowed from other disease treatments. All of these offer new hope of turning the tables on malaria. In parallel with the effort to develop vaccines that interrupt malaria transmission, drugs that target the parasite during transmission to the mosquito or during its pre-erythrocytic development in the liver, may allow us to terminate the parasite’s spread.
To investigate whether women with congenital heart defect (CHD)-affected pregnancies were more likely to have functional single nucleotide polymorphisms (SNPs) in genes encoding enzymes in folate-dependent pathways.
A population-based case-control study of 572 women with CHD-affected pregnancies and 363 control women was conducted. DNA samples were genotyped for SNPs in three genes encoding for folate pathway enzymes. Maternal lifestyle factor information was obtained using standardized interviews.
Case women were 1.5 times more likely to be obese (BMI of 30 or higher) compared to control women. Obese women carrying the MTHFR TT genotype were 4.6 times more likely to have an affected pregnancy compared to normal weight women carrying a CC genotype. Obese women carrying one or two copies of the A allele in the BHMT polymorphism were 1.8 times more likely to have a CHD-affected pregnancy than normal weight women carrying a BHMT GG genotype. Among women who smoked, those carrying a TCII CG or GG genotype were 1.8 times more likely to have an affected fetus than women who smoked and carried a CC genotype. Among women who drank alcohol, those carrying a TCII CG or GG genotype were 1.7 times more likely to have an affected fetus than women who drank and carried a CC genotype.
Results indicate that functional polymorphisms in folate-related genes increase the risk of having a fetus with CHD when maternal lifestyle factors that alter folate metabolism are present.
The majority of congenital heart defects (CHDs) are thought to result from the interaction between multiple genetic, epigenetic, environmental, and lifestyle factors. Epigenetic mechanisms are attractive targets in the study of complex diseases because they may be altered by environmental factors and dietary interventions. We conducted a population based, case-control study of genome-wide maternal DNA methylation to determine if alterations in gene-specific methylation were associated with CHDs. Using the Illumina Infinium Human Methylation27 BeadChip, we assessed maternal gene-specific methylation in over 27,000 CpG sites from DNA isolated from peripheral blood lymphocytes. Our study sample included 180 mothers with non-syndromic CHD-affected pregnancies (cases) and 187 mothers with unaffected pregnancies (controls). Using a multi-factorial statistical model, we observed differential methylation between cases and controls at multiple CpG sites, although no CpG site reached the most stringent level of genome-wide statistical significance. The majority of differentially methylated CpG sites were hypermethylated in cases and located within CpG islands. Gene Set Enrichment Analysis (GSEA) revealed that the genes of interest were enriched in multiple biological processes involved in fetal development. Associations with canonical pathways previously shown to be involved in fetal organogenesis were also observed. We present preliminary evidence that alterations in maternal DNA methylation may be associated with CHDs. Our results suggest that further studies involving maternal epigenetic patterns and CHDs are warranted. Multiple candidate processes and pathways for future study have been identified.
Alterations in maternal folate and homocysteine metabolism are associated with neural tube defects (NTDs). The role that specific micronutrients and metabolites play in the causal pathway leading to NTDs is not fully understood.
We conducted a case-control study to investigate the association between NTDs and maternal alterations in plasma micronutrients and metabolites in two metabolic pathways, the methionine remethylation and glutathione transsulfuration. Biomarkers were measured in a population-based sample of women who had NTD-affected pregnancies (n = 43) and a control group of women who had a pregnancy unaffected by a birth defect (n = 160). Plasma concentrations of folate, Vitamin B12, Vitamin B6, methionine, S-adenosylmethionine (SAM), s- adenosylhomocysteine (SAH), adenosine, homocysteine, cysteine, and reduced and oxidized glutathione were compared between cases and controls after adjusting for lifestyle and sociodemographic factors.
Women with NTD-affected pregnancies had significantly higher plasma concentrations of SAH (29.12 vs. 23.13 nmol/L, P = 0.0011), adenosine (0.323 vs. 0.255 μmol/L, P = 0.0269), homocysteine (9.40 vs. 7.56 μmol/L, P < 0.001), and oxidized glutathione (0.379 vs. 0.262μmol/L, P = 0.0001), but lower plasma SAM concentration (78.99 vs. 83.16 nmol/L, P = 0.0172) than controls. This metabolic profile is consistent with reduced methylation capacity and increased oxidative stress in women with affected pregnancies.
Increased maternal oxidative stress and decreased methylation capacity may contribute to the occurrence of NTDs. Further analysis of relevant genetic and environmental factors is required to define the basis for these observed alterations.
Neural tube defects; maternal biomarkers; folate; methionine; homocysteine; glutathione
We examined the association between maternal age and chromosome 21 nondisjunction by origin of the meiotic error. We analyzed data from two population-based, case–control studies: Atlanta Down Syndrome Project (1989–1999) and National Down Syndrome Project (2001–2004). Cases were live born infants with trisomy 21 and controls were infants without trisomy 21 delivered in the same geographical regions. We enrolled 1,215 of 1,881 eligible case families and 1,375 of 2,293 controls. We report four primary findings. First, the significant association between advanced maternal age and chromosome 21 nondisjunction was restricted to meiotic errors in the egg; the association was not observed in sperm or in post-zygotic mitotic errors. Second, advanced maternal age was significantly associated with both meiosis I (MI) and meiosis II (MII). For example, compared to mothers of controls, mothers of infants with trisomy 21 due to MI nondisjunction were 8.5 times more likely to be ≥40 years old than 20–24 years old at the birth of the index case (95% CI = 5.6–12.9). Where nondisjunction occurred in MII, mothers were 15.1 times more likely to be ≥40 years (95% CI = 8.4–27.3). Third, the ratio of MI to MII errors differed by maternal age. The ratio was lower among women <19 years of age and those ≥40 years (2.1, 2.3, respectively) and higher in the middle age group (3.6). Lastly, we found no effect of grand-maternal age on the risk for maternal nondisjunction. This study emphasizes the complex association between advanced maternal age and nondisjunction of chromosome 21 during oogenesis.
Rates of neural tube defects have decreased since folic acid fortification of the food supply in the United States. The authors’ objective was to evaluate the associations between neural tube defects and maternal folic acid intake among pregnancies conceived after fortification. This is a multicenter, case-control study that uses data from the National Birth Defects Prevention Study, 1998–2003. Logistic regression was used to compute crude and adjusted odds ratios between cases and controls assessing maternal periconceptional use of folic acid and intake of dietary folic acid. Among 180 anencephalic cases, 385 spina bifida cases, and 3, 963 controls, 21.1%, 25.2%, and 26.1%, respectively, reported periconceptional use of folic acid supplements. Periconceptional supplement use did not reduce the risk of having a pregnancy affected by a neural tube defect. Maternal intake of dietary folate was not significantly associated with neural tube defects. In this study conducted among pregnancies conceived after mandatory folic acid fortification, the authors found little evidence of an association between neural tube defects and maternal folic acid intake. A possible explanation is that folic acid fortification reduced the occurrence of folic acid-sensitive neural tube defects. Further investigation is warranted to possibly identify women who remain at increased risk of preventable neural tube defects.
folic acid; neural tube defects
Recent studies have demonstrated that human immunodeficiency virus (HIV) protease inhibitors (PIs) exert inhibitory effects on erythrocytic stages of the human-malaria parasite Plasmodium falciparum in vitro and on erythrocytic stages of the rodent-malaria parasite Plasmodium chabaudi in vivo. Although it remains unclear how HIV PIs inhibit the parasite, the effect seen on parasite development in the erythrocytic stages is potent. The effect on preerythrocytic stages has not yet been investigated. Using the rodent parasite Plasmodium berghei, we screened a panel of HIV PIs in vitro for effects on the preerythrocytic stages. Our data indicated that the HIV PIs lopinavir and saquinavir affect preerythrocytic-stage parasite development in vitro. We then evaluated the effect of HIV PIs on preerythrocytic stages in vivo using the rodent parasite Plasmodium yoelii. We found that lopinavir/ritonavir had a dose-dependent effect on liver-stage parasite development. Given that sub-Saharan Africa is where the HIV/AIDS pandemic intersects with malaria, these results merit analysis in clinical settings.
The National Down Syndrome Project (NDSP), based at Emory University in Atlanta, Georgia, represents a multi-site, population-based, case-control study with two major aims: (1) to identify molecular and epidemiological factors contributing to chromosome nondisjunction and the consequent packaging of an extra chromosome into an egg or sperm, and (2) to identify risk factors for Down syndrome-associated birth defects.
The six national sites represent approximately 11% of U.S. births. Cases were newborns with Down syndrome (trisomy 21), and controls were infants without major birth defects randomly selected from the same birth populations. Biological samples were collected from case infants and their parents, and genetic markers were typed to determine the parental origin of chromosome 21 nondisjunction. Each site interviewed parents of case and control infants addressing pregnancy, medical and family history, occupation, and exposures. Sites collected medical information on case infants.
The NDSP enrolled 907 infants as cases and 977 infants as controls (participation rates: 60.7% for cases; 56.9% for controls). Participation rates varied widely by site as did important demographic factors such as maternal age, race, and education. Nondisjunction during oogenesis accounted for 93.2% of the cases. Errors in spermatogenesis were found in 4.1%, and 2.7% were post-zygotic errors.
This exceptional compilation of questionnaire, clinical, and molecular data makes the NDSP a unique resource for ongoing studies of the etiology and phenotypic consequences of trisomy 21. The combined approach increases study power by defining subgroups of cases by the origin of nondisjunction. This report describes the design and successful implementation of the NDSP.