A non-mosaic PORCN mutation in a male with severe congenital anomalies overlapping focal dermal hypoplasia.

Mutations in the PORCN gene cause the X-linked dominant condition focal dermal hypoplasia (FDH). Features of FDH include striated pigmentation of the skin, ocular and skeletal malformations. FDH is generally associated with in utero lethality in non-mosaic males and most of the currently reported male patients show mosaicism due to de novo post-zygotic mutations in the PORCN gene. There is only one previous report of a surviving male with an inherited mutation in the PORCN gene. Here, we report two male siblings with multiple malformations including skeletal, ocular and renal defects overlapping with FDH. A novel PORCN mutation (p.Ser250Phe) was identified in a non-mosaic, hemizygous state in one of the siblings who survived to 8 years of age. The mother is a heterozygous carrier, has a random X-inactivation pattern and is asymptomatic. Findings unusual for FDH include dysplastic clavicles and bilateral Tessier IV facial clefts. This is the second case report of a non-mosaic PORCN mutation in a male individual with multiple congenital anomalies. While the pathogenicity of this mutation remains to be further investigated, the survival of a male with a non-mosaic mutation in PORCN is suggestive of a functionally mild mutation leading to an X-linked recessive mode of inheritance.

Gene variants as risk factors for gastroschisis.

In a population-based case-control study in California of 228 infants, we investigated 75 genetic variants in 20 genes and risk of gastroschisis with regard to maternal age, race/ethnicity, vitamin use, and smoking exposure. We hypothesized that genes related to vascular compromise may interact with environmental factors to affect the risk of gastroschisis. Haplotypes were constructed for 75 gene variants using the HaploView program. Risk for gastroschisis associated with each gene variant was calculated for both the homozygotes and the heterozygotes, with the homozygous wildtypes as the referent. Risks were estimated as odds ratios (ORs) with 95% confidence intervals (CIs) by logistic regression. We found 11 gene variants with increased risk and four variants with decreased risk of gastroschisis for heterozygous (ORh ) or homozygous variants (ORv ) genotypes. These included NOS3 (rs1036145) ORh = 0.4 (95% CI: 0.2-0.7); NOS3 (rs10277237) ORv = 2.7 (95% CI: 1.3-6.0); ADD1 (rs12503220) ORh = 2.9 (95% CI: 1.6-5.4), GNB3 (rs5443) ORh = 0.2 (95% CI: 0.1-0.5), ORv = 0.4 (95% CI: 0.2-0.9); ICAM1 (rs281428) ORv = 6.9 (95% CI: 2.1-22.9), ICAM1 (rs3093030) ORv = 2.6 (95% CI: 1.2-5.6); ICAM4 (rs281438) ORv = 4.9 (95% CI: 1.4-16.6), ICAM5 (rs281417) ORh = 2.1 (95% CI: 1.1-4.1), ORv = 4.8 (95% CI: 1.7-13.6); ICAM5 (rs281440) ORh = 23.7 (95% CI: 5.5-102.5), ORv = 20.6 (95% CI: 3.4-124.3); ICAM5 (rs2075741) ORv = 2.2 (95% CI: 1.1-4.4); NAT1 ORv = 0.3 (95% CI: 0.1-0.9). There were additional associations between several gene variants and gastroschisis among women aged 20-24 and among mothers with and without vitamin use. NOS3, ADD1, ICAM1, ICAM4, and ICAM5 warrant further investigation in additional populations and with the interaction of additional environmental exposures. © 2016 Wiley Periodicals, Inc.

Genetic polymorphisms in ESR1 and ESR2 genes, and risk of hypospadias in a multiethnic study population.

PURPOSE: Estrogenic endocrine disruptors acting via estrogen receptors α (ESR1) and ß (ESR2) have been implicated in the etiology of hypospadias, a common congenital malformation of the male external genitalia. We determined the association of single nucleotide polymorphisms in ESR1 and ESR2 genes with hypospadias in a racially/ethnically diverse study population of California births. MATERIALS AND METHODS: We investigated the relationship between hypospadias and 108 ESR1 and 36 ESR2 single nucleotide polymorphisms in 647 cases and 877 population based nonmalformed controls among infants born in selected California counties from 1990 to 2003. Subgroup analyses were performed by race/ethnicity (nonHispanic white and Hispanic subjects) and by hypospadias severity (mild to moderate and severe). RESULTS: Odds ratios for 33 of the 108 ESR1 single nucleotide polymorphisms had p values less than 0.05 (p = 0.05 to 0.007) for risk of hypospadias. However, none of the 36 ESR2 single nucleotide polymorphisms was significantly associated. In stratified analyses the association results were consistent by disease severity but different sets of single nucleotide polymorphisms were significantly associated with hypospadias in nonHispanic white and Hispanic subjects. Due to high linkage disequilibrium across the single nucleotide polymorphisms, haplotype analyses were conducted and identified 6 haplotype blocks in ESR1 gene that had haplotypes significantly associated with an increased risk of hypospadias (OR 1.3 to 1.8, p = 0.04 to 0.00001). Similar to single nucleotide polymorphism analysis, different ESR1 haplotypes were associated with risk of hypospadias in nonHispanic white and Hispanic subjects. No significant haplotype association was observed for ESR2. CONCLUSIONS: The data provide evidence that ESR1 single nucleotide polymorphisms and haplotypes influence the risk of hypospadias in white and Hispanic subjects, and warrant further examination in other study populations.

One-carbon metabolite levels in mid-pregnancy and risks of conotruncal heart defects.

BACKGROUND: Evidence exists for an association between use of vitamin supplements with folic acid in early pregnancy and reduced risk for offspring with conotruncal heart defects. A few observations have been made about nutrients related to one-carbon metabolism other than folate. Our prospective study attempted to extend information on nutrition and conotruncal heart defects by measuring analytes in mid-pregnancy sera. METHODS: This study included data from a repository of women's mid-pregnancy serum specimens based on screened pregnancies in California from 2002-2007. Each woman's specimen was linked with delivery information to determine whether her fetus had a conotruncal heart defect or another structural malformation, or was nonmalformed. We identified 140 conotruncal cases and randomly selected 280 specimens as nonmalformed controls. Specimens were tested for a variety of analytes, including homocysteine, methylmalonic acid, folate, vitamin B12 , pyridoxal phosphate, pyridoxal, pyridoxic acid, riboflavin, total choline, betaine, methionine, cysteine, cystathionine, arginine, asymmetric and symmetric dimethylarginine. RESULTS AND CONCLUSIONS: We did not observe statistical evidence for substantial differences between cases and controls for any of the measured analytes. Analyses specifically targeting B-vitamins also did not reveal differences between cases and controls.

Maternal occupational exposure to polycyclic aromatic hydrocarbons and risk of oral cleft-affected pregnancies.

OBJECTIVE: To evaluate whether there is an association between maternal occupational exposure to polycyclic aromatic hydrocarbons and oral clefts in offspring. This is the first human study of polycyclic aromatic hydrocarbons and clefts of which the authors are aware. DESIGN: Case-control study. SETTING, PARTICIPANTS: Data for 1997 to 2002 from the National Birth Defects Prevention Study, a large population-based case-control study in the United States, were analyzed. Maternal telephone interviews yielded information on jobs held in the month before through 3 months after conception. Two industrial hygienists independently assessed occupational exposure to polycyclic aromatic hydrocarbons; all jobs rated as exposed or with rating difficulty were reviewed with a third industrial hygienist to reach consensus on all exposure parameters. Logistic regression estimated crude and adjusted odds ratios with 95% confidence intervals for cleft lip with or without cleft palate and cleft palate alone. RESULTS: There were 2989 controls (3.5% exposed), 805 cases of cleft lip with or without cleft palate (5.8% exposed), and 439 cases of cleft palate alone (4.6% exposed). The odds of maternal occupational exposure to polycyclic aromatic hydrocarbons (any versus none) during pregnancy was increased for cleft lip with or without cleft palate cases as compared with controls (odds ratio, 1.69; 95% confidence interval, 1.18 to 2.40); the odds ratio was 1.47 (95% confidence interval 1.02 to 2.12) when adjusted for maternal education. There was a statistically significant adjusted exposure-response relationship for cleft lip with or without cleft palate (Ptrend = .02). Odd ratios for cleft palate alone were not statistically significant. CONCLUSIONS: Maternal occupational exposure to polycyclic aromatic hydrocarbons was associated with increased risk of cleft lip with or without cleft palate in offspring.

Maternal and infant gene-folate interactions and the risk of neural tube defects.

Neural tube defects (NTDs) are common, serious malformations with a complex etiology that suggests involvement of both genetic and environmental factors. The authors evaluated maternal or offspring folate-related gene variants and interactions between the gene variants and maternal intake of folates on the risk of NTDs in their offspring. A case-control study was conducted on mothers and/or their fetuses and infants who were born in California from 1999 to 2003 with an NTD (cases n = 222, including 24 mother-infant pairs) or without a major malformation (controls n = 454, including 186 mother-infant pairs). Maternal intake of folates was assessed by food frequency questionnaire and genotyping was performed on samples from mothers and infants. For mothers in the lowest folate-intake group, risk of NTDs in offspring was significantly decreased for maternal MTHFR SNPs rs1476413, rs1801131, and rs1801133 (odds ratio [OR] = 0.55, 80% confidence interval [CI]: 0.20, 1.48; OR = 0.58, 80% CI: 0.24, 1.43; OR = 0.69, 80% CI: 0.41, 1.17, respectively), and TYMS SNPs rs502396 and rs699517 (OR = 0.91, 80% CI: 0.53, 1.56; OR = 0.70, 80% CI: 0.38, 1.29). A gene-only effect was observed for maternal SHMT1 SNP rs669340 (OR = 0.69, 95% CI: 0.49, 0.96). When there was low maternal folate intake, risk of NTDs was significantly increased for infant MTHFD1 SNPs rs2236224, rs2236225, and rs11627387 (OR = 1.58, 80% CI: 0.99, 2.51; OR = 1.53, 80% CI: 0.95, 2.47; OR = 4.25, 80% CI: 2.33, 7.75, respectively) and SHMT1 SNP rs12939757 (OR = 2.01, 80% CI: 1.20, 3.37), but decreased for TYMS SNP rs2847153 (OR = 0.73, 80% CI: 0.37, 1.45). Although power to detect interaction effects was low for this birth defects association study, the gene-folate interactions observed in this study represent preliminary findings that will be useful for informing future studies on the complex etiology of NTDs.

Maternal occupational exposure to polycyclic aromatic hydrocarbons and risk of neural tube defect-affected pregnancies.

BACKGROUND: This study evaluated whether there is an association between maternal occupational exposure to polycyclic aromatic hydrocarbons (PAHs) and neural tube defects (NTDs) in offspring. This is the first such study of which the authors are aware. METHODS: Data were analyzed from 1997 to 2002 deliveries in the National Birth Defects Prevention Study, a large population-based case-control study in the United States. Maternal interviews yielded information on jobs held in the month before through 3 months after conception. Three industrial hygienists blinded to case or control status assessed occupational exposure to PAHs. Crude and adjusted odds ratios (ORs) with 95% confidence intervals (CIs) were estimated using unconditional logistic regression. RESULTS: Of the 520 mothers of children with NTDs, 5.0% were classified as exposed to occupational PAHs, as were 3.5% of the 2989 mothers of controls. The crude OR for PAH exposure was 1.43 (95% CI, 0.92-2.22) for any NTD and 1.71 (95% CI, 1.03-2.83) for spina bifida. Adjusted ORs were smaller in magnitude and not significant. Among women who were normal weight or underweight, the crude OR for spina bifida was 3.13 (95% CI, 1.63-6.03) and adjusted OR was 2.59 (95% CI, 1.32-5.07). Based on estimated cumulative exposure, a statistically significant dose-response trend was observed for spina bifida; however, it was attenuated and no longer significant after adjustment. CONCLUSION: Maternal occupational exposure to PAHs may be associated with increased risk of spina bifida in offspring among women who are normal weight or underweight. Other comparisons between PAHs and NTDs were consistent with no association.

Mutations in KAT6B, encoding a histone acetyltransferase, cause Genitopatellar syndrome.

Genitopatellar syndrome (GPS) is a skeletal dysplasia with cerebral and genital anomalies for which the molecular basis has not yet been determined. By exome sequencing, we found de novo heterozygous truncating mutations in KAT6B (lysine acetyltransferase 6B, formerly known as MYST4 and MORF) in three subjects; then by Sanger sequencing of KAT6B, we found similar mutations in three additional subjects. The mutant transcripts do not undergo nonsense-mediated decay in cells from subjects with GPS. In addition, human pathological analyses and mouse expression studies point to systemic roles of KAT6B in controlling organismal growth and development. Myst4 (the mouse orthologous gene) is expressed in mouse tissues corresponding to those affected by GPS. Phenotypic differences and similarities between GPS, the Say-Barber-Biesecker variant of Ohdo syndrome (caused by different mutations of KAT6B), and Rubinstein-Taybi syndrome (caused by mutations in other histone acetyltransferases) are discussed. Together, the data support an epigenetic dysregulation of the limb, brain, and genital developmental programs.

Craniosynostosis and nutrient intake during pregnancy.

OBJECTIVE: To examine the association of craniosynostosis with maternal intake of folic acid-containing supplements and dietary nutrients. METHODS: The study included deliveries from 1997 to 2005 from the National Birth Defects Prevention Study. Nonsyndromic infants with craniosynostosis (n = 815) were compared to nonmalformed, population-based liveborn control infants (n = 6789), by estimating adjusted odds ratios (AORs) and 95% confidence intervals (CIs) from logistic regression models that included mother's age, parity, race-ethnicity, education, body mass index, smoking, alcohol, fertility treatments, plurality, and study center. We compared quartiles of intake and specified nutrients as continuous. RESULTS: Intake of folic acid-containing supplements was not associated with craniosynostosis (AORs were close to 1). Analyses of dietary nutrients were restricted to mothers who took supplements during the first trimester (i.e., most women). Based on continuous specifications of nutrients, sagittal synostosis risk was significantly lower among women with higher intake of riboflavin and vitamins B6, E, and C; metopic synostosis risk was significantly higher among women with higher intakes of choline and vitamin B12; and coronal synostosis risk was significantly lower among women with higher intake of methionine and vitamin C. As examples, AORs for sagittal synostosis among women with intakes of vitamin B6 and riboflavin in the highest versus lowest quartiles were 0.4 (95% CI, 0.2-0.6) and 0.5 (95% CI, 0.3-0.7), respectively. CONCLUSION: This analysis suggests that dietary intake of certain nutrients may be associated with craniosynostosis, and results may vary by suture type.

Planar cell polarity pathway genes and risk for spina bifida.

Spina bifida, a neural tube closure defect (NTD) involving the posterior portion of what will ultimately give rise to the spinal cord, is one of the most common and serious birth defects. The etiology of spina bifida is thought to be multi-factorial and involve multiple interacting genes and environmental factors. The causes of this congenital malformation remain largely unknown. However, several candidate genes for spina bifida have been identified in lower vertebrates, including the planar cell polarity (PCP) genes. We used data from a case-control study conducted in California to evaluate the association between variation within several key PCP genes and the risk of spina bifida. The PCP genes included in this study were the human homologs of the Xenopus genes Flamingo, Strabismus, Prickle, Dishevelled, and Scrib, two of the homologs of Xenopus Wnt genes, WNT5A and WNT11, and two of the homologs of Xenopus Frizzled, FZD3 and FZD6. None of the 172 SNPs that were evaluated were significantly associated with spina bifida in any racial/ethnic group after correction for multiple testing. However, several SNPs in the PRICKLE2 gene had unadjusted P-value <0.01. In conclusion, our results, though largely negative, suggest that the PRICKLE2 gene may potentially modify the risk of spina bifida and deserves further investigation.

Genetic polymorphisms in the thioredoxin 2 (TXN2) gene and risk for spina bifida.

TXN2 encodes human thioredoxin 2, a small redox protein important in cellular antioxidant defenses, as well as in the regulation of apoptosis. Txn2 knockout mice fail to complete neural tube closure by E10.5 and die in utero. We hypothesized that genetic variation in human TXN2 gene may alter the function of the encoded protein in a manner associated with an increased risk for neural tube defects (NTDs). A DNA re-sequencing effort of the human TXN2 gene was taken. After a variation in the promoter was identified, the transcriptional activity of different alleles was investigated. The possible association between these variations and the risk of spina bifida was further evaluated in a subset of samples obtained from a large population-based case-control study in California in two different ethnicity groups, non-Hispanic white and Hispanic white. We identified a novel promoter insertion polymorphism located 9 base pairs upstream of the transcription start site of exon 1(-9 insertion). The GA, G and GGGA insertions were associated with a marked decrease of transcriptional activity when overexpressed in both U2-OS (an osteosarcoma cell line) and 293 cells (derived from human embryonic kidney). Further analysis revealed that the GA insertion was associated with increased spina bifida risk for Hispanic whites. Our study revealed a novel Ins/Del polymorphism in the human TXN2 gene proximal promoter region that altered the transcriptional activity and is associated with spina bifida risk. This polymorphism may be a genetic modifier of spina bifida risk in this California population.

Autoantibodies to folate receptor during pregnancy and neural tube defect risk.

Periconceptional folic acid can reduce the occurrence of neural tube defects (NTDs) by up to 70%, and autoantibodies for folate receptors (FRs) have been observed in serum from women with a pregnancy complicated by an NTD. This population-based cohort study has examined serum from pregnant mothers for autoantibodies to FRs, antibodies to bovine folate binding protein (FBP), and inhibition of folic acid binding to FR and FBP in association with NTD risk. The mid-gestational maternal serum specimens used for this study were collected during the 15-18th week of pregnancy. Samples were obtained from the California Birth Defects Monitoring Program; 29 mothers had a pregnancy complicated by spina bifida and 76 mothers had unaffected children. The presence of IgG and IgM antibodies to human FR, bovine FBP, and inhibition of folic acid binding to FR and FBP was determined. Higher activity of IgM to FBP in cases verses controls was observed (P=0.04). Higher activity of IgM and IgG autoantibodies to FR was observed (P<0.001 and P=0.04, respectively). Risk estimates at two standard deviations above average control antibody concentrations were OR=2.07 (CI=1.02, 4.06) for anti-FBP IgM, OR=2.15 (CI=1.02, 4.69) for anti-FR IgG and OR=3.19 (CI=1.47, 6.92) for anti-FR IgM. These data support the hypothesis that high titers of antibodies and blocking of folic acid binding to FRs by maternal serum should be regarded as risk factors for NTDs.

Genetic susceptibilities in the association between maternal exposure to tobacco smoke and the risk of nonsyndromic oral cleft.

Maternal tobacco consumption is considered as a risk factor for nonsyndromic oral clefts. However, this risk is moderate and may be modulated by genetic susceptibilities, including variants of the TGFA, TGFB3 and MSX1 developmental genes and polymorphisms of genes of the CYP (1A1, 2E1) and GST (M1, T1) families involved in metabolic pathways of tobacco smoke compounds. This French case-control study (1998-2001; 240 nonsyndromic cases, 236 controls) included a case-parent design (175 triad-families) that made it possible to distinguish the direct effect of the child's genotype and maternally mediated effects. Maternal smoking during the first trimester of pregnancy was not associated with the oral cleft risk in this population, but we observed statistically significant increased risks associated with maternal exposure to environmental tobacco smoke (ETS). No variant of any of the three developmental genes was significantly associated with oral cleft. The fetal CYP1A1*2C variant allele was associated with a statistically significant decreased risk, compared with the homozygous wild-type: relative risk = 0.48, 95% confidence interval: 0.2, 1.0. Suggestive reduced risks were also observed for the maternal CYP1A1*2C allele and the fetal CYP2E1*5 allele. The GSTM1 and GSTT1 deletions appeared to play no role. Our findings suggest some interactions, with the strongest between ETS and CYP1A1 or MSX1 and between maternal smoking and CYP2E1. We did not confirm the maternal smoking-infant GSTT1 null interaction previously reported by other investigators.

Gene-nutrient interactions: importance of folic acid and vitamin B12 during early embryogenesis.

The role that nutritional factors play in mammalian development has received renewed attention over the past two decades as the scientific literature has exploded with reports that folic acid supplementation in the periconceptional period can protect embryos from a number of highly significant malformations. As is often the case, the relationship between B vitamin supplementation and improved pregnancy outcomes is more complicated than initially perceived, as the interaction between nutritional factors and selected genes must be considered. In this review, we attempt to summarize the complex clinical and experimental literature on nutritional factors, their biological transport mechanisms, and interactions with genetic polymorphisms that impact early embryogenesis. While not exhaustive, our goal was to provide an overview of important gene-nutrient interactions, focusing on folic acid and vitamin B12, to serve as a framework for understanding the multiple roles they play in early embryogenesis.

Gastroschisis: a gene-environment model involving the VEGF-NOS3 pathway.

Gastroschisis is a severe major malformation in which an infant is delivered with a portion of intestines and possible other abdominal organs extruding through a defect in the abdominal wall, usually to the right of the umbilical cord. Etiologies of gastroschisis are largely unknown, and even its pathogenesis is poorly understood. Several recent epidemiological studies have identified interactions between maternal smoking during pregnancy, genetic variants of endothelial nitric oxide synthase, and risk for gastroschisis. We present a brief review of the endothelial nitric oxide synthase pathway and its relationship to vasculogenesis, suggesting that disruption of this pathway by environmental exposures or by genetic variation may represent one pathogenetic model for gastroschisis.

Nicotinamide N-methyl transferase (NNMT) gene polymorphisms and risk for spina bifida.

BACKGROUND: Moderate hyperhomocysteinemia is a known risk factor for NTDs in a variety of experimental model systems and is believed to be important in humans as well. The enzyme nicotinamide N-methyl transferase (NNMT) was identified in a genome-wide linkage scan as being an important regulator of homocysteine homeostasis in a Spanish population, making it an interesting candidate gene for NTDs. METHODS: We evaluated 11 SNPs (single nucleotide polymorphism) of the NNMT gene in our study population. In this study, 252 cases (infants with spina bifida) and 335 controls (nonmalformed infants), born during the period 1983-1986 in selected counties in California, were genotyped for variants of the NNMT gene. Allelic, genotype, and haplotype associations with spina bifida risk were evaluated and analyzed. RESULTS: None of the SNPs studied alone showed allelic or genotypic associations with spina bifida. However, the TCAG haplotype for block 3 (rs2852447, ra2852425, rs4646337, and rs11569688) showed a decreased risk for spina bifida among non-Hispanic Whites (OR 0.4; 95%CI: 0.1-1.0). CONCLUSIONS: No association was found between infant NNMT gene variants and risk for spina bifida in our study population. However, small sample sizes for most variant groups and for phase-unknown haplotype data limited the power of the study.

Maternal periconceptional smoking and alcohol consumption and risk for select congenital anomalies.

BACKGROUND: This study examined the association between maternal smoking and alcohol use (including binge drinking) during the periconceptional period (i.e., 2 months before through 2 months after conception) and the risk of orofacial clefts, NTDs, and conotruncal heart defects in offspring. METHODS: Data were drawn from a population-based case-control study of fetuses and live-born infants among a cohort of California births between July 1999 and June 2003. The 1,355 cases comprised of 701 orofacial clefts, 337 NTDs, and 323 conotruncal heart defects. Information on smoking and alcohol consumption was obtained via telephone interviews with mothers of 1,355 (80% of eligibles) cases and 700 (77% of eligibles) nonmalformed, live-born controls. RESULTS: Maternal smoking of five cigarettes or less per day was associated with reduced risks of NTDs (OR 0.7; 95% CI: 0.3, 1.4), whereas the risk associated with higher cigarette consumption was lower for conotruncal heart defects (OR 0.5; 95% CI: 0.2, 1.2). Maternal intake of alcohol less than 1 day per week was associated with a 1.6- to 2.1-fold higher risk of NTDs (95% CI: 0.9, 2.6), d-transposition of the great arteries (95% CI: 1.1, 3.2), and multiple cleft lip with or without cleft palate (CLP) (95% CI: 0.8, 4.5). Risks associated with more frequent alcohol intake were 2.1 for NTDs (95% CI: 1.1, 4.0) and 2.6 for multiple CLP (95% CI: 1.1, 6.1). CONCLUSIONS: This study observed that maternal alcohol intake increased the risk for d-transposition of the great arteries, NTDs, and multiple CLP in infants. By contrast, smoking was associated with a lower risk of NTDs and conotruncal heart defects.

Craniosynostosis and maternal smoking.

BACKGROUND: Several previous studies suggested increased risk of craniosynostosis among infants born to women who smoked. METHODS: This study used data from the National Birth Defects Prevention Study, a multi-state, population-based case-control study of infants delivered from 1997-2003. Nonmalformed, liveborn controls were selected randomly from birth certificates or birth hospitals. Data from maternal telephone interviews were available for 531 cases and 5008 controls. RESULTS: Smoking during the first month of pregnancy was not associated with craniosynostosis. Smoking later in pregnancy was associated with increased risk, but only among mothers who smoked at least one pack/day. For example, during the second trimester, the odds ratio for smoking <5 cigarettes/day was 1.0 (95% confidence interval [CI] 0.6, 1.8), but the odds ratio (OR) for smoking 15 or more cigarettes/day was 1.6 (95% CI 0.9, 2.8), after adjustment for maternal age, education, race-ethnicity, sub-fertility, parity, folic acid supplement intake, body mass index, and study center. Among women who did not smoke, adjusted odds ratios suggested that secondhand smoke exposure at home, but not at work/school, was associated with modestly increased risk; the OR for home exposure was 1.3 (95% CI 0.9, 1.9). Results followed a similar pattern for some, but not all, specific suture types, but numbers for some groupings were small. CONCLUSIONS: The results suggest moderately increased risk of craniosynostosis among mothers who were the heaviest smokers and who continued to smoke after the first trimester. Results are somewhat equivocal, given that most confidence intervals included one.

NAT2 variation and idiopathic talipes equinovarus (clubfoot).

Idiopathic talipes equinovarus (ITEV), or isolated clubfoot, is a common developmental anomaly that is characterized by a rigid foot, adducted forefoot, cavus midfoot, equinovarus of the hindfoot, and hypoplastic calf musculature. The etiology of this common birth defect is largely unknown, but genetic factors have been implicated in population and family studies and maternal smoking during pregnancy has been identified as an environmental risk factor. The biotransformation of exogenous substances, such as tobacco smoke, is modulated by numerous genes including N-acetylation genes, NAT1 and NAT2. Functional variants of these genes exist and can be distinguished by genotyping. We hypothesized that variation in NAT1 and NAT2 genes might be associated with ITEV. To test this hypothesis, NAT1 and NAT2 were genotyped in a sample of 56 multiplex ITEV families, 57 trios with a positive family history and 160 simplex trios with ITEV. The results detected a slight decrease in the expected number of homozygotes for the NAT2 normal allele in the Hispanic simplex trios. In addition, in a pilot case-control study of ITEV, there were significantly more slow NAT2 acetylators among the cases. This suggests that slow acetylation may be a risk factor for ITEV. This study is the first to find evidence suggesting a role for a biotransformation candidate gene in the etiology of ITEV and provides a scientific foundation to further explore the contributions of other tobacco metabolism genes in the etiology of clubfoot.

Maternal smoking during early pregnancy, GSTP1 and EPHX1 variants, and risk of isolated orofacial clefts.

OBJECTIVE: To examine the interactions between four fetal xenobiotic metabolizing gene polymorphisms, maternal cigarette smoking, and risk for oral cleft defects. DESIGN AND PARTICIPANTS: California population-based case-control study of 431 infants born with isolated orofacial clefts and 299 nonmalformed controls. MAIN OUTCOME MEASURES: Infants were genotyped for functional polymorphisms of the detoxification enzymes microsomal epoxide hydrolase-1 (EPHX1 T-->C [Tyr113His], and A-->G [His139Arg]), and glutathione-S transferase Pi-1 (GSTP1 A-->G [Ile105Val] and C-->T [Ala114Val]), and risks for cleft outcomes were measured for gene only and gene-maternal smoking effects. RESULTS: Although smoking was associated with an increased risk for isolated cleft lip+/-palate, we found no independent associations of genotypes of EPHX1-codon 113 or GSTP1-codon 105 polymorphisms for either isolated cleft lip+/-palate or isolated cleft palate. The heterozygote genotype for the EPHX1-codon 139 polymorphism was associated with an increased risk of isolated cleft palate (odds ratio=1.6 [95% confidence interval, 1.0 to 2.6]). Infant EPHX1 and GTSP1 polymorphic variants did not appreciably alter the risks for clefts associated with maternal smoking, nor were any EPHX1 combined genotype-specific risks found. Infant genotypes of the GSTP1-codon 105 polymorphism, combined with glutathione-S-transferase-mu-1 null genotypes, did not appreciably alter the risk of orofacial clefts. CONCLUSIONS: Our results suggest that genetic variation of the detoxification enzymes EPHX1 and GSTP1 did not increase the risks of orofacial clefting, nor do they influence the risks associated with maternal smoking.