Idiopathic clubfoot is a common orthopedic birth defect that affects approximately 135,000 newborns worldwide. It is characterized by equinus, varus and adductus deformities of the ankle and foot. While numerous studies suggest a multifactorial etiology, the specific genetic and environmental components have yet to be delineated. Maternal smoking during pregnancy is the only common environmental factor consistently shown to increase the risk for clubfoot. Moreover, a positive family history of clubfoot, in conjunction with maternal smoking, increases the risk twenty-fold. These findings suggest that genetic variation in smoking metabolism (xenobiotic) genes may increase susceptibility to clubfoot. Based on this reasoning, we interrogated eight candidate genes (CYP1A1, CYP1A2, CYP1B1, CYP2A6, EPHX1, NAT2, GSTM1 and GSTT1), chosen based on their involvement in xenobiotic metabolism. Twenty-two SNPs and two null alleles in these genes were genotyped in a dataset composed of nonHispanic white and Hispanic multiplex and simplex families. Only rs1048943/CYP1A1 had significantly altered transmission in the aggregate and multiplex NHW datasets (p=0.003 and p=0.009, respectively). Perturbation of CYP1A1 can cause an increase in harmful, adduct forming metabolic intermediates. A significant interaction between EPHX1 and NAT2 was also found (p=0.007). Importantly, for CYP1A2, significant maternal (p=0.03; RR=1.24; 95% CI: 1.04–1.44) and fetal (p=0.01; RR=1.33; 95% CI: 1.13–1.54) genotypic effects were identified, suggesting that both maternal and fetal genotypes can negatively impact limb development. No association was found between maternal smoking status and variation in xenobiotic metabolism genes. Together, these results suggest that xenobiotic metabolism genes are unlikely to play a major role in clubfoot, however, perturbation of this pathway may still play a contributory role.
clubfoot; smoking; tobacco; genetics; xenobiotic genes; CYP
The central nervous system is derived from the neural plate that undergoes a series of complex morphogenetic events resulting in formation of the neural tube in a process known as neurulation. The cellular behaviors driving neurulation in the cranial region involve forces generated by the neural tissue itself as well as the surrounding epithelium and mesenchyme. Of interest, the cranial mesenchyme underlying the neural plate undergoes stereotypical rearrangements hypothesized to drive elevation of the neural folds. As the neural folds rise, the hyaluronate-rich extracellular matrix greatly expands resulting in increased space between individual cranial mesenchyme cells. Based on inhibitor studies, expansion of the extracellular matrix has been implicated in driving neural fold elevation; however, since the surrounding neural and epidermal ectoderm were also affected by inhibitor exposure, these studies are inconclusive. Similarly, treatment of neurulating embryos with teratogenic doses of retinoic acid results in altered organization of the cranial mesenchyme but alterations in surrounding tissues are also observed. The strongest evidence for a critical role for the cranial mesenchyme in neural fold elevation comes from studies of genes expressed exclusively in the cranial mesenchyme that when mutated result in exencephaly associated with abnormal organization of the cranial mesenchyme. Twist is the best studied of these and is expressed in both the paraxial mesoderm and neural crest derived cranial mesenchyme. Here we review the evidence implicating the cranial mesenchyme in providing a driving force for neural fold elevation to evaluate whether there is sufficient data to support this hypothesis.
A critical event in neural tube closure is the formation of median (MHP) and dorsolateral hinge points (DLHP). Together, they buckle the ventral midline, elevate and juxtapose the neural folds for proper neural tube closure. Dynamic cell behaviors occur at hinge points (HPs), but their molecular regulation is largely unexplored. Bone Morphogenetic Proteins (BMPs) have been implicated in a variety of neural tube closure defects, although the underlying mechanisms are poorly understood.
In this study we used in vivo electroporations, high-resolution microscopy and biochemical analyses to explore the role of BMP signaling in chick midbrain neural tube closure.
We identified a cell-cycle dependent BMP gradient in the midbrain neural plate, which results in low-level BMP activity at the MHP. We show that while BMP signaling does not play a role in midbrain cell-fate specification, its attenuation is necessary and sufficient for MHP formation and midbrain closure. BMP blockade induces MHP formation by regulating apical constriction and basal nuclear migration. Furthermore, BMP signaling is critically important for maintaining epithelial organization by biochemically interacting with apicobasal polarity proteins (e.g., PAR3). Thus prolonged BMP blockade disrupts apical junctions, desegregating the apical (PAR3+, ZO1+) and basolateral (LGL+) compartments. Direct apical LGL-GFP misexpression in turn is sufficient to induce ectopic HPs.
BMPs play a critical role in maintaining epithelial organization, a role that is conserved across species and tissue-types. Its’ cell-cycle dependent modulation in the neural plate dynamically regulates apicobasal polarity and helps bend, shape and close the neural tube.
Noggin; hinge point; apical constriction; basal nuclear migration; PAR3; Lethal giant larva; tight junctions; adherens junctions; interkinetic nuclear migration; midbrain
Spina bifida is one of the most common of all human structural birth defects. Despite considerable effort over several decades, the causes and mechanisms underlying this malformation remain poorly characterized. In order to better understand the pathogenesis of this abnormality, we conducted a microarray study using Mouse Whole Genome Bioarrays which have ~36,000 gene targets, to compare gene expression profiles between two mouse models; CXL-Splotch and Fkbp8Gt(neo)which express a similar spina bifida phenotype. We anticipated that there would be a collection of overlapping genes or shared genetic pathways at the molecular level indicative of a common mechanism underlying the pathogenesis of spina bifida during embryonic development.
A total of 54 genes were determined to be differentially expressed (25 down regulated, 29 upregulated) in the Fkbp8Gt(neo) mouse embryos; while 73 genes were differentially expressed (56 down regulated, 17 upregulated) in the CXL-Splotch mouse relative to their wildtype controls. Remarkably, the only two genes that showed decreased expression in both mutants were v-ski sarcoma viral oncogene homolog (Ski), and Zic1, a transcription factor member of the zinc finger family. Confirmation analysis using real time qRT-PCR indicated that only Zic1 was significantly decreased in both mutants. Gene Ontology analysis revealed striking enrichment of genes associated with mesoderm and central nervous system development in the CXL-Splotch mutant embryos, whereas in the Fkbp8Gt(neo) mutants, the genes involved in dorsal/ventral pattern formation, cell fate specification, and positive regulation of cell differentiation were most likely to be enriched. These results indicate that there are multiple pathways and gene networks perturbed in mouse embryos with shared phenotypes.
spina bifida; gene expression; microarray; mouse mutants
A common and severe neural tube defect (NTD) phenotype, myelomeningocele (MM), results from the defective closure of the caudal end of the neural tube with herniation of the spinal cord and meninges through the vertebral column. The exact mechanisms for NTDs are unknown, but excessive oxidative stress, particularly in association with maternal diabetes, has been postulated as a mechanism for MM.
The SNPlex Genotyping (ABI, Foster City, CA) platform was used to investigate single nucleotide polymorphisms (SNPs) across the superoxide dismutase (SOD) 1 and 2 genes to assess their association with MM risk. The study population included 329 trio (affected child and both parents) and 281 duo (affected child and one parent) families. Only cases with documented MM were studied. Seventeen SNPs across the SOD1 and SOD2 genes met the quality control criteria to be considered for statistical analysis. Genetic association was assessed using the family-based Transmission Disequilibrium Test in PLINK.
Four SNPs in the SOD1 gene (rs 202446, rs202447, rs4816405 and rs2070424) and one SNP in the SOD2 gene (rs5746105) appeared to be associated with MM risk in our population. After adjusting for multiple testing, these SNPs remained significant.
The study provides the first genetic evidence to support association of myelomeningocele with superoxide scavenging. The rare alleles of the five specific SNPs within SOD1 and SOD2 appear to confer a protective effect on the susceptibility for MM risk in the MM population tested. Further evaluation of the roles of superoxide scavenging and neural tube development is warranted.
Neural tube defects (NTDs) are common human birth defects with a complex etiology. To develop a comprehensive knowledge of the genes expressed during normal neurulation, we established transcriptomes from human neural tube fragments during and after neurulation using long Serial Analysis of Gene Expression (long-SAGE).
Rostral and caudal neural tubes were dissected from normal human embryos aged between 26 and 32 days of gestation. Tissues from the same region and Carnegie stage were pooled (n>=4) and total RNA extracted to construct four long-SAGE libraries. Tags were mapped using the UniGene Homo sapiens 17 bp tag-to-gene best mapping set. Differentially expressed genes were identified by chi-square or Fisher’s exact test and validation was performed for a subset of those transcripts using in situ hybridization. In silico analyses were performed with BinGO and EXPANDER.
We observed most genes to be similarly regulated in rostral and caudal regions, but expression profiles differed during and after closure. In silico analysis found similar enrichments in both regions for biological process terms, transcription factor binding and miRNA target motifs. Twelve genes potentially expressing alternate isoforms by region or developmental stage, and the miRNAs miR-339-5p, miR-141/200a, miR-23ab, and miR-129/129-5p, are among several potential candidates identified here for future research.
Time appears to influence gene expression in the developing central nervous system more than location. These data provide a novel complement to traditional strategies of identifying genes associated with human NTDs, and offer unique insight into the genes associated with normal human neurulation.
gene expression; Homo sapiens; long-SAGE; neurulation; neural tube defects
Nitrosatable drugs can react with nitrite in the stomach to form N-nitroso compounds, and results from animal studies suggest that N-nitroso compounds are teratogens. With data from the National Birth Defects Prevention Study, the relation between prenatal exposure to nitrosatable drugs and limb deficiencies, oral cleft, and heart malformations in offspring was examined.
Maternal reports of drugs taken during the first trimester of pregnancy were classified with respect to nitrosatability for mothers of 741 babies with limb deficiencies, 2,774 with oral cleft malformations, 8,091 with congenital heart malformations, and 6,807 without major congenital malformations. Nitrite intake was estimated from maternal responses to a food frequency questionnaire.
Isolated transverse limb deficiencies and atrioventricular septal defects were associated with secondary amine drug exposures (adjusted odds ratios [aOR] 1.51, 95% confidence limit [CI] 1.11, 2.06 and aOR 1.97, 95% CI 1.19, 3.26, respectively). Tertiary amines were associated with hypoplastic left heart syndrome (aOR 1.50, 95% CI 1.10, 2.04) and single ventricle (aOR 1.61, 95% CI 1.06, 2.45). These two malformations were also significantly associated with amide drugs. For several malformations, the strongest associations with nitrosatable drug use occurred among mothers with the highest estimated dietary nitrite intake, especially for secondary amines and atrioventricular septal defects (highest tertile of nitrite, aOR 3.30, 95% CI 1.44, 7.58).
Prenatal exposure to nitrosatable drugs may be associated with several congenital malformations, especially with higher nitrite intake. The possible interaction between nitrosatable drugs and dietary nitrite on risk of congenital malformations warrants further attention.
congenital heart defects; congenital limb deficiency; nitrosatable drug; oral clefts; nitrites
The current understanding of the effects of hypoxia on early embryogenesis is limited. Potential mediators of hypoxic effects include adenosine, which increases dramatically during hypoxic conditions and activates A1 adenosine receptors (A1ARs).
To examine the influences of hypoxia and adenosine signaling on cardiac development, chicken embryos were studied. Real time RT-PCR assay was used to examine the A1AR gene expression during embryogenesis and after siRNA- mediated knock down. Cell proliferation was determined by counting cell nuclei and PhosphoHistone H3 positive cells. Apoptosis was determined by TUNEL assay.
A1ARs were found to be expressed in chicken embryos during early embryogenesis. Treatment of Hamburger and Hamilton stage 4 embryos with the A1AR agonist N6-cyclopentyladenosine caused cardiac bifida and looping defects in 55% of embryos. Hamburger and Hamilton stage 4 embryos exposed to 10% oxygen for 6, 12, 18, and 24 h followed by recovery in room air until stage 11, exhibited cardia bifida and looping defects in 34, 45, 60, and 86% of embryos respectively. Hypoxia-induced abnormalities were reduced when A1AR signaling was inhibited by the A1AR antagonist 1,3 dipropyl-8-cyclopentylxanthine or by siRNA-targeting A1ARs. Hypoxia treatment did not increase apoptosis, but decreased embryonic cell proliferation.
These data indicate that hypoxia adversely influences cardiac malformations during development, in part by A1AR signaling.
hypoxia; adenosine; adenosine receptor; chicken embryo; heart
This study examines gene-environment interaction (GEI) between the MTHFR C667T polymorphism and folic acid in the etiology of orofacial clefts (OFC). We used a pooled-analyticapproach on four studies that used similar methods.
We used logistic regression to analyse the pooled sample of 1149 isolated cases and 1161 controls. Fetal and maternal MTHFR C677T genotypes, and maternal periconceptional exposure to smoking, alcohol, vitamin containing folic acid and folic acid supplements were contrasted between the cleft types [non-syndromic clefts lip or without cleft palate (CL(P)) and non syndromic cleft palate (CP)] and control groups.
There was a reduced risk of CL(P) with maternal folic acid use (p=0.008; OR=0.70, 95% CI: 0.65–0.94) and with supplements containing folic acid (p=0.028, OR=0.80, 95% CI: 0.65–0.94). Maternal smoking increased the risk of both CL(P) (p<10e−3; OR=1.62, 95% CI: 1.35–1.95) and CP (p=0.028; OR=1.38, 95% CI: 1.04–1.83). No significant risk was observed with either maternal or fetal MTHFR C677T genotypes.
This individual paticipant data (IPD) meta-analysis affords greater statistical power and can help alleviate the problems associated with aggregate-level data-sharing. The result of this IPD meta-analysis is consistent with previous reports suggesting that folic acid and smoking influence OFC outcomes.
Cleft lip and palate; MTHFR; Folic acid; Individual patient data; pooled-analysis
The left ventricular outflow tract (LVOT) defects aortic valve stenosis (AVS), coarctation of the aorta (COA), and hypoplastic left heart syndrome (HLHS) represent an embryologically related group of congenital cardiovascular malformations. They are common and cause substantial morbidity and mortality. Prior evidence suggests a strong genetic component in their causation.
We selected NRG1, ERBB3, and ERBB4 of the epidermal growth factor receptor (EGFR) signaling pathway as candidate genes for investigation of association with LVOT defects based on the importance of this pathway in cardiac development and the phenotypes in knockout mouse models. Single nucleotide polymorphism (SNP) genotyping was performed on 343 affected case-parent trios of European ancestry.
We identified a specific haplotype in intron 3 of ERBB4 that was positively associated with the combined LVOT defects phenotype (p = 0.0005) and in each anatomic defect AVS, COA, and HLHS separately. Mutation screening of individuals with an LVOT defect failed to identify a coding sequence or splice site change in ERBB4. RT-PCR on lymphoblastoid cells from LVOT subjects did not show altered splice variant ratios among those homozygous for the associated haplotype.
These results suggest ERBB4 is associated with LVOT defects. Further replication will be required in separate cohorts to confirm the consistency of the observed association.
genetics of cardiovascular disease; heart defects; congenital; congenital abnormalities; cardiovascular abnormalities; analysis; genetic association
The objective of this report is to estimate the benefits of universal meconium screening for maternal drinking during pregnancy. Fetal alcohol spectrum disorder (FASD), including its most severe manifestation fetal alcohol syndrome (FAS), is preventable and remains a public health tragedy. The incidences of FAS and FASD have been conservatively estimated to be 0.97 and 10 per 1000 births, respectively. Meconium testing has been demonstrated to be a promising at-birth method for detection of drinking during pregnancy.
The current costs of FAS and FASD, alcohol treatment programs, and meconium screening were estimated by literature review. Monetary values were converted roughly to equal dollars in 2006.
Costs of adding meconium analysis to the current newborn screening program and of treatment for the identified mothers were estimated and compared to potential averted costs that may result from identification and intervention for mothers and affected infants. Three potential maternal treatment strategies are analyzed. Depending on the treatment type, the savings may range from $6 to $97 for every $1 spent on screening and treatment.
It needs to be emphasized, however, that such screening is premature and that to be effective this screening can be implemented only if there is a societal willingness to institute prevention and intervention programs to improve both women’s and children’s health. Future research should be directed at improving detection and developing in-depth prevention and remedial intervention programs. A thorough consideration of the ethical issues involved in such a screening program is also needed.
fetal alcohol spectrum disorder; meconium; fatty acid ethyl esters; newborn screening; benefits; maternal alcohol use
The role of maternal zinc nutrition in human oral clefts (OCs) is unclear. We measured plasma zinc concentrations (PZn) of case- and control-mothers to evaluate the associations between PZn and risk of OCs with and without other malformations.
Case-mothers were ascertained by the Utah Birth Defects Network and control-mothers were selected from Utah birth certificates by matching for child gender and delivery month and year. Maternal blood was collected > 1 year after the last pregnancy. PZn was available for 410 case-mothers who were divided into four subgroups; isolated cleft lip with or without cleft palate (CL/P-I, n = 231), isolated cleft palate (CP-I, n = 74), CL/P with other malformations (CLP-M, n = 42), and CP with other malformations (CP-M, n = 63). PZn was available for 447 control-mothers. The mean age of children at blood sampling was 3.7 years for all cases combined and 4.3 years for controls.
Mean PZn of all groups were similar, and low PZn (< 11.0 μmol/l) was found in 59% of cases and 62% of controls. Risk of OCs did not vary significantly across PZn quartiles for the four subgroups individually and all OC groups combined.
We previously reported that poor maternal zinc status was a risk factor for OCs in the Philippines, where OC prevalence is high and maternal PZn is low. In Utah, however, no such association was found, suggesting that poor maternal zinc status may become a risk factor only when zinc status is highly compromised.
zinc status; plasma zinc; cleft lip; cleft palate; malformations; human; Utah
Oral clefts are common congenital anomalies and result from defects during embryogenesis. The complex etiology is evident by the number of genes and signaling pathways involved in craniofacial development. Matrix metalloproteinases (MMPs) and their inhibitors TIMPs are responsible for tissue remodeling during craniofacial development.
In this study, we investigated the association of polymorphisms in 14 biologically relevant MMP and TIMP genes in 494 individuals with oral clefts and 413 control individuals from Brazil. Genotypes were generated using Taqman chemistry. Analyses were performed using PLINK software.
Polymorphisms in MMP3 (rs522616) and TIMP2 (rs8179096) showed significant association with all cleft types (all clefts, cleft lip/palate, and cleft palate) (P≤0.002). An additional family-based dataset (881 case-parent trios) from the US was used for confirmation of the association findings (P<0.05). Analysis of gene-gene interaction suggests that MMP3 and TIMP2 may interactively contribute to a cleft phenotype.
This study provides new evidence that variation in MMP3 may contribute to nonsyndromic oral clefts and further supports the involvement of TIMP2 as a cleft susceptibility gene. Although additional studies are still necessary to unveil the exact mechanism by which MMP3 and TIMP2 would contribute to a cleft phenotype, allelic polymorphisms in these genes and their interactions may partly explain the variance of individual susceptibility to oral clefts.
MMP; TIMP; cleft lip; palate; polymorphism; promoter
This review evaluates current knowledge related to trends in the prevalence of hypospadias, the association of hypospadias with endocrine-disrupting exposures, and the potential contribution of genetic susceptibility to its etiology. The review focuses on epidemiologic evidence. Increasing prevalence of hypospadias has been observed, but such increases tend to be localized to specific regions or time periods. Thus, generalized statements that hypospadias is increasing are unsupported. Due to limitations of study designs and inconsistent results, firm conclusions cannot be made regarding the association of endocrine-disrupting exposures with hypospadias. Studies with more rigorous study designs (e.g., larger and more detailed phenotypes) and exposure assessment that encompasses more breadth as well as depth (e.g., specific endocrine-related chemicals) will be critical to make better inferences about these important environmental exposures. Many candidate genes for hypospadias have been identified, but few of them have been examined to an extent that enables solid conclusions. Further studies are needed that include larger sample sizes, comparison groups that are more representative of the populations from which the cases were derived, phenotype-specific analyses, and more extensive exploration of variants. In conclusion, examining the associations of environmental and genetic factors with hypospadias remain important areas of inquiry, although our actual understanding of their contribution to hypospadias risk in humans is currently limited.
hypospadias; urogenital; genes; environment
Genome-wide association studies are now used routinely to identify genes implicated in complex traits. The panels used for such analyses can detect single nucleotide polymorphisms and copy number variants, both of which may help to identify small deleted regions of the genome that may contribute to a particular disease
We performed a candidate gene analysis involving 1221 SNPs in 333 candidate genes for orofacial clefting using 2823 samples from 725 two- and three-generation families with a proband with clefts of the lip and/or palate. We used SNP genotyping, DNA sequencing, high-resolution DNA microarray analysis and long-range PCR to confirm and characterize the deletion events
This dataset had a high duplicate reproducibility rate (99.98%), high Mendelian consistency rate (99.93%), and low missing data rate (0.55%), which provided a powerful opportunity for deletion detection. Apparent Mendelian inconsistencies between parents and child suggested deletion events in 15 individuals in 11 genomic regions. We confirmed deletions involving CYP1B1, FGF10, SP8, SUMO1, TBX1, TFAP2A, and UGT7A1, including both de novo and familial cases. Deletions of SUMO1, TBX1, and TFAP2A are likely to be etiologic
These deletions suggest the potential roles of genes or regulatory elements contained within deleted regions in the etiology of clefting. Our analysis took advantage of genotypes from a candidate-gene-based SNP survey and proved to be an efficient analytical approach to interrogate genes potentially involved in clefting. This can serve as a model to find genes playing a role in complex traits in general.
cleft lip; cleft palate; microdeletion; SNPs; CNV; candidate genes
Prenatal alcohol exposure can alter physical and behavioral development, leading to a range of fetal alcohol spectrum disorders (FASD). Despite warning labels, pregnant women continue to drink alcohol, creating a need to identify effective interventions to reduce the severity of alcohol’s teratogenic effects. Choline is an essential nutrient that influences brain and behavioral development. Recent studies indicate that choline supplementation can reduce the teratogenic effects of developmental alcohol exposure. The present study examined whether choline supplementation during prenatal ethanol treatment could mitigate the adverse effects of ethanol on behavioral development.
Pregnant Sprague-Dawley rats were intubated with 6 g/kg/day ethanol in a binge-like manner from gestational days 5–20; pair-fed and ad lib chow controls were included. During treatment, subjects from each group were intubated with either 250 mg/kg/day choline chloride or vehicle. Spontaneous alternation, parallel bar motor coordination, Morris water maze, and spatial working memory were assessed in male and female offspring.
Subjects prenatally exposed to alcohol exhibited delayed development of spontaneous alternation behavior and deficits on the working memory version of the Morris water maze during adulthood, effects that were mitigated with prenatal choline supplementation. Neither alcohol nor choline influenced performance on the motor coordination task.
These data indicate that choline supplementation during prenatal alcohol exposure may reduce the severity of fetal alcohol effects, particularly on alterations in tasks that require behavioral flexibility. These findings have important implications for children of women who drink alcohol during pregnancy.
fetal alcohol spectrum disorders; treatment; nutrition; ethanol; fetal alcohol syndrome
The Hedgehog signalling pathway is essential for many aspects of normal embryonic development, including formation and patterning of the neural tube. Absence of Shh ligand is associated with the midline defect holoprosencephaly, while increased Shh signalling is associated with exencephaly and spina bifida. To complicate this apparently simple relationship, mutation of proteins required for function of cilia often leads to impaired Shh signalling and to disruption of neural tube closure. In this manuscript, we review the literature on Shh pathway mutants and discuss the relationship between Shh signalling, cilia and neural tube defects.
Neural tube defects; Hedgehog Signalling Pathway; Cilia; Exencephaly; Spina bifida; Intraflagellar Transport; Neural Tube Patterning
Folate one-carbon metabolism has been implicated as a determinant of susceptibility to neural tube defects (NTDs), owing to the preventive effect of maternal folic acid supplementation and the higher risk associated with markers of diminished folate status.
Folate one-carbon metabolism was compared in curly tail (ct/ct) and genetically matched congenic (+ct/+ct) mouse strains using the deoxyuridine suppression test in embryonic fibroblast cells and by quantifying s-adenosylmethionine (SAM) and s-adenosylhomocysteine (SAH) in embryos using liquid chromatography tandem mass spectrometry. A possible genetic interaction between curly tail and a null allele of 5,10-methylenetetrahydrofolate reductase (MTHFR) was investigated by generation of compound mutant embryos.
There was no deficit in thymidylate biosynthesis in ct/ct cells but incorporation of exogenous thymidine was lower than in +ct/+ct cells. In +ct/+ct embryos the SAM/SAH ratio was diminished by dietary folate deficiency and normalised by folic acid or myor-inositol treatment, in association with prevention of NTDs. In contrast, folate deficiency caused a significant increase in SAM/SAH ratio in ct/ct embryos. Loss of MTHFR function in curly tail embryos significantly reduced the SAM/SAH ratio but did not cause cranial NTDs or alter the frequency of caudal NTDs.
Curly tail fibroblasts and embryos, in which Grhl3 expression is reduced, display alterations in one-carbon metabolism, particularly in the response to folate deficiency, compared with genetically-matched congenic controls in which Grhl3 is unaffected. However, unlike folate deficiency, diminished methylation potential appears to be insufficient to cause cranial NTDs in the curly tail strain, and neither does it increase the frequency of caudal NTDs.
Neural tube defects; folate; methylation; curly tail; inositol
Adhesion and fusion of epithelial sheets marks the completion of many morphogenetic events during embryogenesis. Neural tube closure involves an epithelial fusion sequence in which the apposing neural folds adhere initially via cellular protrusions, proceed to a more stable union, and subsequently undergo remodelling of the epithelial structures to yield a separate neural tube roof plate and overlying non-neural ectoderm. Cellular protrusions comprise lamellipodia and filopodia, and studies in several different systems emphasise the critical role of RhoGTPases in their regulation. How epithelia establish initial adhesion is poorly understood but, in neurulation, may involve interactions between EphA receptors and their ephrinA ligands. Epithelial remodelling is spatially and temporally correlated with apoptosis in the dorsal neural tube midline, but experimental inhibition of this cell death does not prevent fusion and remodelling. A variety of molecular signalling systems have been implicated in the late events of morphogenesis, but genetic redundancy, for example among the integrins and laminins, makes identification of the critical players challenging. An improved understanding of epithelial fusion can provide insights into normal developmental processes, and may also indicate the mode of origin of clinically important birth defects.
epithelium; adhesion; fusion; lamellipodia; filopodia; apoptosis; neurulation; neural tube defects; spina bifida; morphogenesis
Antihistamines are commonly used during pregnancy. There is little evidence that they have teratogenic effects, but there are knowledge gaps with respect to newer products, as well as the relationship between specific antihistamines and specific birth defects.
Using the National Birth Defects Prevention Study (1997-2003), the authors examined associations between maternal use of 14 antihistamines during early pregnancy and 26 isolated major birth defects. A Bayesian analysis incorporating prior knowledge about the relationships between the antihistamines, birth defects, and measured covariates was conducted.
Of the 364 associations investigated, 24 had 95% posterior intervals excluding 1.0. All 24 associations were positive; 23 associations were of weak to moderate magnitude (posterior odds ratio [OR] < 3.0) and one was strong (OR > 6.0) but very imprecise. Of the 24 associations, 20 were with non-cardiac defects. Eight associations involved the antihistamine diphenhydramine.
The results of this study generally were consistent with no association between birth defects and antihistamine use during early pregnancy. Several of the findings might warrant further investigation, although the observed elevated associations should be interpreted in the context of the number of associations investigated and the analysis of retrospective, self-reported data.
antihistamine; Bayesian methods; birth defects; H1 blockers; pregnancy
This study examined the association between 49 markers in the Runt-related transcription factor 2 (RUNX2) gene and nonsyndromic cleft lip with/without cleft palate (CL/P) among 326 Chinese case-parent trios, while considering gene-environment (GxE) interaction and parent-of-origin effects. Five single-nucleotide polymorphisms (SNPs) showed significant evidence of linkage and association with CL/P and these results were replicated in an independent European sample of 825 case-parent trios. We also report compelling evidence for interaction between markers in RUNX2 and environmental tobacco smoke (ETS). Although most marginal SNP effects (i.e., ignoring maternal exposures) were not statistically significant, eight SNPs were significant when considering possible interaction with ETS when testing for gene (G) and GxE interaction simultaneously or when considering GxE alone. Independent samples from European populations showed consistent evidence of significant GxETS interaction at two SNPs (rs6904353 and rs7748231). Our results suggest genetic variation in RUNX2 may influence susceptibility to CL/P through interacting with ETS.
RUNX2; oral clefts; gene-environment interaction; parent-of-origin effects; imprinting
The presence of strain-specific modifier genes is known to modulate the phenotype and pathophysiology of mice harboring genetically engineered mutations. Thus, identification of genetic modifier genes is requisite to understanding control of phenotypic expression. c-Ski is a transcriptional regulator. Ski−/− mice on a C57BL6J (B6) background exhibit facial clefting, while Ski−/− mice on a 129P3 (129) background present with exencephaly.
In the present study, oligonucleotide-based gene expression profiling was utilized to identify potential strain-specific modifier gene candidates present in wild-type mice of B6 and 129 genetic backgrounds. Changes in gene expression were verified by TaqMan quantitative real-time PCR.
Steady-state levels of 89 genes demonstrated a significantly higher level of expression, and those of 68 genes demonstrated a significantly lower level of expression in the developing neural tubes from E8.5, B6 embryos when compared to expression levels in neural tubes derived from E8.5, 129 embryos.
Based on the results from the current comparative microarray study, and taking into consideration a number of relevant published reports, several potential strain-specific gene candidates, likely to modify the craniofacial phenotypes in various knockout mouse models have been identified.
Neural tube; embryo; mouse; modifier gene; microarray
Intracranial lipomas are rare, but 45% of them occur along the midline cisterns between the hemispheres and are often associated with corpus callosum hypoplasia and craniofacial defects. They are difficult to detect, as they are generally asymptomatic and visible by MRI or by postmortem examination. The exact cause of these interhemispheric lipomas is not known, but they arise from a developmental defect resulting in the maldifferentiation of mesenchymal cells into mesodermal derivatives that are not normally present. We have identified a new mouse mutant called tuft, exhibiting a forebrain, intracranial lipoma with midline craniofacial defects resembling frontonasal dysplasia (FND) that arose spontaneously in our wild-type 3H1 colony. The tuft trait appears to be transmitted in recessive fashion, but approximately 80% less frequent than the expected Mendelian 25%, due to either incomplete penetrance or prenatal lethality. MRI and histological analysis revealed that the intracranial lipoma occurred between the hemispheres and often protruded through the sagittal suture. We also observed a lesion at the lamina terminalis that may indicate improper closure of the anterior neuropore. We have mapped the tuft trait to within an 18 cM region on mouse chromosome 10 by microsatellite linkage analysis and identified several candidate genes involved with craniofacial development and cellular differentiation of adipose tissue. tuft is the only known mouse model for midline craniofacial defects with an intracranial lipoma. Identifying the gene(s) and mutation(s) causing this early developmental defect will help us understand the pathogenesis of FND and related craniofacial disorders.
intracranial lipoma; craniofacial defect; hypertelorism; corpus callosum; interfrontal bone