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Birth defects are the leading cause of infant mortality in the United States, accounting for more than 20% of all infant deaths.[1] Birth defects also contribute substantially to morbidity and […]

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. [ABSTRACT FROM AUTHOR]

BACKGROUND: Women who have had pregnancies that were affected by nonsyndromic congenital heart defects have alterations in the homocysteine-methionine pathway that may indicate increased exposure to oxidative stress or reduced antioxidant defense or both. OBJECTIVE: Our goal was to establish a maternal metabolic risk profile for nonsyndromic congenital heart defects that would enhance current preventive strategies. DESIGN: Using a case-control design, we measured biomarkers of the transsulfuration pathway in a population-based sample of women whose pregnancies were affected by congenital heart defects (331 cases) and in a control group of women (125 controls). Plasma concentrations of reduced and oxidized glutathione, vitamin B-6, homocysteine, cysteine, cysteinylglycine (CysGly), and glutamylcysteine (GluCys) were compared between cases and controls after adjustment for lifestyle and sociodemographic variables. RESULTS: After covariate adjustment, cases had significantly lower mean plasma concentrations of reduced glutathione (P < 0.0001), GluCys (P < 0.0001), and vitamin B-6 (P = 0.0023) and significantly higher mean concentrations of homocysteine (P < 0.0001) and oxidized glutathione (P < 0.0001) than did controls. CONCLUSIONS: Biomarkers of oxidative stress involved in the transsulfuration pathway were significantly higher in women with pregnancies affected by congenital heart defects than in women without such a history. Further analysis of relevant biomarkers of oxidative stress and genetic and environmental factors is required to define the basis for the observed alterations. Identifying the nature and extent of alterations in biomarkers of oxidative stress may suggest primary intervention strategies and provide clues to a greater understanding of the pathogenesis of congenital heart defects. Copyright © 2005 American Society for Clinical Nutrition [ABSTRACT FROM AUTHOR]

Conotruncal heart defects (CTDs) are heart malformations that affect the cardiac outflow tract and typically cause significant morbidity and mortality. Evidence from epidemiological studies suggests that maternal folate intake is associated with a reduced risk of heart defects, including CTD. However, it is unclear if folate-related gene variants and maternal folate intake have an interactive effect on the risk of CTDs. In this study, we performed targeted sequencing of folate-related genes on DNA from 436 case families with CTDs who are enrolled in the National Birth Defects Prevention Study and then tested for common and rare variants associated with CTD. We identified risk alleles in maternal MTHFS (OR meta = 1.34; 95% CI 1.07 to 1.67), maternal NOS2 (OR meta = 1.34; 95% CI 1.05 to 1.72), fetal MTHFS (OR meta = 1.35; 95% CI 1.09 to 1.66), and fetal TCN2 (OR meta = 1.38; 95% CI 1.12 to 1.70) that are associated with an increased risk of CTD among cases without folic acid supplementation. We detected putative de novo mutations in genes from the folate, homocysteine, and transsulfuration pathways and identified a significant association between rare variants in MGST1 and CTD risk. Results suggest that periconceptional folic acid supplementation is associated with decreased risk of CTD among individuals with susceptible genotypes.

Rapid genome sequencing in critically ill infants is increasingly identified as a crucial test for providing targeted and informed patient care. We report the outcomes of a pilot study wherein eight critically ill neonates received rapid whole genome sequencing with parental samples in an effort to establish a prompt diagnosis. Our pilot study resulted in a 37.5% diagnostic rate by whole genome sequencing alone and an overall 50% diagnostic rate for the cohort. We describe how the diagnoses led to identification of additional affected relatives and a change in management, the limitations of rapid genome sequencing, and some of the challenges with sample collection. Alongside this pilot study, our site simultaneously established a research protocol pipeline that will allow us to conduct research-based genomic testing in the cases for which a diagnosis was not reached by rapid genome sequencing or other available clinical testing. Here we describe the benefits, limitations, challenges, and potential for rapid whole genome sequencing to be incorporated into routine clinical evaluation in the neonatal period.

As SARS-CoV-2 continues to spread and evolve, detecting emerging variants early is critical for public health interventions. Inferring lineage prevalence by clinical testing is infeasible at scale, especially in areas with limited resources, participation, or testing and/or sequencing capacity, which can also introduce biases 1-3 . SARS-CoV-2 RNA concentration in wastewater successfully tracks regional infection dynamics and provides less biased abundance estimates than clinical testing 4,5 . Tracking virus genomic sequences in wastewater would improve community prevalence estimates and detect emerging variants. However, two factors limit wastewater-based genomic surveillance: low-quality sequence data and inability to estimate relative lineage abundance in mixed samples. Here we resolve these critical issues to perform a high-resolution, 295-day wastewater and clinical sequencing effort, in the controlled environment of a large university campus and the broader context of the surrounding county. We developed and deployed improved virus concentration protocols and deconvolution software that fully resolve multiple virus strains from wastewater. We detected emerging variants of concern up to 14 days earlier in wastewater samples, and identified multiple instances of virus spread not captured by clinical genomic surveillance. Our study provides a scalable solution for wastewater genomic surveillance that allows early detection of SARS-CoV-2 variants and identification of cryptic transmission., (© 2022. The Author(s).)

Newborn screening (NBS) dramatically improves outcomes in severe childhood disorders by treatment before symptom onset. In many genetic diseases, however, outcomes remain poor because NBS has lagged behind drug development. Rapid whole-genome sequencing (rWGS) is attractive for comprehensive NBS because it concomitantly examines almost all genetic diseases and is gaining acceptance for genetic disease diagnosis in ill newborns. We describe prototypic methods for scalable, parentally consented, feedback-informed NBS and diagnosis of genetic diseases by rWGS and virtual, acute management guidance (NBS-rWGS). Using established criteria and the Delphi method, we reviewed 457 genetic diseases for NBS-rWGS, retaining 388 (85%) with effective treatments. Simulated NBS-rWGS in 454,707 UK Biobank subjects with 29,865 pathogenic or likely pathogenic variants associated with 388 disorders had a true negative rate (specificity) of 99.7% following root cause analysis. In 2,208 critically ill children with suspected genetic disorders and 2,168 of their parents, simulated NBS-rWGS for 388 disorders identified 104 (87%) of 119 diagnoses previously made by rWGS and 15 findings not previously reported (NBS-rWGS negative predictive value 99.6%, true positive rate [sensitivity] 88.8%). Retrospective NBS-rWGS diagnosed 15 children with disorders that had been undetected by conventional NBS. In 43 of the 104 children, had NBS-rWGS-based interventions been started on day of life 5, the Delphi consensus was that symptoms could have been avoided completely in seven critically ill children, mostly in 21, and partially in 13. We invite groups worldwide to refine these NBS-rWGS conditions and join us to prospectively examine clinical utility and cost effectiveness., Competing Interests: Declaration of interests K.P.H., C.M.K., S.S.M., and D.T. are employees and shareholders of Illumina, Inc. G.D,A., B.M., S.L., and T.D. are employees and shareholders of Alexion Pharmaceuticals. E.F. and M.G.R. are employees and shareholders of Fabric Genomics, Inc. M.K. and S.S. are employees and shareholders of Genomenon, Inc. C.K., G.P., S.S., S.P., and A.R.W. are employees and shareholders of TileDB, Inc. S.K. is an employee and shareholder of Luna PBC, Inc. S.K. has filed a patent related to this work., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Motivation: CpG sites within the same genomic region often share similar methylation patterns and tend to be co-regulated by multiple genetic variants that may interact with one another., Results: We propose a multi-trait methylation random field (multi-MRF) method to evaluate the joint association between a set of CpG sites and a set of genetic variants. The proposed method has several advantages. First, it is a multi-trait method that allows flexible correlation structures between neighboring CpG sites (e.g. distance-based correlation). Second, it is also a multi-locus method that integrates the effect of multiple common and rare genetic variants. Third, it models the methylation traits with a beta distribution to characterize their bimodal and interval properties. Through simulations, we demonstrated that the proposed method had improved power over some existing methods under various disease scenarios. We further illustrated the proposed method via an application to a study of congenital heart defects (CHDs) with 83 cardiac tissue samples. Our results suggested that gene BACE2, a methylation quantitative trait locus (QTL) candidate, colocalized with expression QTLs in artery tibial and harbored genetic variants with nominal significant associations in two genome-wide association studies of CHD., Availability and Implementation: https://github.com/chenlyu2656/Multi-MRF., Supplementary Information: Supplementary data are available at Bioinformatics online., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

While many genetic diseases have effective treatments, they frequently progress rapidly to severe morbidity or mortality if those treatments are not implemented immediately. Since front-line physicians frequently lack familiarity with these diseases, timely molecular diagnosis may not improve outcomes. Herein we describe Genome-to-Treatment, an automated, virtual system for genetic disease diagnosis and acute management guidance. Diagnosis is achieved in 13.5 h by expedited whole genome sequencing, with superior analytic performance for structural and copy number variants. An expert panel adjudicated the indications, contraindications, efficacy, and evidence-of-efficacy of 9911 drug, device, dietary, and surgical interventions for 563 severe, childhood, genetic diseases. The 421 (75%) diseases and 1527 (15%) effective interventions retained are integrated with 13 genetic disease information resources and appended to diagnostic reports ( https://gtrx.radygenomiclab.com ). This system provided correct diagnoses in four retrospectively and two prospectively tested infants. The Genome-to-Treatment system facilitates optimal outcomes in children with rapidly progressive genetic diseases., (© 2022. The Author(s).)

DNA methylation may be regulated by genetic variants within a genomic region, referred to as methylation quantitative trait loci (mQTLs). The changes of methylation levels can further lead to alterations of gene expression, and influence the risk of various complex human diseases. Detecting mQTLs may provide insights into the underlying mechanism of how genotypic variations may influence the disease risk. In this article, we propose a methylation random field (MRF) method to detect mQTLs by testing the association between the methylation level of a CpG site and a set of genetic variants within a genomic region. The proposed MRF has two major advantages over existing approaches. First, it uses a beta distribution to characterize the bimodal and interval properties of the methylation trait at a CpG site. Second, it considers multiple common and rare genetic variants within a genomic region to identify mQTLs. Through simulations, we demonstrated that the MRF had improved power over other existing methods in detecting rare variants of relatively large effect, especially when the sample size is small. We further applied our method to a study of congenital heart defects with 83 cardiac tissue samples and identified two mQTL regions, MRPS10 and PSORS1C1, which were colocalized with expression QTL in cardiac tissue. In conclusion, the proposed MRF is a useful tool to identify novel mQTLs, especially for studies with limited sample sizes., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The emergence of the COVID-19 epidemic in the United States (U.S.) went largely undetected due to inadequate testing. New Orleans experienced one of the earliest and fastest accelerating outbreaks, coinciding with Mardi Gras. To gain insight into the emergence of SARS-CoV-2 in the U.S. and how large-scale events accelerate transmission, we sequenced SARS-CoV-2 genomes during the first wave of the COVID-19 epidemic in Louisiana. We show that SARS-CoV-2 in Louisiana had limited diversity compared to other U.S. states and that one introduction of SARS-CoV-2 led to almost all of the early transmission in Louisiana. By analyzing mobility and genomic data, we show that SARS-CoV-2 was already present in New Orleans before Mardi Gras, and the festival dramatically accelerated transmission. Our study provides an understanding of how superspreading during large-scale events played a key role during the early outbreak in the U.S. and can greatly accelerate epidemics., Competing Interests: Declaration of interests M.A.S. reports grants from the National Institutes of Health, European Research Council, and Wellcome Trust during the conduct of this research and grants and contracts from the Bill & Melinda Gates Foundation, Janssen Research and Development, Private Health Management, IQVIA, and the U.S. Department of Veterans Affairs outside the submitted work. S.L.L., R.R., and D.J.N. are employed by BioInfoexperts LLC. R.F.G. reports grants from the National Institutes of Health, the Coalition for Epidemic Preparedness Innovations, the Burroughs Wellcome Fund, the Wellcome Trust, the Center for Disease Prevention and Control, and the European & Developing Countries Clinical Trials Partnership. He is the co-founder and Chief Scientific Advisor of Zalgen Labs, a biotechnology company developing countermeasures to emerging viruses, including SARS-CoV-2. K.G.A. has received consulting fees and compensated expert testimony on SARS-CoV-2 and the COVID-19 pandemic., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Most congenital heart defects (CHDs) result from complex interactions among genetic susceptibilities, epigenetic modifications, and maternal environmental exposures. Characterizing the complex relationship between genetic, epigenetic, and transcriptomic variation will enhance our understanding of pathogenesis in this important type of congenital disorder. We investigated cis-acting effects of genetic single nucleotide polymorphisms (SNPs) on local DNA methylation patterns within 83 cardiac tissue samples and prioritized their contributions to CHD risk by leveraging results of CHD genome-wide association studies (GWAS) and their effects on cardiac gene expression., Results: We identified 13,901 potential methylation quantitative trait loci (mQTLs) with a false discovery threshold of 5%. Further co-localization analyses and Mendelian randomization indicated that genetic variants near the HLA-DRB6 gene on chromosome 6 may contribute to CHD risk by regulating the methylation status of nearby CpG sites. Additional SNPs in genomic regions on chromosome 10 (TNKS2-AS1 gene) and chromosome 14 (LINC01629 gene) may simultaneously influence epigenetic and transcriptomic variations within cardiac tissues., Conclusions: Our results support the hypothesis that genetic variants may influence the risk of CHDs through regulating the changes of DNA methylation and gene expression. Our results can serve as an important source of information that can be integrated with other genetic studies of heart diseases, especially CHDs.

Previous research on risk factors for obstructive heart defects (OHDs) focused on maternal and infant genetic variants, prenatal environmental exposures, and their potential interaction effects. Less is known about the role of paternal genetic variants or environmental exposures and risk of OHDs. We examined parent-of-origin effects in transmission of alleles in the folate, homocysteine, or transsulfuration pathway genes on OHD occurrence in offspring. We used data on 569 families of liveborn infants with OHDs born between October 1997 and August 2008 from the National Birth Defects Prevention Study to conduct a family-based case-only study. Maternal, paternal, and infant DNA were genotyped using an Illumina Golden Gate custom single nucleotide polymorphism (SNP) panel. Relative risks (RR), 95% confidence interval (CI), and likelihood ratio tests from log-linear models were used to estimate the parent-of-origin effect of 877 SNPs in 60 candidate genes in the folate, homocysteine, and transsulfuration pathways on the risk of OHDs. Bonferroni correction was applied for multiple testing. We identified 3 SNPs in the transsulfuration pathway and 1 SNP in the folate pathway that were statistically significant after Bonferroni correction. Among infants who inherited paternally-derived copies of the G allele for rs6812588 in the RFC1 gene, the G allele for rs1762430 in the MGMT gene, and the A allele for rs9296695 and rs4712023 in the GSTA3 gene, RRs for OHD were 0.11 (95% CI: 0.04, 0.29, P = 9.16x10-7), 0.30 (95% CI: 0.17, 0.53, P = 9.80x10-6), 0.34 (95% CI: 0.20, 0.57, P = 2.28x10-5), and 0.34 (95% CI: 0.20, 0.58, P = 3.77x10-5), respectively, compared to infants who inherited maternally-derived copies of the same alleles. We observed statistically significant decreased risk of OHDs among infants who inherited paternal gene variants involved in folate and transsulfuration pathways., Competing Interests: The authors have declared that no competing interests exist.

Hypospadias is a common birth defect where the urethral opening forms on the ventral side of the penis. We performed integrative methylomic, genomic, and transcriptomic analyses to characterize sites of DNA methylation that influence genital development. In case-control and case-only epigenome-wide association studies (EWAS) of preputial tissue we identified 25 CpGs associated with hypospadias characteristics and used one-sample two stage least squares Mendelian randomization (2SLS MR) to show a causal relationship for 21 of the CpGs. The largest difference was 15.7% lower beta-value at cg14436889 among hypospadias cases than controls (EWAS P = 5.4e-7) and is likely causal (2SLS MR P = 9.8e-15). Integrative annotation using two-sample Mendelian randomization of these methylation regions highlight potentially causal roles of genes involved in germ layer differentiation (WDHD1, DNM1L, TULP3), beta-catenin signaling (PKP2, UBE2R2, TNKS), androgens (CYP4A11, CYP4A22, CYP4B1, CYP4X1, CYP4Z2P, EPHX1, CD33/SIGLEC3, SIGLEC5, SIGLEC7, KLK5, KLK7, KLK10, KLK13, KLK14), and reproductive traits (ACAA1, PLCD1, EFCAB4B, GMCL1, MKRN2, DNM1L, TEAD4, TSPAN9, KLK family). This study identified CpGs that remained differentially methylated after urogenital development and used the most relevant tissue sample available to study hypospadias. We identified multiple methylation sites and candidate genes that can be further evaluated for their roles in regulating urogenital development.

Background: The development of conotruncal heart defects (CTDs) involves a complex relationship among genetic variants and maternal lifestyle factors. In this article, we focused on the interactions between 13 candidate genes within folate, homocysteine, and transsulfuration pathways for potential association with CTD risk., Methods: Targeted sequencing was used for 328 case-parental triads enrolled in the National Birth Defects Prevention Study (NBDPS). To evaluate the interaction of two genes, we applied a conditional logistic regression model for all possible SNP pairs within two respective genes by contrasting the affected infants with their pseudo-controls. The findings were replicated in an independent sample of 86 NBDPS case-parental triads genotyped by DNA microarrays. The results of two studies were further integrated by a fixed-effect meta-analysis., Results: One SNP pair (i.e., rs4764267 and rs6556883) located in gene MGST1 and GLRX, respectively, was found to be associated with CTD risk after multiple testing adjustment using simpleM, a modified Bonferroni correction approach (nominal p-value of 4.62e-06; adjusted p-value of .04). Another SNP pair (i.e., rs11892646 and rs56219526) located in gene DNMT3A and MTRR, respectively, achieved marginal significance after multiple testing adjustment (adjusted p-value of .06)., Conclusion: Further studies with larger sample sizes are needed to confirm and elucidate these potential interactions., (© 2019 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Gardenia blue is widely used in Eastern Asia as a natural food colorant. To evaluate the genotoxic potential of gardenia blue, as well as genipin, the natural starting material from which it is produced, a GLP-compliant test battery was conducted according to OECD guidelines. No evidence of mutagenicity of gardenia blue was detected in a 5-strain bacterial reverse mutation assay, with or without metabolic activation; an equivocal response for genipin occurred in S. typhimurium TA97a without metabolic activation. In in vitro micronucleus and chromosome aberration assays, genipin tested positive under some test conditions; however, gardenia blue tested negative in both assays. In combined micronucleus/comet assays conducted in male and female B6C3F1 mice, exposure to genipin at doses reaching maximal toxicity (74 and 222 mg/kg bw/day for males and females, respectively) or gardenia blue tested up to the limit dose (2000 mg/kg bw/day) did not induce micronuclei in peripheral blood or DNA damage in several examined tissues. Modified ("reverse") comet assays showed no evidence of DNA crosslinking potential of either genipin, known to form crosslinks with other macromolecules, or gardenia blue. Our results indicate that consumption of gardenia blue in food products does not pose a significant genotoxic concern for humans., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Obstructive heart defects (OHDs) are a major health concern worldwide. The platelet-derived growth factor (PDGF) genes are known to have regulatory functions that are essential for proper heart development. In a zebrafish model, Pdgfra was further demonstrated to interact with ethanol during craniofacial development. In this article, we investigated interactions between variants in PDGF genes and periconceptional alcohol exposure on the risk of OHDs by applying log-linear models to 806 OHD case and 995 control families enrolled in the National Birth Defects Prevention Study. The interactions between four variants in PDGFA and maternal binge drinking reached a nominal significance level. The maternal T allele of rs869978 was estimated to increase OHD risk among women who binge drink, while infant genotypes of rs2291591, rs2228230, rs1547904, and rs869978 may reduce the risk. Although none of these associations remain statistically significant after multiple testing adjustment and the estimated maternal effect may be influenced by unknown confounding factors, such as maternal smoking, these findings are consistent with previous animal studies supporting potential interactions between the PDGFRA gene and maternal alcohol exposure. Replication studies with larger sample sizes are needed to further elucidate this potential interplay and its influence on OHD risks.

When a comet assay, an increasingly popular in vivo genotoxicity test, shows a positive test result, interpretation of that response requires ruling out any confounding tissue site toxicity. Since the comet assay typically uses only two or three daily doses of test agent, precursor tissue changes indicative of toxicity may be easily overlooked. Using case examples for two flavoring agents, perillaldehyde and 4,5-epoxydec-2( trans )-enal, we highlight the role of pathology peer review in verifying precursor tissue changes indicative of tissue site toxicity, thereby increasing confidence in final interpretation of comet assay results. Given global deliberation regarding safety assessment of compounds entering the marketplace, we recommend consideration of pathology peer review for equivocal and positive comet assays so that interpretations are universally consistent.

Quercetin and its glycosides possess potential benefits to human health. Several flavonols are available to consumers as dietary supplements, promoted as anti-oxidants; however, incorporation of natural quercetin glycosides into food and beverage products has been limited by poor miscibility in water. Enzymatic conjugation of multiple glucose moieties to isoquercitrin to produce alpha-glycosyl isoquercitrin (AGIQ) enhances solubility and bioavailability. AGIQ is used in Japan as a food additive and has been granted generally recognized as safe (GRAS) status. However, although substantial genotoxicity data exist for quercetin, there is very little available data for AGIQ and isoquercitrin. To support expanded global marketing of food products containing AGIQ, comprehensive testing of genotoxic potential of AGIQ and isoquercitrin was conducted according to current regulatory test guidelines. Both chemicals tested positive in bacterial reverse mutation assays, and exposure to isoquercitrin resulted in chromosomal aberrations in CHO-WBL cells. All other in vitro mammalian micronucleus and chromosomal aberration assays, micronucleus and comet assays in male and female B6C3F1 mice and Sprague Dawley rats, and Muta™ Mouse mutation assays evaluating multiple potential target tissues, were negative for both chemicals. These results supplement existing toxicity data to further support the safe use of AGIQ in food and beverage products., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Magnesium stearate is widely used in the production of dietary supplement and pharmaceutical tablets, capsules and powders as well as many food products, including a variety of confectionery, spices and baking ingredients. Although considered to have a safe toxicity profile, there is no available information regarding its potential to induce genetic toxicity. To aid safety assessment efforts, magnesium sulfate was evaluated in a battery of tests including a bacterial reverse mutation assay, an in vitro chromosome aberration assay, and an in vivo erythrocyte micronucleus assay. Magnesium stearate did not produce a positive response in any of the five bacterial strains tested, in the absence or presence of metabolic activation. Similarly, exposure to magnesium stearate did not lead to chromosomal aberrations in CHL/IU Chinese hamster lung fibroblasts, with or without metabolic activation, or induce micronuclei in the bone marrow of male CD-1 mice. These studies have been used by the Japanese government and the Joint FAO/WHO Expert Committee on Food Additives in their respective safety assessments of magnesium stearate. These data indicate a lack of genotoxic risk posed by magnesium stearate consumed at current estimated dietary exposures. However, health effects of cumulative exposure to magnesium via multiple sources present in food additives may be of concern and warrant further evaluation.

Acrylamide (AA) exposure in 2-year cancer bioassays leads to thyroid, but not liver, adenomas and adenocarcinomas in rats. Hypothesized modes of action (MOAs) include genotoxicity/mutagenicity, or thyroid hormone dysregulation. To examine the plausibility of these two or any alternative MOAs, RNA-sequencing was performed on the thyroids and livers of AA-exposed rats, in parallel with measurement of genotoxicity (blood micronucleus and Pig-a mutant frequency) and serum thyroid hormone levels, following the exposure of male Fischer 344/DuCrl rats to 0.0, 0.5, 1.5, 3.0, 6.0, or 12.0 mg AA/kg bw-day in drinking water for 5, 15, or 31 days. Differentially expressed genes in both tissues provided marginal support for hormonal and genotoxic MOAs, which was consistent with negative/equivocal genotoxicity assay and marginal changes in thyroid hormone levels. Instead, there was a pronounced effect on calcium signaling/cytoskeletal genes in the thyroid. Benchmark dose modeling of RNA-sequencing data for the calcium signaling pathway suggests a point of departure (POD) of 0.68 mg/kg bw-day, which is consistent with a POD of 0.82 mg/kg bw-day derived from the thyroid 2-year cancer bioassay data. Overall, this study suggests a novel MOA for AA-induced thyroid carcinogenicity in male rats centered around perturbation of calcium signaling., (Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.)

Acrylamide is used in many industrial processes and is present in a variety of fried and baked foods. In rodent carcinogenicity assays, acrylamide exposure leads to tumour formation at doses lower than those demonstrated to induce genotoxic damage. We evaluated the potential of acrylamide to induce structural DNA damage and gene mutations in rodents using highly sensitive flow cytometric analysis of micronucleus and Pig-a mutant frequencies, respectively. Male F344 rats and B6C3F1 mice were administered acrylamide in drinking water for 30 days at doses spanning and exceeding the range of acrylamide exposure tested in cancer bioassays-top dose of 12.0 and 24.0mg/kg/day in mice and in rats, respectively. A positive control, N-ethyl-N-nitrosourea, was administered at the beginning and end of the study to meet the expression time for the two DNA damage phenotypes. The results of the micronucleus and Pig-a assays were negative and equivocal, respectively, for male rats exposed to acrylamide at the concentrations tested. In contrast, acrylamide induced a dose-dependent increase in micronucleus formation but tested negative in the Pig-a assay in mice. Higher plasma concentrations of glycidamide in mice than rats are hypothesized to explain, at least in part, the differences in the response. Benchmark dose modelling indicates that structural DNA damage as opposed to point mutations is most relevant to the genotoxic mode of action of acrylamide-induced carcinogenicity. Moreover, the lack of genotoxicity detected at <6.0mg/kg/day is consistent with the notion that non-genotoxic mechanisms contribute to acrylamide-induced carcinogenicity in rodents., (© The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Perillaldehyde, a natural monocyclic terpenoid found most abundantly in the herb perilla, has a long history of use as a flavouring ingredient to add spiciness and citrus taste to foods. Previously, it was judged to be safe by several international expert panels. To confirm the safety of flavourings placed on the European Union list of flavourings, perillaldehyde was selected by the European Food Safety Authority as a representative of a subgroup of alicyclic aldehyde flavouring substances to be evaluated for genotoxic potential. Perillaldehyde was tested in a bacterial reverse mutation assay, an in vitro micronucleus assay in human lymphocytes, an HPRT assay in mouse lymphoma cells, and a micronucleus/comet assay in Han Wistar rats. In contrast to previously published results, perillaldehyde induced mutation in Salmonella typhimurium strain TA98 in the absence of metabolic activation. The comet assay was negative for duodenum and weakly positive for liver but only at a hepatotoxic dose of perillaldehyde. All other genotoxicity assays were negative. These data do not provide an indication of any genotoxic potential for perillaldehyde, and they provide the primary basis for recent scientific opinions regarding perillaldehyde genotoxicity announced by several international organizations responsible for safety assessment of food additives and flavourings., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Introduction: Microbiologically derived cyclodextrin glucanotransferase (CGTase) is used commercially as a processing agent in manufacture of food, pharmaceuticals, and cosmetics. Its toxic potential was evaluated in anticipation of use in the production of alpha -glycosyl isoquercitrin, a water-soluble form of quercetin., Methods: Following OECD guidelines, CGTase, produced by Bacillus pseudalcaliphilus DK-1139, was evaluated in a genotoxicity battery consisting of a bacterial reverse mutation assay, an in vitro micronucleus (MN) assay and MN and comet assays using B6C3F1 male and female mice. These same genotoxicity assays were also conducted for sodium sulfate, a contaminant of CGTase preparation. In a 90-day Sprague Dawley rat toxicity study, CGTase was administered by gavage in water at daily doses of 0, 250, 500, and 1000 mg/kg/day., Results: CGTase did not induce mutations with or without metabolic activation in the bacterial reverse mutation assay. Formation of micronuclei was not induced in either in vitro or in vivo MN assays with or without metabolic activation. No induction of DNA damage was detected in male or female mouse liver, stomach, or duodenum in the comet assay. Sodium sulfate also tested negative in these same genotoxicity assays. In the 90-day repeated dose rat study there were no treatment-related adverse clinical or pathological findings., Conclusion: The genotoxicity assays and repeated dose toxicity study support the safe use of CGTase in production of alpha -glycosyl isoquercitrin.

Congenital heart defects (CHDs) develop through a complex interplay between genetic variants, epigenetic modifications, and maternal environmental exposures. Genetic studies of CHDs have commonly tested single genetic variants for association with CHDs. Less attention has been given to complex gene-by-gene and gene-by-environment interactions. In this study, we applied a recently developed likelihood-ratio Mann-Whitney (LRMW) method to detect joint actions among maternal variants, fetal variants, and maternal environmental exposures, allowing for high-order statistical interactions. All subjects are participants from the National Birth Defect Prevention Study, including 623 mother-offspring pairs with CHD-affected pregnancies and 875 mother-offspring pairs with unaffected pregnancies. Each individual has 872 single nucleotide polymorphisms encoding for critical enzymes in the homocysteine, folate, and trans-sulfuration pathways. By using the LRMW method, three variants (fetal rs625879, maternal rs2169650, and maternal rs8177441) were identified with a joint association to CHD risk (nominal P-value = 1.13e-07). These three variants are located within genes BHMT2, GSTP1, and GPX3, respectively. Further examination indicated that maternal SNP rs2169650 may interact with both fetal SNP rs625879 and maternal SNP rs8177441. Our findings suggest that the risk of CHD may be influenced by both the intragenerational interaction within the maternal genome and the intergenerational interaction between maternal and fetal genomes., (© 2015 John Wiley & Sons Ltd/University College London.)

Myricitrin, a flavonoid extracted from the fruit, leaves, and bark of Chinese bayberry (Myrica rubra SIEBOLD), is currently used as a flavor modifier in snack foods, dairy products, and beverages in Japan. Myricitrin is converted to myricetin by intestinal microflora; myricetin also occurs ubiquitously in plants and is consumed in fruits, vegetables, and beverages. The genotoxic potential of myricitrin and myricetin was evaluated in anticipation of worldwide marketing of food products containing myricitrin. In a bacterial reverse mutation assay, myricetin tested positive for frameshift mutations under metabolic activation conditions whereas myricitrin tested negative for mutagenic potential. Both myricitrin and myricetin induced micronuclei formation in human TK6 lymphoblastoid cells under conditions lacking metabolic activation; however, the negative response observed in the presence of metabolic activation suggests that rat liver S9 homogenate may detoxify reactive metabolites of these chemicals in mammalian cells. In 3-day combined micronucleus/Comet assays using male and female B6C3F1 mice, no induction of micronuclei was observed in peripheral blood, or conclusive evidence of damage detected in the liver, glandular stomach, or duodenum following exposure to myricitrin or myricetin. Our studies did not reveal evidence of genotoxic potential of myricitrin in vivo, supporting its safe use in food and beverages., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Objective: The aim of this study is to investigate the association between birth defects (BDs), prematurity and small-for-gestational age (SGA) in a population-based sample., Study Design: Participants were singleton live births enrolled in the National Birth Defects Prevention Study, including 18 737 case infants with one or more BD and 7999 controls. Logistic regression models to evaluate associations between BDs, prematurity and fetal growth were computed while adjusting for covariates., Result: Cases were significantly more likely to be born prematurely than controls, particularly at 24 to 28 weeks of gestation. The highest odds ratios for preterm birth were found for intestinal atresia, anencephaly, gastroschisis and esophageal atresia. Infants with BDs were also significantly more likely to be SGA than controls (17.2 and 7.8%)., Conclusion: Infants with BDs are more likely than controls to be born prematurely and SGA. Findings from this study present additional evidence demonstrating a complex interaction between the development of BDs, prematurity and intrauterine growth.

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.

The development of congenital heart defects (CHDs) involves a complex interplay between genetic variants, epigenetic variants, and environmental exposures. Previous studies have suggested that susceptibility to CHDs is associated with maternal genotypes, fetal genotypes, and maternal-fetal genotype (MFG) interactions. We conducted a haplotype-based genetic association study of obstructive heart defects (OHDs), aiming to detect the genetic effects of 877 SNPs involved in the homocysteine, folate, and transsulfuration pathways. Genotypes were available for 285 mother-offspring pairs with OHD-affected pregnancies and 868 mother-offspring pairs with unaffected pregnancies. A penalized logistic regression model was applied with an adaptive least absolute shrinkage and selection operator (lasso), which dissects the maternal effect, fetal effect, and MFG interaction effects associated with OHDs. By examining the association between 140 haplotype blocks, we identified 9 blocks that are potentially associated with OHD occurrence. Four haplotype blocks, located in genes MGMT, MTHFS, CBS, and DNMT3L, were statistically significant using a Bayesian false-discovery probability threshold of 0.8. Two blocks in MGMT and MTHFS appear to have significant fetal effects, while the CBS and DNMT3L genes may have significant MFG interaction effects.

The epithelial sodium channel (ENaC) is responsible for Na(+) and fluid absorption across colon, kidney, and airway epithelia. Short palate lung and nasal epithelial clone 1 (SPLUNC1) is a secreted, innate defense protein and an autocrine inhibitor of ENaC that is highly expressed in airway epithelia. While SPLUNC1 has a bactericidal permeability-increasing protein (BPI)-type structure, its NH2-terminal region lacks structure. Here we found that an 18 amino acid peptide, S18, which corresponded to residues G22-A39 of the SPLUNC1 NH2 terminus inhibited ENaC activity to a similar degree as full-length SPLUNC1 (∼2.5 fold), while SPLUNC1 protein lacking this region was without effect. S18 did not inhibit the structurally related acid-sensing ion channels, indicating specificity for ENaC. However, S18 preferentially bound to the βENaC subunit in a glycosylation-dependent manner. ENaC hyperactivity is contributory to cystic fibrosis (CF) lung disease. Unlike control, CF human bronchial epithelial cultures (HBECs) where airway surface liquid (ASL) height was abnormally low (4.2 ± 0.6 μm), addition of S18 prevented ENaC-led ASL hyperabsorption and maintained CF ASL height at 7.9 ± 0.6 μm, even in the presence of neutrophil elastase, which is comparable to heights seen in normal HBECs. Our data also indicate that the ENaC inhibitory domain of SPLUNC1 may be cleaved away from the main molecule by neutrophil elastase, suggesting that it may still be active during inflammation or neutrophilia. Furthermore, the robust inhibition of ENaC by the S18 peptide suggests that this peptide may be suitable for treating CF lung disease.

The ability to maintain proper airway surface liquid (ASL) volume homeostasis is vital for mucus hydration and clearance, which are essential aspects of the mammalian lung's innate defense system. In cystic fibrosis (CF), one of the most common life-threatening genetic disorders, ASL dehydration leads to mucus accumulation and chronic infection. In normal airways, the secreted protein short palate lung and nasal epithelial clone 1 (SPLUNC1) effectively inhibits epithelial Na(+) channel (ENaC)-dependent Na(+) absorption and preserves ASL volume. In CF airways, it has been hypothesized that increased ENaC-dependent Na(+) absorption contributes to ASL depletion, and hence increased disease. However, this theory is controversial, and the mechanism for abnormal ENaC regulation in CF airways has remained elusive. Here, we show that SPLUNC1 is a pH-sensitive regulator of ENaC and is unable to inhibit ENaC in the acidic CF airway environment. Alkalinization of CF airway cultures prevented CF ASL hyperabsorption, and this effect was abolished when SPLUNC1 was stably knocked down. Accordingly, we resolved the crystal structure of SPLUNC1 to 2.8 Å. Notably, this structure revealed two pH-sensitive salt bridges that, when removed, rendered SPLUNC1 pH-insensitive and able to regulate ASL volume in acidic ASL. Thus, we conclude that ENaC hyperactivity is secondary to reduced CF ASL pH. Together, these data provide molecular insights into the mucosal dehydration associated with a range of pulmonary diseases, including CF, and suggest that future therapy be directed toward alkalinizing the pH of CF airways.

Airway epithelia absorb Na+ through the epithelial Na+ channel (ENaC) and secrete Cl- through the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. This balance maintains sufficient airway surface liquid hydration to permit efficient mucus clearance, which is needed to maintain sterility of the lung. Cystic fibrosis (CF) is a common autosomal recessive inherited disease caused by mutations in the CFTR gene that lead to the reduction or elimination of the CFTR protein. CF is a multi-organ disease that affects epithelia lining the intestines, lungs, pancreas, sweat ducts and vas deferens, among others. CF lungs are characterized by viscous, dehydrated mucus, persistent neutrophilia and chronic infections. ENaC is negatively regulated by CFTR and, in patients with CF, the absence of CFTR results in a double hit of reduced Cl-/HCO3- and H2O secretion as well as ENaC hyperactivity and increased Na+ and H2O absorption. Together, these effects are hypothesized to trigger mucus dehydration, resulting in a failure to clear mucus. Rehydrating CF mucus has become a recent clinical focus and yields important end-points for clinical trials. However, while ENaC hyperactivity in CF airways has been detected in vivo and in vitro, recent data have brought the role of ENaC in CF lung disease pathogenesis into question. This review will focus on our current understanding of the contribution of ENaC to CF pathogenesis.

Purpose: Advanced maternal age and altered recombination are known risk factors for Down syndrome cases due to maternal nondisjunction 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., Methods: 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., Results: 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)., Conclusion: We detected a significant association between low maternal socioeconomic status and meiosis II chromosome 21 nondisjunction. 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.

The breast cancer 1 (BRCA1) protein is a tumor suppressor playing roles in DNA repair and cell cycle regulation. Studies of DNA repair functions of BRCA1 have focused on double-strand break (DSB) repair pathways and have recently included base excision repair (BER). However, the function of BRCA1 in BER is not well defined. Here, we examined a BRCA1 role in BER, first in relation to alkylating agent (MMS) treatment of cells and the BER enzyme DNA polymerase β (pol β). MMS treatment of BRCA1 negative human ovarian and chicken DT40 cells revealed hypersensitivity, and the combined gene deletion of BRCA1 and pol β in DT40 cells was consistent with these factors acting in the same repair pathway, possibly BER. Using cell extracts and purified proteins, BRCA1 and pol β were found to interact in immunoprecipitation assays, yet in vivo and in vitro assays for a BER role of BRCA1 were negative. An alternate approach with the human cells of immunofluorescence imaging and laser-induced DNA damage revealed negligible BRCA1 recruitment during the first 60 s after irradiation, the period typical of recruitment of pol β and other BER factors. Instead, 15 min after irradiation, BRCA1 recruitment was strong and there was γ-H2AX co-localization, consistent with DSBs and repair. The rapid recruitment of pol β was similar in BRCA1 positive and negative cells. However, a fraction of pol β initially recruited remained associated with damage sites much longer in BRCA1 positive than negative cells. Interestingly, pol β expression was required for BRCA1 recruitment, suggesting a partnership between these repair factors in DSB repair.

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 U.S. 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.

Background: 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., Methods: 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., Results: 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)., Conclusions: 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).

Gum ghatti is a food additive in some parts of the world, serving as an emulsifier, a stabilizer, and a thickening agent. To evaluate its genotoxic potential, we conducted Good Laboratory Practice compliant in vitro and in vivo studies in accordance with the Organisation for Economic Co-operation and Development (OECD) guidelines. No evidence of toxicity or mutagenicity was detected in a bacterial reverse mutation assay using five tester strains evaluating gum ghatti at up to 6 mg/plate, with or without metabolic activation. Gum ghatti also did not induce chromosome structural damage in a chromosome aberration assay using Chinese hamster ovary cells. To assess the ability to induce DNA damage in rodents, a combined micronucleus/Comet assay was conducted in male B6C3F1 mice. Gum ghatti was administered at 1000, 1500, and 2000 mg/kg/day by gavage once daily for 4 days and samples were collected 4h after the final dosing. No effect of gum ghatti was measured on micronucleated reticulocyte (MN-RET) frequency in peripheral blood, or DNA damage in blood leukocytes or liver as assessed by the Comet assay. Our results show no evidence of genotoxic potential of gum ghatti administered up to the maximum concentrations recommended by OECD guidelines., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

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.

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 >or=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 >or=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 >or=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.

Background: With >1% of US births occurring following use of assisted reproductive technology (ART), it is critical to examine whether ART is associated with birth defects., Methods: We analyzed data from the National Birth Defects Prevention Study, a population-based, multicenter, case-control study of birth defects. We included mothers of fetuses or live-born infants with a major birth defect (case infants) and mothers who had live-born infants who did not have a major birth defect (control infants), delivered during the period October 1997-December 2003. We compared mothers who reported ART use (IVF or ICSI) with those who had unassisted conceptions. Multiple logistic regression was used to adjust for the following confounders: maternal race/ethnicity, maternal age, smoking and parity; we stratified by plurality., Results: ART was reported by 1.1% of all control mothers, and by 4.5% of control mothers 35 years or older. Among singleton births, ART was associated with septal heart defects (adjusted odds ratio [aOR] = 2.1, 95% confidence intervals [CI] 1.1-4.0), cleft lip with or without cleft palate (aOR = 2.4, 95% CI 1.2-5.1), esophageal atresia (aOR = 4.5, 95% CI 1.9-10.5) and anorectal atresia (aOR = 3.7, 95% CI 1.5-9.1). Among multiple births, ART was not significantly associated with any of the birth defects studied., Conclusions: These findings suggest that some birth defects occur more often among infants conceived with ART. Although the mechanism is not clear, couples considering ART should be informed of all potential risks and benefits.

Purpose: The population-based National Down Syndrome Project combined epidemiological and molecular methods to study congenital heart defects in Down syndrome., Methods: Between 2000 and 2004, six sites collected DNA, clinical, and epidemiological information on parents and infants. We used logistic regression to examine factors associated with the most common Down syndrome-associated heart defects., Results: Of 1469 eligible infants, major cardiac defects were present in 44%; atrioventricular septal defect (39%), secundum atrial septal defect (42%), ventricular septal defect (43%), and tetralogy of Fallot (6%). Atrioventricular septal defects showed the most significant sex and ethnic differences with twice as many affected females (odds ratio, 1.93; 95% confidence interval, 1.40-2.67) and, compared with whites, twice as many blacks (odds ratio, 2.06; 95% confidence interval, 1.32-3.21) and half as many Hispanics (odds ratio, 0.48; 95% confidence interval, 0.30-0.77). No associations were found with origin of the nondisjunction error or with the presence of gastrointestinal defects., Conclusions: Sex and ethnic differences exist for atrioventricular septal defects in Down syndrome. Identification of genetic and environmental risk factors associated with these differences is essential to our understanding of the etiology of congenital heart defects.

Objective: 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., Methods: 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., Results: 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., Conclusions: 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

Elevated expression of ornithine decarboxylase (ODC) and increased synthesis of polyamines are hallmarks of epithelial tumorigenesis. The skin and tumors of K6/ODC and ODC/Ras transgenic mice, in which overexpression of ODC has been targeted to hair follicles, were found to exhibit intrinsically high histone acetyltransferase (HAT) activity. We identified Tip60 as a candidate enzyme for contributing significantly to this abnormally high HAT activity. Compared with normal littermate controls, the levels of Tip60 protein and an alternative splice variant Tip53 were found to be greater in K6/ODC mouse skin. Furthermore, skin tumors that spontaneously develop in ODC/Ras bigenic mice typically have substantially more Tip60 protein than adjacent non-tumor-bearing skin and exhibit a unique pattern of Tip60 size variants and chemically modified protein isoforms. Steady-state Tip60 and Tip53 mRNA levels were not affected in ODC-overexpressing skin and tumors, implying novel posttranscriptional regulation by polyamines. Given the diverse roles of Tip60, the overabundance of Tip60 protein is predicted to have biological consequences. Compared with normal littermate skin, we detected altered association of Tip60 with E2F1 and a subset of newly identified Tip60-interacting transcription factors in ODC transgenic mouse skin and tumors. E2F1 was shown to be bound in greater amounts to up-regulated target genes in ODC-overexpressing skin. Thus, up-regulation of Tip60 protein, influencing the expression of Tip60-regulated genes, could play a contributing role in polyamine-mediated tumor promotion. (

Cost-effectiveness evaluations of interventions to prevent or treat spina bifida require quality of life information measured as preference scores. Preference scores of caregivers also may be relevant. This study tested whether the preference scores of children with spina bifida and their caregivers would decrease as disability in the child increased. Families of children aged 0-17 with spina bifida (N = 98) were identified using a birth defect surveillance system in the state of Arkansas. Primary caregivers of children with spina bifida identified other families with an unaffected child (N = 49). Preference scores for child health states were determined using the Health Utilities Index--Mark 2 (HUI2). Caregiver preference scores were determined using the Quality of Well-Being (QWB) scale. Children with spina bifida were categorized into three disability levels according to the location of the child's lesion. Mean preference scores declined for both affected children and the primary caregiver as disability in the child increased. In multivariate analysis, the preference score of the child was a significant and positive predictor of the primary caregiver's preference score. A more modest association was found for caregiver health preference scores by lesion location. The findings can inform cost-effectiveness evaluations of interventions to treat or prevent spina bifida.