A Common Variant in Methionine Synthase Reductase Combined with Low Cobalamin (Vitamin B12) Increases Risk for Spina Bifida

Impairment of folate and cobalamin (vitamin B(12)) metabolism has been observed in families with neural tube defects (NTDs). Genetic variants of enzymes in the homocysteine remethylation pathway might act as predisposing factors contributing to NTD risk. The first polymorphism linked to increased NTD risk was the 677C-->T mutation in methylenetetrahydrofolate reductase (MTHFR). We now report a polymorphism in methionine synthase reductase (MTRR), the enzyme that activates cobalamin-dependent methionine synthase. This polymorphorism, 66A-->G (I22M), has an allele frequency of 0.51 and increases NTD risk when cobalamin status is low or when the MTHFR mutant genotype is present. Genotypes and cobalamin status were assessed in 56 patients with spina bifida, 58 mothers of patients, 97 control children, and 89 mothers of controls. Cases and case mothers were almost twice as likely to possess the homozygous mutant genotype when compared to controls, but this difference was not statistically significant. However, when combined with low levels of cobalamin, the risk for mothers increased nearly five times (odds ratio (OR) = 4.8, 95% CI 1.5-15.8); the OR for children with this combination was 2.5 (95% CI 0.63-9.7). In the presence of combined MTHFR and MTRR homozygous mutant genotypes, children and mothers had a fourfold and threefold increase in risk, respectively (OR = 4.1, 95% CI 1.0-16.4; and OR = 2.9, 95% CI 0.58-14.8). This study provides the first genetic link between vitamin B(12) deficiency and NTDs and supports the multifactorial origins of these common birth defects. Investigation of this polymorphism in other disorders associated with altered homocysteine metabolism, such as vascular disease, is clearly warranted.     

Maternal and neonatal outcomes in dichorionic twin pregnancies following IVF treatment: a hospital-based comparative study

Aim: To compare maternal, and neonatal outcomes in IVF/ICSI and spontaneously conceived dichorionic twin pregnancy. Method: We collected data regarding dichorionic twin pregnancies following in vitro fertilization/ intracytoplasmic sperm injection (IVF/ICSI, n=162) with the transfer of fresh embryos as well as data regarding spontaneously conceived pregnancies (n=213) delivered after 28 weeks of gestation at the Department of Obstetrics and Gynecology, Renmin Hospital in Wuhan in the years of 2010-2013. We then compared maternal and neonatal outcomes between IVF/ICSI and spontaneous dichorionic twin pregnancies, with a subgroup analysis separating traditional IVF from ICSI pregnancies. Odds ratios (OR) for associations between IVF/ICSI and pregnancy outcomes were adjusted for maternal factors. Results: The mean maternal age and the percentage of primiparous women were significantly higher in the IVF/ICSI group. Multivariate analysis revealed that maternal outcomes were comparable in both groups with/without adjustment for maternal age and parity. However, IVF/ICSI twins were less likely to have birth weight discordance than those spontaneously conceived (unadjusted OR=0.526, 95% CI 0.297-0.932; adjusted OR=0.486, 95% CI 0.255-0.856). In subgroup analyses, these associations were confirmed in the IVF (adjusted OR=0.496, 95% CI 0.265-0.926), but not in the ICSI group (adjusted OR=0.500, 95% CI 0.139-1.807). Conclusion: IVF/ICSI treatment was not a risk factor for adverse maternal neonatal outcomes, but the risk for birth weight discordance is lower among IVF/ICSI twins.     

Risk factors associated with neural tube defects: exposure during the first trimester of gestation

The neural tube defects (NTD) are a group of malformations of multifactorial etiology. Their high incidence in Mexico and the etiologic heterogeneity observed in several studies, prompted the present investigation with the main objective of looking for risk factors associated to NTD. We analyzed maternal exposure during the first trimester of pregnancy to different environmental factors, such as acute or chronic illnesses, immunizations, smoking, alcoholism, maternal or paternal occupation and exposure to chemicals. The sample include 360 patients with anencephaly, 249 with spina bifida and 44 with encephalocele, ascertained from a total of 230 635 live births and 4,020 stillborns, studied in the Mexican program of Registro y Vigilancia Epidemiológica de Malformaciones Congénitas Externa. Of the risk factors considered, significant differences with the control group were found for anencephaly in relation to maternal viral upper respiratory infection, hyperthermia, ingestion of analgesics, antiemetics and paternal occupation. In the case of spina bifida, significant differences were found only for viral upper respiratory infections.     

A targeted sequencing panel identifies rare damaging variants in multiple genes in the cranial neural tube defect,anencephaly

Neural tube defects (NTDs) affecting the brain (anencephaly) are lethal before or at birth, whereas lower spinal defects (spina bifida) may lead to lifelong neurological handicap. Collectively, NTDs rank among the most common birth defects worldwide. This study focuses on anencephaly, which despite having a similar frequency to spina bifida and being the most common type of NTD observed in mouse models, has had more limited inclusion in genetic studies. A genetic influence is strongly implicated in determining risk of NTDs and a molecular diagnosis is of fundamental importance to families both in terms of understanding the origin of the condition and for managing future pregnancies. Here we used a custom panel of 191 NTD candidate genes to screen 90 patients with cranial NTDs (n = 85 anencephaly and n = 5 craniorachischisis) with a targeted exome sequencing platform. After filtering and comparing to our in‐house control exome database (N = 509), we identified 397 rare variants (minor allele frequency, MAF < 1%), 21 of which were previously unreported and predicted damaging. This included 1 frameshift (PDGFRA), 2 stop‐gained (MAT1A; NOS2) and 18 missense variations. Together with evidence for oligogenic inheritance, this study provides new information on the possible genetic causation of anencephaly.     

Risk factors associated with neural tube defects

Spina bifida represents a broad category of neural tube defects (NTD) which affects approximately 1–4/1,000 live births. Since effective prenatal diagnostic testing for 90 % of NTD is available through measurement of alpha-fetoprotein (AFP) in amniotic fluid, ascertainment of high risk factors associated with the occurrence of NTD would be both desirable and important. At the present time, generally, the major indication for prenatal testing for NTD is the presence of a first-degree relative with some form of NTD. To date, few other factors have been utilized to identify a family as "at risk".
We have studied a group of 19 families of 10 female and 9 male index cases with NTD. The parents of each index case were interviewed and pedigrees were prepared on each family. Conditions screened for in these families included spina bifida and other NTD, pilonidal cysts, scoliosis, kyphosis and other vertebral disorders which were hypothesized to be possibly related to NTD. There were 58 first-, 171 second-, and 802 third-degree relatives screened in this study. This sample population was similarly characteristic with regard to sex, maternal age and birth order distributions as compared to previous populations of NTD described and was therefore considered to be representative. Our results indicate that: (1) pilonidal cysts are 6 times more frequent in the fathers and twice as frequent in the mothers of children with spina bifida than in the general population;
These preliminary studies suggest that several minor clinical conditions in parents may be important to consider as possible risk signs suggesting couples be considered for prenatal evaluation for the prevention of NTD.     

Homocysteine remethylation enzyme polymorphisms and increased risks for neural tube defects

Folic acid supplementation can effectively reduce the risk of neural tube defects (NTDs); however, the mechanism underlying this beneficial effect remains unclear. Recent evidence suggests that certain folate pathway genes, as well as those related to homocysteine metabolism might be contributing to this effect. The purpose of this study is to investigate whether gene polymorphisms of methionine synthase (MTR) and methionine synthase reductase (MTRR) are involved in the risk for NTDs, specifically spina bifida. We detected MTR A2756G and MTRR A66G polymorphisms using PCR-RFLP analysis in a group of NTD infants, their mothers and normal controls. We found that infants with the MTRR mutant genotype had a 2.6-fold higher risk of NTDs when compared to the AA genotype (OR = 2.6, 95%CI = 1.3-5.3). Mothers with the MTRR mutant genotype also had a 1.9-fold higher risk of having an NTD baby compared to AA genotype (OR = 1.9, 95%CI = 1.1-3.1). Infants who carry mutant alleles for both MTRR and MTR had exceptionally elevated NTD risks, with odds ratios of 5.1 compared to infants with the wild type genotype at both loci (AA + AA) (OR = 5.1, 95%CI = 1.7-15.4). A comparable result was observed in the mothers of NTD cases (OR = 2.1, 95%CI = 1.0-4.7). Our results indicate that MTRR and MTR genes may interact to increase the infants' NTD risks. These results did not appear to be influenced by maternal periconceptional folic acid intake. However,the sample size of this study was limited, and a larger population study is needed to pursue these initial observations.     

Evaluation of 64 candidate single nucleotide polymorphisms as risk factors for neural tube defects in a large Irish study population

Individual studies of the genetics of neural tube defects (NTDs) contain results on a small number of genes in each report. To identify genetic risk factors for NTDs, we evaluated potentially functional single nucleotide polymorphisms (SNPs) that are biologically plausible risk factors for NTDs but that have never been investigated for an association with NTDs, examined SNPs that previously showed no association with NTDs in published studies, and tried to confirm previously reported associations in folate‐related and non‐folate‐related genes. We investigated 64 SNPs in 34 genes for association with spina bifida in up to 558 case families (520 cases, 507 mothers, 457 fathers) and 994 controls in Ireland. Case‐control and mother‐control comparisons of genotype frequencies, tests of transmission disequilibrium, and log‐linear regression models were used to calculate effect estimates. Spina bifida was associated with over‐transmission of the LEPR (leptin receptor) rs1805134 minor C allele [genotype relative risk (GRR): 1.5; 95% confidence interval (CI): 1.0–2.1; P = 0.0264] and the COMT (catechol‐O‐methyltransferase) rs737865 major T allele (GRR: 1.4; 95% CI: 1.1–2.0; P = 0.0206). After correcting for multiple comparisons, these individual test P‐values exceeded 0.05. Consistent with previous reports, spina bifida was associated with MTHFR 677C>T, T (Brachyury) rs3127334, LEPR K109R, and PDGFRA promoter haplotype combinations. The associations between LEPR SNPs and spina bifida suggest a possible mechanism for the finding that obesity is a NTD risk factor. The association between a variant in COMT and spina bifida implicates methylation and epigenetics in NTDs. © 2010 Wiley‐Liss, Inc.     

The 19-bp deletion polymorphism in intron-1 of dihydrofolate reductase (DHFR) may decrease rather than increase risk for spina bifida in the Irish population

Periconceptional maternal folic acid supplementation can prevent up to 70% of pregnancies affected with neural tube defects (NTDs), including spina bifida. This has focused attention on folate-related genes such as dihydrofolate reductase (DHFR) in a bid to identify the genetic factors that influence NTD risk through either the fetal or maternal genotype. We considered a novel intronic 19-bp deletion polymorphism and two polymorphisms within the 3' untranslated region (721A>T and 829C>T) of the DHFR gene as candidates for NTD risk. We studied NTD cases (n=283), mothers of cases (n=280), fathers of cases (n=279), and controls (n=256). We did not find the DHFR 829C>T polymorphism to be variable within the Irish population. The 19-bp intron deletion and the 721A>T polymorphisms were found to be in linkage disequilibrium. In contrast to a previous study, the 19-bp intron deletion allele did show a significant protective effect in mothers of NTD cases when present in one (relative risk 0.59 [95%CI: 0.39-0.89], P=0.01) or two copies (relative risk 0.52 [95%CI: 0.32-0.86], P=0.01). Analysis of mRNA levels revealed a small increase in expression ( approximately 1.5-fold) associated with the 19-bp intron deletion polymorphism, but this was not significant. In conclusion, the DHFR intron 19-bp deletion allele may be a protective NTD genetic factor by increasing DHFR mRNA levels in pregnant women.     

Spontaneous abortion–high risk factor for neural tube defects in subsequent pregnancy

An increased spontaneous abortion rate has been observed in pregnancies preceding that of fetuses or newborn infants with neural tube defects (NTDs). There are 2 suggested explanations for this observation. One is that a trophoblastic cell rest, remaining from a previous aborted pregnancy, interferes with normal embryogenesis. The second is that the previous lost fetus was affected with NTD. We studied the obstetric history of mothers of newborn infants with NTDs compared to those with other birth defects, in low and high risk groups for NTD (Jew and Bedouins). A significantly higher spontaneous abortion rate (48%) in the preceding pregnancy was found in the NTD group compared to the group with other birth defects (20%). This was especially remarkable for spina bifida cases in the Jewish study population. A significantly higher rate of preceding spontaneous abortion was also found in congenital heart defects (CHD) when compared to other congenital malformations. A hypothesis based on the multifactorial threshold model is put forward to explain these findings. Based on the realization that spontaneous abortion constitutes a high risk factor for NTD and possibly also CHD, we recommend a delay of subsequent pregnancy and periconceptional treatment with folic acid following spontaneous abortion. © 1994 Wiley-Liss, Inc.     

The methionine synthase reductase 66A>G polymorphism is a maternal risk factor for spina bifida

The methionine synthase reductase (MTRR) enzyme restores methionine synthase (MTR) enzyme activity and therefore plays an essential role in homocysteine remethylation. In some studies, the 66A>G polymorphism in the MTRR gene was associated with increased neural tube defect (NTD) risk. Using a case-control design, we studied the association between the MTRR 66A>G polymorphism and spina bifida risk in 121 mothers, 109 spina bifida patients, 292 control women, and 234 pediatric controls. Possible interactions between the MTRR 66A>G variant and the MTR 2756A>G polymorphism, the MTHFR 677C>T variant, plasma vitamin B12, and plasma methylmalonic acid (MMA) levels were examined in the 121 mothers and 292 control women. Meta-analyses were conducted to set the results of the case-control study in the context of eligible literature on the relation between the MTRR 66A>G variant and NTD risk. Finally, a transmission disequilibrium test was performed for 82 complete mother–father–child triads to test for preferential transmission of the MTRR risk allele. In our case-control study, the MTRR 66A>G polymorphism had no influence on spina bifida risk in children [odds ratio (OR) 0.6, 95% confidence interval (CI) 0.4–1.1]. The MTRR 66GG genotype increased maternal spina bifida risk by 2.1-fold (OR 2.1, 95% CI 1.3–3.3). This risk became more pronounced in combination with the MTHFR 677TT genotype (OR 4.0, 95% CI 1.3–12.5). Moreover, we demonstrate a possible interaction between the MTRR 66GG genotype and high plasma MMA levels (OR 5.5, 95% CI 2.2–13.5). The meta-analyses demonstrated that the maternal MTRR 66GG genotype was associated with an overall 55% (95% CI 1.04–2.30) increase in NTD risk and that the MTRR 66GG genotype did not increase NTD risk in children (OR 0.96, 95% CI 0.46–2.01). These data show that the MTRR 66GG genotype is a maternal risk factor for spina bifida especially when intracellular vitamin B12 status is low.     

Molecular genetic analysis of the gene encoding the trifunctional enzyme MTHFD (methylenetetrahydrofolate-dehydrogenase, methenyltetrahydrofolate-cyclohydrolase, formyltetrahydrofolate synthetase) in patients with neural tube defects

It is now well recognized that periconceptional folic acid or folic acid containing multivitamin supplementation reduces the risk of neural tube defects (NTDs). Recently we were able to show that homozygosity for a thermolabile variant of the enzyme methylenetetrahydrofolate reductase is associated with an increased risk for spina bifida in patients recruited from the Dutch population. However, this genetic risk factor could not account for all folic acid preventable NTDs. In an attempt to identify additional folate related enzymes that contribute to NTD etiology we now studied the methylenetetrahydrofolate dehydrogenase gene on chromosome 14q24 which encodes a single protein with three catalytic properties important in the folate metabolism. The cDNA sequence of 38 familial and 79 sporadic patients was screened for the presence of mutations by single strand conformation polymorphism (SSCP) analysis followed by sequencing. Two amino acid substitutions were identified. The first one (R293H) was detected in a patient with familial spina bifida and not in 300 control individuals. The mutation was inherited from the unaffected maternal grandmother and was also present in two younger brothers of the index patient, one of them displaying spina bifida occulta and the other being unaffected. The second change turned out to be an amino acid polymorphism (R653Q) that was present in both patients and controls with similar frequencies. Our results so far provide no evidence for a major role of the methylenetetrahydrofolate-dehydrogenase (MTHFD) gene in NTD etiology. However, the identification of a mutation in one family suggests that this gene can act as a risk factor for human NTD.     

Multigeneration maternal transmission in Italian families with neural tube defects

Periconceptional vitamin supplementation with folate prevents about three-quarters of expected cases of neural tube defects (NTDs) in clinical trials. However, vitamin action may be regulated at the level of the gene, and individual susceptibility to environmental agents, including dietary components, also may be under genetic control. We investigated the presence of familial factors in a retrospective case control study of neural tube defects in Genoa, Italy. Cases included all patients treated at a single pediatric neurosurgical service. Controls matched on age and sex came from the same hospital. We found strong evidence for the contribution of genetic factors in this study. There was an excess risk of 14 for the occurrence of NTDs in first-degree relatives compared to controls (P < .0005). There was no difference in sex ratio in any group of relatives, but maternal grandparents of children with a high spinal lesion had 14% fewer offspring than paternal grandparents (P < .005), possibly because of excess miscarriages. Our study is the first to show complex patterns of inheritance in spina bifida families affecting three generations in one clinical subgroup and preferentially on the mother's side. These results support a role for genomic imprinting and highlight the value of multidisciplinary epidemiologic and clinical studies that include multiple generations. New studies incorporating dietary and genetic approaches will help clarify and extend these findings. © 1996 Wiley-Liss, Inc.     

Evaluation of genetic variants in the reduced folate carrier and in glutamate carboxypeptidase II for spina bifida risk

Genetic variants in folate metabolism have been reported to increase risk for neural tube defects (NTD). The first such sequence change was the 677C-->T substitution in methylenetetrahydrofolate reductase (MTHFR), but additional sequence changes have been identified in enzymes or transporters for folates. Two recently identified variants are the 1561C-->T (H475Y) mutation in glutamate carboxypeptidase II (GCPII) and the 80A-->G (H27R) change in the reduced folate carrier RFC-1. We examined a group of mothers of spina bifida offspring, and a group of control women, for the above polymorphisms to assess their impact on NTD risk as well as on homocysteine and nutrient (RBC folate, serum folate, and serum cobalamin) levels. The GCPII variant (in the heterozygous state) did not influence NTD risk or metabolite levels; homozygous mutant (YY) women were not observed in our study group. The homozygous mutant (RR) genotype for the RFC-1 gene was not associated with a significant difference in NTD risk (OR=1.39, 95% CI=0.55-3.54), but there was a borderline significant (p=0.065) decrease in RBC folate levels, compared with the HH genotype. However, the combination of the RR genotype for RFC-1 and low RBC folate was associated with a significant 4.6-fold increase in NTD risk (OR=4.6, 95% CI=1.47-14.37). Since this small study is the first to demonstrate increased risk for women with the RFC-1 variant for having a child with a NTD, additional larger studies are required to confirm this change as another potential genetic modifier for spina bifida risk.     

Upper and lower neural tube defects: an alternate hypothesis.

It has been suggested that neural tube defects (NTDs) of the upper type (anencephaly, encephalocele, and thoracic spina bifida) may have a pathogenesis different from those of the lower type (lumbosacral spina bifida), since recurrent cases within a sibship were said always to be concordant with respect to NTD type. Also, spontaneous abortion, additional malformation, and recurrence rate were observed to be higher in the upper group, and there was an excess of females in upper NTD probands. To test this hypothesis, we measured the above variables in upper and lower NTDs in a sample from Quebec. We found less than full concordance (50%) of NTD type in 18 sib pairs. Recurrence rate was not significantly lower in the lower NTD group (5.6 v 5.8%). The other variables were in general agreement with previous studies, inconsistent findings possibly attributable to different NTD population incidences. These findings can be accounted for if upper and lower NTDs share a similar pathogenesis and the embryo is more susceptible during early than late neural tube formation.     

Trends in periconceptional folic acid use by relatives in Irish families with neural tube defects

Close relatives in families who have a child with a neural tube defect (NTD) are at greatly increased risk of having an affected child. Periconceptional folic acid reduces the risk of both occurrence and recurrence of NTDs substantially. Public health authorities currently recommend that the diets of all women between the ages of 15 and 44 who are capable of becoming pregnant be supplemented with folic acid tablets daily. We wondered if relatives in NTD families were more likely to use folic acid. From data obtained by interview with uncles and aunts in Irish NTD families we evaluated folic acid use in 144 of their pregnancies occurring between 1990 and 2000. There was a significant trend towards increasing use of folic acid both before and during pregnancy over the 10 years covered by the study. During the most recent years, 1998-2000, 57.9% of pregnancies reported by aunts were supplemented beforehand and 89.5% during the pregnancy. Pregnancies to smokers were significantly less likely to be supplemented with folic acid. In this study close relatives of an NTD child were more likely to report periconceptional folic acid use than the general public. While these results are encouraging, more remains to be done to ensure in this high risk group to ensure that the full prevention potential of folic acid is realised.     

Genetic susceptibility to neural tube defect pregnancy varies with offspring phenotype

Neural tube defects (NTDs) have a well-established genetic basis, although no single genetic factor has been identified as a major risk factor in NTD susceptibility. A large number of association studies have been conducted to investigate the possibility that NTD susceptibility is linked to polymorphic variation in genes involved in early embryonic development or in the absorption or metabolism of folate, a nutrient that has been clearly associated with a reduction in the risk of NTD pregnancy. A study of three candidate gene polymorphisms at loci implicated in folate absorption and metabolism has been conducted on a population of 211 mothers of a heterogeneous mix of NTD phenotypes: 59% spina bifida aperta (SBA), 20.3% spina bifida occulta (SBO), 17% anencephaly, and 3.7% other NTD. Allele and genotype frequencies were stratified according to offspring NTD phenotype, and variation in the level of NTD risk was associated with different phenotypes. All the three variants (MTHFR 677C > T, GCPII 1561C > T, and RFC-1 80G > A) were shown to significantly influence the risk of anencephalic pregnancy. In addition, the MTHFR 677C > T variant conferred a modest protective effect in SBO mothers and the total NTD mother group, but not in SBA mothers. The RFC-1 80G > A variant elevated the risk of SBO and anencephalic pregnancy. The findings of this study suggest that NTD phenotypic heterogeneity may help explain the mixed findings of previous association studies and that different polymorphisms may hold differing degrees of significance for the various NTD phenotypes.     

Epidemiology of neural tube defects

The epidemiological investigation of the common open neural tube defects (NTDs), anencephaly, and spina bifida, has a long history. The most significant finding from these past studies of NTDs was the identification of the protective effect of maternal, periconceptional supplementation with folic acid. Fortuitously, the association between folic acid and NTDs became widely accepted in the early 1990s, at a time when genetic association studies of complex traits were becoming increasingly feasible. The confluence of these events has had a major impact on the direction of epidemiological, NTD research. Association studies to evaluate genes that may influence the risk of NTDs through their role in folate-related processes, or through other metabolic or developmental pathways are now commonplace. Moreover, the study of genetic as well as non-genetic, factors that may influence NTD risk through effects on the nutrient status of the mother or embryo has emerged as a major research focus. Research efforts over the past decade indicate that gene-gene, gene-environment, and higher-order interactions, as well as maternal genetic effects influence NTD risk, highlighting the complexity of the factors that underlie these conditions. The challenge for the future is to design studies that address these complexities, and are adequately powered to detect the factors or combination of factors that influence the development of NTDs.     

Susceptibility to spina bifida; an association study of five candidate genes

Clues regarding candidate genes which influence susceptibility to spina bifida and anencephaly come from the identification of folate-associated risk factors and from studies of mouse mutants showing neural tube anomalies. On this basis we selected five candidate genes; CBS , MS , MTHFR , T ( Brachyury ) and BRCA1 for genetic analysis in 31 Dutch and 48 British NTD families. Ten polymorphisms, two for each gene, were used in transmission tests for disequilibrium (TDT). In six instances more than 50 transmissions from heterozygous parents could be examined. Using TDT we find evidence for an association between an allele at the T gene and liability to NTD in the embryo. Data from British and Dutch populations showed the same trend and in combination gave a χ²_TDT=4.89, P =0.03 (OR 2.39, CI 95% 1.02–5.61). No association, in either population group, was found for CBS , MS and MTHFR , the enzymes most directly associated with the known risk factors in folate metabolism. The possibility of complex genetic interactions was explored; the data show that a Gly919 MS variant occurs more frequently in combination with the MTHFR thermolabile variant in mothers of NTD offspring (OR 3.94, CI 95% 1.0–16.3).     

Interactions between breast-feeding, specific parental atopy, and sex on development of asthma and atopy

BACKGROUND: The influence of breast-feeding on the risk of developing atopy and asthma remains controversial. OBJECTIVE: To examine asthma and atopy outcomes by sex, reported specific parental history of atopy, and breast-feeding. METHODS: In a birth cohort, we examined childhood asthma and atopy (positive skin prick tests) by sex and breast-feeding in relation to maternal and paternal atopy. Interactions were explored in logistic regression models. RESULTS: For boys, breast-feeding (odds ratio [OR], 1.63; 95% CI, 0.93-2.87; P = .09) and maternal atopy (OR, 1.95; 95% CI, 0.93-4.08; P = .08) were each associated with atopy at age 13 years. Breast-feeding increased the risk for atopy among boys with paternal atopy (OR, 7.39; 95% CI, 2.21-24.66) compared with non-breast-fed boys with paternal atopy, but did not significantly further increase risk among subjects with maternal atopy. For girls, breast-feeding (OR, 0.74; 95% CI, 0.41-1.31) and maternal and paternal atopy were not independent risk factors for atopy at age 13 years. However, breast-feeding increased the risk for atopy in girls with maternal atopy (OR, 3.13; 95% CI, 1.20-8.14) compared with non-breast-fed girls with maternal atopy. There was no such effect among subjects with paternal atopy. Results for the outcome of asthma followed a similar pattern. CONCLUSION: The influence of breast-feeding on development of atopy and asthma differs by sex and by maternal and paternal atopy, and is most significant among subjects at lower baseline risk. CLINICAL IMPLICATIONS: Analyses of environmental risk factors for asthma and atopy should be stratified by specific parental atopy and sex.