Development Of Neural Tube Defects Research Articles

Multiplexing of TMT labeling reveals folate-deficient diet-specific proteome changes in NTDs.

Introduction: In the early stage of embryonic development, the neural tube (NT) cannot be closed properly due to some complex factors, including environmental factors, genetic factors, and the relationship between various factors, leading to the occurrence of neural tube defects (NTDs). Methods: In this study, we induced a mouse model of NTDs by feeding mice with a low-folate diet and intraperitoneally injecting them with 1.5mg/kg methotrexate on E7.5. Fetal mice were achieved at E13.5, and we extracted proteins from brain tissues with trypsin digestion. After enzymatic digestion, peptides were labeled with TMT/iTRAQ and separated in high-performance liquid chromatography (HPLC) for subsequent liquid chromatography tandem mass spectroscopy (LC-MS/MS) analysis. We used gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation to analyze proteomic changes and analyze the functional enrichment of differentially expressed proteins (DEPs) in the NTD mice tissues. Results: A low-folate-induced mouse model was successfully constructed. Folate was used as a sensitizing agent, and the teratogenicity rate of the NTD fetal mice increased to 36.5% when the concentration of methotrexate was at 1.5mg/kg. Mass spectrometry was used to identify 6,614 proteins, and among them, 5,656 proteins were quantified. In the following proteomic analysis, GO classification and KEGG pathway enrichment analysis were conducted, and heatmaps were drawn for differentially expressed proteins (DEPs). The main pathways associated with NTDs, such as the Hedgehog, Wnt, p53, and Hippo signaling pathways and the one-carbon pool mediated by folate, can be identified through a protein-protein interaction (PPI) network. It was also found that the regulation of ribosomal proteins, such as RPL13 and RPL14, which are upregulated in NTDs, has a certain impact on neural tube development. Discussion: Our results revealed proteomic changes in the tissues of low-folate-induced NTD mice. Validation showed that ribosomal proteins play a regulatory role during the development of NTDs and provides new ideas for the pathogenesis and preventive measures of NTDs.

Inpp5e Regulated the Cilium-Related Genes Contributing to the Neural Tube Defects Under 5-Fluorouracil Exposure.

Primary cilia are crucial for neurogenesis, and cilium-related genes are involved in the closure of neural tubes. Inositol polyphosphate-5-phosphatase (Inpp5e) was enriched in primary cilia and closely related to the occurrence of neural tube defects (NTDs). However, the role of Inpp5e in the development of NTDs is not well-known. To investigate whether Inpp5e gene is associated with the neural tube closure, we established a mouse model of NTDs by 5-fluorouracil (5-FU) exposure at gestational day 7.5 (GD7.5). The Inpp5e knockdown (Inpp5e-/-) mouse embryonic stem cells (mESCs) were produced by CRISPR/Cas9 system. The expressions of Inpp5e and other cilium-related genes including intraflagellar transport 80 (Ift80), McKusick-Kaufman syndrome (Mkks), and Kirsten rat sarcoma viral oncogene homolog (Kras) were determined, utilizing quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), western blot, PCR array, and immunofluorescence staining. The result showed that the incidence of NTDs was 37.10% (23 NTDs/62 total embryos) and significantly higher than that in the control group (P < 0.001). The neuroepithelial cells of neural tubes were obviously disarranged in NTD embryos. The mRNA and protein levels of Inpp5e, Ift80, Mkks, and Kras were significantly decreased in NTD embryonic brain tissues, compared to the control (P < 0.05). Knockdown of the Inpp5e (Inpp5e-/-) reduced the expressions of Ift80, Mkks, and Kras in mESCs. Furthermore, the levels of α-tubulin were significantly reduced in NTD embryonic neural tissue and Inpp5e-/- mESCs. These results suggested that maternal 5-FU exposure inhibited the expression of Inpp5e, which resulted in the downregulation of cilium-related genes (Ift80, Mkks, and Kras), leading to the impairment of primary cilium development, and ultimately disrupted the neural tube closure.

Effect of Zuclopenthixol Acetate on Neural Tube Development in Early Chick Embryos

Objective: Neural tube defects are one of the congenital malformations of the central nervous system. Although the factors that cause the development of neural tube defects and their mechanisms of action are still not clearly explained, genetic predisposition, drug use and some environmental factors are thought to play a role. In this study, it was aimed to investigate the effects of zuclopenthixol acetate (ZA) on neural tube development in a chick embryo model.&#x0D; Methods: Fourty specific pathogen-free (SPF) eggs were used in the study. The eggs were incubated for 28 hours and divided into four groups of 10 eggs each. At the end of the 28th hours, saline was injected to the control group, while ZA was administered subblastodermically to the experimental groups in 3 different doses (0.7, 1.4, 2.1 mg/kg). At the end of the 48th hours, all the eggs were opened and the embryos were dissected from the embryonic membranes and evaluated morphologically and histopathologically.&#x0D; Results: When the study groups were evaluated according to the neural tube positions (open or closed), it was found that the neural tube patency increased depending on the ZA dose, which was statistically significant (p &lt; 0.05). In addition, morphological developments of embryos were evaluated. Compared to the control group, a statistically significant decrease was observed in the mean somite numbers in all ZA-treated groups, while a significant decrease was found in the mean cranio-caudal length only in the high-dose group.&#x0D; Conclusion: In this study, it was observed that neural tube and morphological development were adversely affected in the groups treated with ZA in the chick embryo model. It was shown that neural tube closure defects in embryos increased in direct proportion with ZA doses. However, we believe that it will not be possible to fully adapt the results of this study, which was carried out in the chick embryo model, to humans and that more comprehensive research should be conducted.

Maternal folic acid supplementation and the risk of ankyloglossia (tongue-tie) in infants; a systematic review.

Maternal folic acid supplementation is protective against the development of neural tube defects (NTDs) in babies. However, recent public-facing communications have raised concerns about a causal relationship between folic acid supplementation, particularly after the first trimester, and ankyloglossia (tongue-tie) in infants. Non-evidence-based communications are potentially harmful because they could adversely affect adherence to folic acid supplementation, increasing NTD occurrence. This study aimed to review evidence on the relationships between maternal folic acid supplementation during preconception and/or pregnancy and the risk of ankyloglossia in infants. We searched the databases MEDLINE, EMBASE, Cochrane CENTRAL, and Scopus. We searched for observational, and interventional studies, and systematic reviews investigating the effect of maternal folic acid supplementation during preconception or pregnancy on the occurrence of ankyloglossia in offspring. The search was registered on PROSPERO on 01/12/2022, ID: CRD42022375862. The database searches yielded 93 articles. After removing duplicates and screening titles and abstracts, 26 remained. One article was judged relevant for inclusion in analyses; a case-control study that directly mentions the relationship between folic acid supplementation and ankyloglossia. This study reported that regular intake of folic acid supplements was higher in women with infants with ankyloglossia. However, this study has limitations regarding design, selection bias, and confounding, calling the findings into question. Insufficient evidence exists for a relationship between folic acid supplementation and ankyloglossia. Currently, the benefits of folic acid supplementation far outweigh the risks. This must be clearly communicated to patients by their clinicians during preconception and antenatal care.

Effects of sodium fluoride on neural tube development in chick embryos

ObjectiveVarious environmental factors encountered in daily life are associated with the development of neural tube defects. This study aims to investigate the effects of fluoride on neural tube development in chick embryos. MethodsA total of 60 specific pathogen-free, fertile, zero-day Leghorn-type eggs were used in the study. Group 1 was the control group, in which only saline was administered. Group 2 was the low-dose group, in which 0.003 mg of fluoride was administered, and Group 3 was the high-dose group, in which 0.006 mg of fluoride was administered. After 72 h of incubation, the embryonic disc was evaluated microscopically. ResultsIn the control group, the surface ectoderm of all sections was intact, the neural tube was closed, and the neuroepithelium, the basement membrane surrounding the neuroepithelium, the somites, and the notochord displayed standard structure. Neural tube defects were observed in 3 of the chick embryos, that was given low-dose fluoride. In Group 3, which was administered high doses of fluoride, neural tube defects were observed in 4 embryos. It was observed that the development of neural tube defects was no statistically significantly higher in low and high-dose fluoride group compared to the control group. ConclusionLow and high-dose fluoride exposure was associated with developing neural tube defects, but there was no statisticaly significance.

Frequency of fetal anomalies on antenatal scans in pediatric surgery department of mayo hospital.

Objective: To know the prevalence of congenital anomalies in foeti on antenatal ultrasonography and role of preconceptual folic acid intake in preventing development of neural tube defects. Study Design: Cross-sectional Descriptive study. Setting: Department of Pediatric Surgery, Mayo Hospital, Lahore. Period: January 2022 to September 2022. Material &amp; Methods: The data of 57 patients was collected, who were referred to Pediatric Surgery Department of Mayo Hospital, Lahore from lady Wallington Hospital and Lady Aitchison Hospital, with fetal anomaly detected on antenatal scan. Women with pregnancy in second and third trimester, ranging in age from 20 to 45 years, were enrolled in the study. A questionnaire was used to collect the data. We analyzed the data of study usingSPSS 20(Statistical Package for Social Sciences version 20). Results: Data analysis showed higher incidence (34%) of anomalies of central nervous system including neural tube defects. 89% (51/57) women had not taken preconceptual folic acid supplements. Consanguinity was also noted to be an important factor and significant correlation was seen 70% (40/57). Conclusion: The incidence of central nervous system anomalies including neural tube defects was higher (34%) among all anomalies. This might be probably due to preconceptual deficiency of folic acid.

Double whammy: the genetic variants in CECR2 and high Hcy on the development of neural tube defects.

Introduction: Neural tube defects (NTDs) are serious congenital malformations. The etiology of NTDs involves both genetic and environmental factors. Loss of CECR2 in mice has been shown to result in NTDs. Our previous study indicated that high homocysteine (HHcy) levels could further reduced the expression level of CECR2. This investigation aims to explore the genetic influence of the chromatin remodeling gene, CECR2, in humans and determine if HHcy can have a synergistic effect on protein expression. Methods: We conducted Next-Generation Sequencing (NGS) of the CECR2 gene in 373 NTD cases and 222 healthy controls, followed by functional assay application to select and evaluate CECR2 missense variants and subsequent Western blotting to identify protein expression levels. Results: From the analysis, we identified nine rare, NTD-specific mutations within the CECR2 gene. Significantly, four missense variants (p.E327V, p.T521S, p.G701R, and p.G868R) were selected via functional screening. The E9.5 mouse ectodermal stem cell line NE-4C, transfected with plasmids expressing p.E327V, p.T521S, p.G868R variants or a recombinant harboring all four (named as 4Mut), exhibited notable reductions in CECR2 protein expression. Furthermore, exposure to homocysteine thiolactone (HTL), an extremely reactive homocysteine metabolite, amplified the reduction in CECR2 expression, accompanied by a significant increase in the apoptotic molecule Caspase3 activity, a potential NTD inducer. Importantly, folic acid (FA) supplementation effectively counteracted the CECR2 expression decline induced by CECR2 mutation and HTL treatment, leading to reduced apoptosis. Discussion: Our observations underscore a synergistic relationship between HHcy and genetic variations in CECR2 concerning NTDs, thereby reinforcing the concept of gene-environment interaction phenomena in NTD etiology.

Maternal Calcium and Vitamin D Deficiency and Their Relation with Fetal Neural Tube Defects

Background: NTDs are major birth defects associated with significant fetal loss, mortality, morbidity, and healthcare costs. Although periconceptional folic acid supplementation has reduced their occurrence, NTDs remain a global concern. Recent studies have suggested a possible link between low maternal serum calcium and vitamin D levels and the development of NTDs. Objective: The objective of this prospective observational study was to investigate the relationship between maternal calcium and vitamin D deficiency during early pregnancy and the development of fetal neural tube defects (NTDs). Materials and Method: A total of 207 pregnant mothers in their first trimester were included in the study. Data on age, BMI, gestational age, and serum levels of total calcium and 25-hydroxyvitamin D were collected. Calcium deficiency was identified in 61 (29.5%) participants, while vitamin D deficiency was present in 139 (67%). Ultrasound scans between 18-22 weeks of gestation were conducted in 173 participants to search for fetal NTDs. Results: Among the study population, only 1 (0.6%) fetus was found to have an NTD on sonographic scan. The mother of this fetus exhibited both calcium and vitamin D deficiencies. However, statistical analysis did not reveal a significant relationship between maternal calcium and vitamin D deficiency and the development of fetal NTDs. Conclusion: Deficiency of calcium and vitamin D during early pregnancy does not appear to be associated with the development of fetal NTDs. Further research is needed to explore other potential risk factors for NTDs

An optimal combination of five main monomer components in Wuzi Yanzong Pill that prevents neural tube defects and reduces apoptosis and oxidative stress

Ethnopharmacological relevanceWuzi Yanzong Pill (WYP) is a classic traditional Chinese medicine (TCM) formula that is used for reproductive system diseases. Previous studies showed that WYP had a preventive effect on the development of neural tube defects (NTDs) induced by all-trans retinoic acid (atRA) in mice. Aim of the studyThis study aimed to determine the optimal combination of main monomer components in WYP on preventing NTDs and to understand the underlying mechanism. Materials and methodsAn optimal combination was made from five representative components in WYP including hyperoside, acteoside, schizandrol A, kaempferide and ellagic acid by orthogonal design method. In a mouse model of NTDs induced by intraperitoneal injection of atRA, pathological changes of neural tube tissues were observed by Hematoxylin & Eosin (HE) staining, neural tube epithelial cells apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), protein changes related to apoptosis, anti-apoptosis, and antioxidant factors were detected with Western blot. Potential targets and mechanisms of monomer compatibility group (MCG) acting on NTDs were analyzed by bioinformatics. ResultsTreatment with different combinations of WYP bioactive ingredients resulted in varying decreases in the incidence of NTDs in mice embryos. The combination of MCG15 (200 mg/kg of hyperoside, 100 mg/kg of acteoside, 10 mg/kg of schizandrol A, 100 mg/kg of kaempferide and 1 mg/kg of ellagic acid) showed the most significant reduction in NTD incidence. Mechanistically, MCG15 inhibited apoptosis and oxidative stress, as evidenced by reduced TUNEL-positive cells, downregulation of caspase-9, cleaved caspase-3, Bad, and Bax, and upregulation of Bcl-2, as well as decreased MDA and increased SOD, CAT, GSH, HO-1, and GPX1 levels. Bioinformatics analysis showed that MCG15 acted on the PI3K/Akt signaling pathway, which was confirmed by Western blot analysis showing increased expression of p-PI3K, p-Akt/Akt, and Nrf2 related indicators. ConclusionWe have identified an optimal combination of five bioactive components in WYP (MCG15) that prevented NTDs in mice embryos induced by atRA by activating the PI3K/Akt signaling pathway and inhibiting apoptosis and oxidative stress.

Intended and Unintended Benefits of Folic Acid Fortification-A Narrative Review.

Inadequate folate intake during pregnancy is the leading cause of the development of neural tube defects (NTDs) in newborns. For this reason, mandatory fortification of folic acid, a synthetic, easily bioavailable form, in processed cereals and cereal products has been implemented in the US since 1 January 1998 to reduce the risk of NTD in newborn children. This report aimed to review the literature related to the impact of mandated folic acid fortification on the intended and unintended benefits to health. Potential adverse effects were also discussed. We searched Pubmed, Google Scholar, Embase, SCOPUS, and Cochrane databases for reports. About 60 reports published between January 1998 and December 2022 were reviewed, summarized, and served as background for this review. The intended benefit was decreased prevalence of NTDs, while unintended benefits were reduction in anemia, blood serum homocysteine, and the risk of developing cardiovascular diseases. Potential issues with folic acid fortification are the presence of unmetabolized folic acid in circulation, increased risk of cancer, and the masking of vitamin B-12 deficiency. From a health perspective, it is important to monitor the impact of folic acid fortification periodically.

A Randomized Trial of Quadruple-Fortified Salt for Anemia and Birth Defects Prevention in Southern India: Protocol Design and Methods

BackgroundWomen of reproductive age are at an increased risk of anemia and micronutrient deficiencies. Evidence supports the role of periconceptional nutrition in the development of neural tube defects (NTDs) and other pregnancy complications. Vitamin B12 deficiency is a risk factor for NTDs and may modify folate biomarkers that predict NTD risk at the population level. There is an interest in mandatory fortification with vitamin B12 and folic acid for anemia and birth defect prevention. However, there are limited population-representative data needed to inform policy and guidelines. ObjectivesThis randomized trial will be conducted to evaluate the efficacy of quadruple-fortified salt (QFS; iron, iodine, folic acid, vitamin B12) in 1,000 households in Southern India. MethodsWomen 18 to 49 y who are not pregnant or lactating and reside within the catchment area of our community-based research site in Southern India will be screened and invited to participate in the trial. After informed consent, women and their households will be randomized to receive one of the following 4 interventions: 1) double-fortified salt (DFS; iron, iodine), 2) DFS + folic acid (iron, iodine, folic acid), 3) DFS + vitamin B12 (iron, iodine, vitamin B12), or 4) DFS + folic acid and vitamin B12 (QFS; iron, iodine, folic acid, vitamin B12) for 12 mo. Structured interviews will be conducted by trained nurse enumerators to collect sociodemographic, anthropometric, dietary, health, and reproductive history data. Biological samples will be collected at baseline, midpoint, and endpoint. Whole blood will be analyzed for hemoglobin using Coulter Counter. Total vitamin B12 will be measured by chemiluminescence; red blood cell folate and serum folate will be evaluated using the World Health Organization-recommended microbiologic assay. ConclusionsThe results of this randomized trial will help to evaluate the efficacy of QFS to prevent anemia and micronutrient deficiencies. Clinical trial registration numbers: NCT03853304 and Clinical Trial Registry of India REF/2019/03/024479. Registration numberNCT03853304 and REF/2019/03/024479.

Distinct H3K27me3 and H3K27ac Modifications in Neural Tube Defects Induced by Benzo[a]pyrene.

The pathological mechanisms of neural tube defects (NTDs) are not yet fully understood. Although the dysregulation of histone modification in NTDs is recognized, it remains to be fully elucidated on a genome-wide level. We profiled genome-wide H3K27me3 and H3K27ac occupancy by CUT&Tag in neural tissues from ICR mouse embryos with benzo[a]pyrene (BaP)-induced NTDs (250 mg kg-1) at E9.5. Furthermore, we performed RNA sequencing (RNA-seq) to investigate the regulation of histone modifications on gene expressions. Gene ontology and KEGG analysis were conducted to predict pathways involved in the development of NTDs. Our analysis of histone 3 lysine 27 modification in BaP-NTD neural tissues compared to BaP-nonNTD revealed 6045 differentially trimethylated regions and 3104 acetylated regions throughout the genome, respectively. The functional analysis identified a number of pathways uniquely enriched for BaP-NTD embryos, including known neurodevelopment related pathways such as anterior/posterior pattern specification, ephrin receptor signaling pathway, neuron migration and neuron differentiation. RNA-seq identified 423 differentially expressed genes (DEGs) between BaP-NTD and BaP-nonNTD group. The combination analysis of CUT&Tag and RNA-seq found that 55 DEGs were modified by H3K27me3 and 25 by H3K27ac in BaP-NTD, respectively. In the transcriptional regulatory network, transcriptional factors including Srsf1, Ume6, Zbtb7b, and Cad were predicated to be involved in gene expression regulation. In conclusion, our results provide an overview of histone modifications during neural tube closure and demonstrate a key role of genome-wide alterations in H3K27me3 and H3K27ac in NTDs corresponding with changes in transcription profiles.

Neural Tube Defects and Folate Deficiency: Is DNA Repair Defective?

Neural tube defects (NTDs) are complex congenital malformations resulting from failure of neural tube closure during embryogenesis, which is affected by the interaction of genetic and environmental factors. It is well known that folate deficiency increases the incidence of NTDs; however, the underlying mechanism remains unclear. Folate deficiency not only causes DNA hypomethylation, but also blocks the synthesis of 2'-deoxythymidine-5'-monophosphate (dTMP) and increases uracil misincorporation, resulting in genomic instabilities such as base mismatch, DNA breakage, and even chromosome aberration. DNA repair pathways are essential for ensuring normal DNA synthesis, genomic stability and integrity during embryonic neural development. Genomic instability or lack of DNA repair has been implicated in risk of development of NTDs. Here, we reviewed the relationship between folate deficiency, DNA repair pathways and NTDs so as to reveal the role and significance of DNA repair system in the pathogenesis of NTDs and better understand the pathogenesis of NTDs.

BMP/Smad Pathway Is Involved in Lithium Carbonate-Induced Neural-Tube Defects in Mice and Neural Stem Cells

Neural-tube defects (NTDs) are one type of the most serious birth defects. Studies have shown that inositol deficiency is closely related to the occurrence of NTDs. Bone morphogenetic protein (BMP)-mediated Smad signaling pathways have been implicated in neurogenesis and neural-tube closure. However, the role of the BMP/Smad pathway in inositol-deficiency-induced NTDs remains unclear. Inositol-deficiency models in C57 mice and mouse neural stem cells (mNSCs) were induced with Li2CO3 treatment or inositol withdrawal. The role of the BMP/Smad pathway in the regulation of cell proliferation and the development of NTDs was determined utilizing qRT-PCR, HE staining, Western blot, immunostaining, MTT assay, EdU staining, and flow cytometry. The intraperitoneal injection of Li2CO3 at Embryonic Day 7.5 induced the occurrence of NTDs. The mRNA levels of Bmp2, Bmp4, Smad1, Smad5, Smad8 and Runx2, the phosphorylation of Smad1/5/8, and the nuclear translocation of Runx2 were significantly increased in NTD embryonic brain tissues and mNSCs exposed to Li2CO3 or an inositol-free medium, which were suppressed by BMP receptor selective inhibitor LDN-193189. The Li2CO3-induced phosphorylation of Smad1/5/8 was inhibited by inositol supplementation. Cell proliferation was significantly promoted by Li2CO3 exposure or the absence of inositol in mNSCs, which was reversed by LDN-193189. These results suggest that the activation of the BMP/Smad signaling pathway might play an important role in the development of NTDs induced by maternal Li2CO3 exposure via inositol deficiency.

Identifying key m6A-methylated lncRNAs and genes associated with neural tube defects via integrative MeRIP and RNA sequencing analyses.

Objective: N6-methyladenosine (m6A) is a common post-transcriptional modification of messenger RNAs (mRNAs) and long non-coding RNAs (lncRNAs). However, m6A-modified lncRNAs are still largely unexplored. This study aimed to investigate differentially m6A-modified lncRNAs and genes involved in neural tube defect (NTD) development. Methods: Pregnant Kunming mice (9-10 weeks of age) were treated with retinoic acid to construct NTD models. m6A levels and methyltransferase-like 3 (METTL3) expression were evaluated in brain tissues of the NTD models. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq) were performed on the NovaSeq platform and Illumina HiSeq 2,500 platform, respectively. Differentially m6A-methylated differentially expressed lncRNAs (DElncRNAs) and differentially expressed genes (DEGs) were identified, followed by GO biological process and KEGG pathway functional enrichment analyses. Expression levels of several DElncRNAs and DEGs were evaluated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for validation. Results: m6A levels and METTL3 expression levels were significantly lower in the brain tissues of the NTD mouse model than in controls. By integrating MeRIP-seq and RNA-seq data, 13 differentially m6A-methylated DElncRNAs and 170 differentially m6A-methylated DEGs were identified. They were significantly enriched in the Hippo signaling pathway and mannose-type O-glycan biosynthesis. The qRT-PCR results confirmed the decreased expression levels of lncRNAs, such as Mir100hg, Gm19265, Gm10544, and Malat1, and genes, such as Zfp236, Erc2, and Hmg20a, in the NTD group. Conclusion: METTL3-mediated m6A modifications may be involved in NTD development. In particular, decreased expression levels of Mir100hg, Gm19265, Gm10544, Malat1, Zfp236, Erc2, and Hmg20a may contribute to the development of NTD.

Single nucleotide polymorphisms of PCP pathway related genes participate in the occurrence and development of neural tube defect.

To screen the single nucleotide polymorphisms (SNPs) in the coding regions of VANGL and FZD family members related to the plane cell polarity (PCP) signaling pathway in neural tube defects (NTDs) patients, so as to provide theoretical and experimental basis for the prevention and treatment of NTDs by intervening PCP signal transduction. 112 NTDs patients were collected as the case group and 112 craniocerebral trauma patients as control. Afterwards, blood genomic DNA was extracted and sequenced. The distribution of SNP alleles and genotypes between case and control groups was analyzed. Finally, the NTD rat model was constructed, and the effect of SNPs on the expression level of VANGL and FZD genes was verified by qRT-PCR. GC genotype was newly found at VANGL1 c.346G>A, as well as AT genotype in FZD6 c.97A>G. The distribution of VANGL1 c.346g>A allele and genotype was statistically different between the case and control groups (p < 0.05). The newly found genotype GC increased the risk of NTDs (OR=9.918, 95% CI: 1.234%-79.709%). The results of qRT-PCR showed that the expression level of FZD6 in E11 NTD fetuses were significantly increased (p < 0.05), but there was no obvious difference in the expression of VANGL1. We found a new variant of VANGL1 c.346G>A, whose GC genotype might play an important role in the pathogenesis of NTDs. The SNPs of VANGL1 had no significant effect on its expression level, indicating that it may induce NTDs through other ways. FZD6 was significantly overexpressed in NTDs fetuses.

On paleoepidemiology of spina bifida sacralis: prevalence of the anomaly in Late Scythians of lower Dnieper region

Spina bifida is a developmental anomaly that is thought to be caused by a combination of hereditary and en-vironmental factors. Though the most significant association was found with the folic acid deficit during early em-bryogenesis, numerous genetic variants were also found to be in some association with the development of neu-ral tube defects, but this data is inconsistent. It is still debated whether the unremarkable anomaly (spina bifida occulta) often observed in osteological samples is truly a form of neural tube defects forming in early embryo-genesis, or is a minor variation that is forming later in postnatal life. Also, data is equivocal regarding the neuro-logical consequences of sacral spina bifida occulta, some pointing to its clinical significance, others stating that it does not affect the wellbeing in any way. Though paleopathological studies on spina bifida are numerous, the frequency of the anomaly remains largely unknown for the Early Iron Age population of the Eurasian steppes. This study examines presence of sacral spina bifida in a Late Scynthian sample (3rd c. BC — 3rd с. AD) from the northern Black Sea region. Totally, 89 skeletons were examined, originating from cemeteries near Nikolayevka and Zolotaya Balka villages (Kherson Oblast, Ukrain). These cemeteries are located along the Dnieper river bank about 90 km apart. The people they represent were settled agriculturalists. The defect was scored when either or all of the three upper sacral vertebrae were affected (S1–S3), its level was recorded. Totally, 16 % of individuals were affected. The majority of defects were observed solely on S1 level. Only two cases of spina bifida sacralis totalis were recorded. The frequency of the anomaly was not significantly different either between males and fe-males, or between younger (&lt;35 years) and older (&gt;35 years) adults. In two instances, two individuals buried in a common grave both had spina bifida sacralis which may point to a biological relationship between them. The fre-quency of spina bifida sacralis is very close to the mean values, reported in the literature. Thus, no evidence of an increased selection against individuals in this group with this form of skeletal anomaly, or a significant impact of envi-ronmental factors, leading to its development in ontogenesis, is present. Also, contrary to literature data, it seems that in this sample age and sex were not the factors significantly contributing to the trait’s variation, though the age dynamics of this trait needs further consideration using a sample with higher proportion of senile individuals.

Cilia and their role in neural tube development and defects

Abstract Cilia are microtubule-based filamentous organelles that play a vital role in embryogenesis. Multiple signal transduction pathways are orchestrated by cilia, such as Hedgehog and planar cell polarity signals. Various studies, spanning over last 2 decades, have emphasized the role of cilia-mediated signaling cascades in regulating neural tube patterning and development. Moreover, the deficiency of certain ciliary genes have been reported to cause neural tube defects (NTDs), which are a set of disorders that occur due to perturbation of normal neural tube closure. However, the mechanisms underlying cilia dysfunction resulting NTDs remain unclear. Recent studies have highlighted the association of phosphoinositide signaling with cilia, thereby conferring novel insights into the function of cilia during neural tube development. Here, we have reviewed recent studies on cilia, focusing on the molecular mechanism underlying the involvement of cilia in neural tube development and the role of ciliary disruption in the development of NTDs.

Metabolic Analysis of Methylenetetrahydrofolate Reductase Single Nucleotide Polymorphisms (MTHFR 677C

Neural tube defects (NTDs)are among the most prevalent and debilitating birth defects with their causes are still unknown, despite mounting evidence that genetic and/or environmental factors may play a role. We aimed to analyze two single nucleotide polymorphisms of methylenetetrahydrofolate reductase (MTHFR) gene, serum folate and vitamin B12 status among a cohort of Egyptian children with NTDs and their mothers. A case-control study has been conducted on 50 Egyptian children with various types of NTDs and their mothers. They were comparable with 50 unrelated healthy, age and sex matched children and their mothers (50) selected as controls. Pediatric and neurosurgical assessments were performed to the included cases. Serum folate and vitamin B12 were measured using ELISA kits. MTHFR 677C<T (rs1801133) and MTHFR1298A<C (rs1801131) were analyzed by restriction fragment length polymorphism using polymerase chain reaction. Lumbosacral meningomyelocele was the most frequent NTDs (50%). Significantly lower serum folate and vitamin B12 among cases and case mothers compared to the control and control mothers (p<0.05 for all). Significantly higher frequencies of both heterozygous mutant (CT) and homozygous mutant (TT) genotypes, and mutant T allele of MTHFR 677C<T among case mothers compared to control mothers (p<0.05 for all), with lack of significant differences of this SNP between pediatric groups. Mutant homozygous (AA) genotype and mutant A allele of MTHFR 1298A<C among control mothers were significantly frequent compared to case mothers (p<0.05 for both), with OR 6.081 and 7.071, [95%CI were 3.071-11.287 and 3.296-15.172, respectively]. Significantly frequent wild homozygous (CC) genotype and normal C allele of MTHFR 1298A<C among children with NTDs compared to the controls (p<0.05 for both), with OR 0.231 and 0.754, [95%CI were 0.095-0.561and 0.432-1.317, respectively].Low serum folate and B12 are frequently common among children with NTDs and their mothers. MTHFR 677C<T in mothers could be considered as genetic risk factors for development of NTDs in their children, while MTHFR1298A<C could be protective genetic factor against NTDs development.

Identification of immune-related biomarkers in embryos with neural tube defects via a bioinformatics analysis.

BackgroundNeural tube defects (NTDs) are one of the most common types of birth defects. Oral folic acid (FA) prophylaxis is currently available, but the pathogenesis of NTDs is not fully understood. We conducted this study to examine the role of the immune landscape of NTDs and identify novel diagnostic and therapeutic biomarkers.MethodsWe downloaded the GSE33111 data set of 12 NTD embryos and 12 healthy embryos in the same period of fetal development from the Gene Expression Omnibus (GEO) database. We compared the healthy embryos and NTD embryos to identify the differentially expressed genes (DEGs). We also performed a functional enrichment analysis of the DEGs using the clusterProfiler package. We extracted the top 10 ranked genes as hub immune-related biomarkers. We then used receiver operating characteristic (ROC) curves to determine the expression levels of the hub immune-related genes and the accuracy of the diagnosis of NTDs. Finally, we analyzed the immune landscape of the NTD embryos and healthy embryos via a single-sample gene set enrichment analysis (ssGSEA).ResultsA total of 611 DEGs were identified by the differential analysis, including 95 immune genes. The functional enrichment analysis indicated that Epstein-Barr virus infection, antigen processing and presentation, inflammatory bowel disease, and type I diabetes mellitus were associated with NTDs. The results of the expression level analysis showed that the hub immune-related genes were more highly expressed in the NTD embryos than the healthy embryos. Additionally, the ROC curve analysis also indicated that the expression levels of the 10 hub immune-related genes were highly accurate in the diagnosis of NTDs [area under the curve (AUC) range, 0.708–0.812]. The immune infiltration analysis demonstrated that 20 of the 28 immune cell types were more highly infiltrated in the NTD embryos than the healthy embryos.ConclusionsImmune-related genes are important regulators of the occurrence and development of NTDs.