Hindbrain neuropore tissue geometry determines asymmetric cell-mediated closure dynamics in mouse embryos.

Gap closure is a common morphogenetic process. In mammals, failure to close the embryonic hindbrain neuropore (HNP) gap causes fatal anencephaly. We observed that surface ectoderm cells surrounding the mouse HNP assemble high-tension actomyosin purse strings at their leading edge and establish the initial contacts across the embryonic midline. Fibronectin and laminin are present, and tensin 1 accumulates in focal adhesion-like puncta at this leading edge. The HNP gap closes asymmetrically, faster from its rostral than caudal end, while maintaining an elongated aspect ratio. Cell-based physical modeling identifies two closure mechanisms sufficient to account for tissue-level HNP closure dynamics: purse-string contraction and directional cell motion implemented through active crawling. Combining both closure mechanisms hastens gap closure and produces a constant rate of gap shortening. Purse-string contraction reduces, whereas crawling increases gap aspect ratio, and their combination maintains it. Closure rate asymmetry can be explained by asymmetric embryo tissue geometry, namely a narrower rostral gap apex, whereas biomechanical tension inferred from laser ablation is equivalent at the gaps' rostral and caudal closure points. At the cellular level, the physical model predicts rearrangements of cells at the HNP rostral and caudal extremes as the gap shortens. These behaviors are reproducibly live imaged in mouse embryos. Thus, mammalian embryos coordinate cellular- and tissue-level mechanics to achieve this critical gap closure event.

Reduced H3K27me3 leads to abnormal Hox gene expression in neural tube defects.

BACKGROUND: Neural tube defects (NTDs) are severe, common birth defects that result from failure of normal neural tube closure during early embryogenesis. Accumulating strong evidence indicates that genetic factors contribute to NTDs etiology, among them, HOX genes play a key role in neural tube closure. Although abnormal HOX gene expression can lead to NTDs, the underlying pathological mechanisms have not fully been understood. METHOD: We detected that H3K27me3 and expression of the Hox genes in a retinoic acid (RA) induced mouse NTDs model on E8.5, E9.5 and E10.5 using RNA-sequencing and chromatin immunoprecipitation sequencing assays. Furthermore, we quantified 10 Hox genes using NanoString nCounter in brain tissue of fetuses with 39 NTDs patients including anencephaly, spina bifida, hydrocephaly and encephalocele. RESULTS: Here, our results showed differential expression in 26 genes with a > 20-fold change in the level of expression, including 10 upregulated Hox genes. RT-qPCR revealed that these 10 Hox genes were all upregulated in RA-induced mouse NTDs as well as RA-treated embryonic stem cells (ESCs). Using ChIP-seq assays, we demonstrate that a decrease in H3K27me3 level upregulates the expression of Hox cluster A-D in RA-induced mouse NTDs model on E10.5. Interestingly, RA treatment led to attenuation of H3K27me3 due to cooperate between UTX and Suz12, affecting Hox gene regulation. Further analysis, in human anencephaly cases, upregulation of 10 HOX genes was observed, along with aberrant levels of H3K27me3. Notably, HOXB4, HOXC4 and HOXD1 expression was negatively correlated with H3K27me3 levels. CONCLUSION: Our results indicate that abnormal HOX gene expression induced by aberrant H3K27me3 levels may be a risk factor for NTDs and highlight the need for further analysis of genome-wide epigenetic modification in NTDs.

Gangliosidome of human anencephaly: A high resolution multistage mass spectrometry study.

Widely dispersed throughout the entire body tissues, gangliosides (GGs) are essential components of neuronal cell membranes, where exhibit a vital role in neuronal function and brain development, directly influencing the neural tube formation, neurogenesis, neurotransmission, etc. Due to several factors, partial or complete closing faults of the fetal neural tube may occur in the first trimester of pregnancy, generating a series of neural tube defects (NTD), among which anencephaly. The absence in anencephaly of the forebrain and skull bones determines the exposure to the amniotic fluid of the remaining brain tissue and the spinal cord, causing the degeneration of the nervous system tissue. Based on the previously achieved information related to the direct alteration of neural development with deficient concentration of several GGs, a systematic and comparative mass spectrometry (MS) mapping assay on GGs originating from fetuses in different intrauterine developmental stages, i.e. the 29th (denoted An29), 35th (An35) and the 37th (An37) gestational weeks was here conducted. Our approach, based on Orbitrap MS under high sensitivity, resolution and mass accuracy conditions, enabled for the first time the nanoelectrospray ionization, detection and identification of over 150 glycoforms, mainly novel, polysialylated species. Such a pattern, specific for incipient developmental stages reliably documents the brain development stagnation, characteristic for anencephaly. Further, the fragmentation MS2-MS3 experiments by collision induced dissociation (CID) confirmed the incidence in all three samples of GT2(d18:1/16:2) as a potential biomarker. Therefore, this fingerprinting of the anencephalic gangliosidome may serve in development of approaches for routine screening and early diagnosis.

Neural tube closure and embryonic metabolism.

Neural tube closure (NTC) is an embryonic process during formation of the mammalian central nervous system. Disruption of the dynamic, sequential events of NTC can cause neural tube defects (NTD) leading to spina bifida and anencephaly in the newborn. NTC is affected by inherent factors such as genetic mutation or if the mother is exposed to certain environmental factors such as intake of harmful chemicals, maternal infection, irradiation, malnutrition, and inadequate or excessive intake of specific nutrients. Although effects of these stress factors on NTC have been intensively studied, the metabolic state of a normally developing embryo remains unclear. State-of-the art mass spectrometry techniques have enabled detailed study of embryonic metabolite profiles and their distribution within tissues. This approach has demonstrated that glucose metabolism is altered during NTC stages involving chorioallantoic branching. An understanding of embryonic metabolic rewiring would help reveal the etiology of NTD caused by environmental factors.

Nutrition, One-Carbon Metabolism and Neural Tube Defects: A Review.

Neural tube defects (NTDs) are a group of severe congenital malformations, induced by the combined effects of genes and the environment. The most valuable finding so far has been the protective effect of folic acid supplementation against NTDs. However, many women do not take folic acid supplements until they are pregnant, which is too late to prevent NTDs effectively. Long-term intake of folic acid-fortified food is a good choice to solve this problem, and mandatory folic acid fortification should be further promoted, especially in Europe, Asia and Africa. Vitamin B2, vitamin B-6, vitamin B-12, choline, betaine and n-3 polyunsaturated fatty acids (PUFAs) can also reduce the NTD risk by interacting with the one-carbon metabolism pathway. This suggest that multivitamin B combined with choline, betaine and n-3 PUFAs supplementation may have a better protective effect against NTDs than folic acid alone. Genetic polymorphisms involved in one-carbon metabolism are associated with NTD risk, and gene screening for women of childbearing age prior to pregnancy may help prevent NTDs induced by the risk allele. In addition, the consumption of alcohol, tea and coffee, and low intakes of fruit and vegetable are also associated with the increased risk of NTDs, and should be avoided by women of childbearing age.

Modeling anterior development in mice: diet as modulator of risk for neural tube defects.

Head morphogenesis is a complex process that is controlled by multiple signaling centers. The most common defects of cranial development are craniofacial defects, such as cleft lip and cleft palate, and neural tube defects, such as anencephaly and encephalocoele in humans. More than 400 genes that contribute to proper neural tube closure have been identified in experimental animals, but only very few causative gene mutations have been identified in humans, supporting the notion that environmental influences are critical. The intrauterine environment is influenced by maternal nutrition, and hence, maternal diet can modulate the risk for cranial and neural tube defects. This article reviews recent progress toward a better understanding of nutrients during pregnancy, with particular focus on mouse models for defective neural tube closure. At least four major patterns of nutrient responses are apparent, suggesting that multiple pathways are involved in the response, and likely in the underlying pathogenesis of the defects. Folic acid has been the most widely studied nutrient, and the diverse responses of the mouse models to folic acid supplementation indicate that folic acid is not universally beneficial, but that the effect is dependent on genetic configuration. If this is the case for other nutrients as well, efforts to prevent neural tube defects with nutritional supplementation may need to become more specifically targeted than previously appreciated. Mouse models are indispensable for a better understanding of nutrient-gene interactions in normal pregnancies, as well as in those affected by metabolic diseases, such as diabetes and obesity.

Amniotic fluid and serum biomarkers from women with neural tube defect-affected pregnancies: a case study for myelomeningocele and anencephaly: clinical article.

OBJECT: The authors sought to identify novel biomarkers for early detection of neural tube defects (NTDs) in human fetuses. METHODS: Amniotic fluid and serum were drawn from women in the second trimester of pregnancy. The study group included 2 women pregnant with normal fetuses and 4 with fetuses displaying myelomeningocele (n = 1), anencephaly (n = 1), holoprosencephaly (n = 1), or encephalocele (n = 1). Amniotic fluid stem cells (AFSCs) were isolated and cultured. The cells were immunostained for the stem cell markers Oct4, CD133, and Sox2; the epigenetic biomarkers H3K4me2, H3K4me3, H3K27me2, H3K27me3, H3K9Ac, and H3K18Ac; and the histone modifiers KDM6B (a histone H3K27 demethylase) and Gcn5 (a histone acetyltransferase). The levels of 2 markers for neural tube development, bone morphogenetic protein-4 (BMP4) and sonic hedgehog (Shh), were measured in amniotic fluid and serum using an enzyme-linked immunosorbent assay. RESULTS: The AFSCs from the woman pregnant with a fetus affected by myelomeningocele had higher levels of H3K4me2, H3K4me3, H3K27me2, and H3K27me3 and lower levels of KDM6B than the AFSCs from the women with healthy fetuses. The levels of H3K9ac, H3K18ac, and Gcn5 were also decreased in the woman with the fetus exhibiting myelomeningocele. In AFSCs from the woman carrying an anencephalic fetus, levels of H3K27me3, along with those of H3K9Ac, H3K18ac, and Gcn5, were increased, while that of KDM6B was decreased. Compared with the normal controls, the levels of BMP4 in amniotic fluid and serum from the woman with a fetus with myelomeningocele were increased, whereas levels of Shh were increased in the woman pregnant with a fetus displaying anencephaly. CONCLUSIONS: The levels of epigenetic marks, such as H3K4me, H3K27me3, H3K9Ac, and H3K18A, in cultured AFSCs in combination with levels of key developmental proteins, such as BMP4 and Shh, are potential biomarkers for early detection and identification of NTDs in amniotic fluid and maternal serum.

Association study of PARD3 gene polymorphisms with neural tube defects in a Chinese Han population.

Partitioning defective 3 homolog (PARD3) is an attractive candidate gene for screening neural tube defect (NTD) risk. To investigate the role of genetic variants in PARD3 on NTD risk, a case-control study was performed in a region of China with a high prevalence of NTDs. Total 53 single-nucleotide polymorphisms (SNPs) in PARD3 were genotyped in 224 fetuses with NTDs and in 253 normal fetuses. We found that 6 SNPs (rs2496720, rs2252655, rs3851068, rs118153230, rs10827337, and rs12218196) were statistically associated with NTDs (P < .05). After stratifying participants by NTD phenotypes, the significant association only existed in cases with anencephaly rather than spina bifida. Further haplotype analysis confirmed the association between PARD3 polymorphisms and NTD risk (global test P = 3.41e-008). Our results suggested that genetic variants in PARD3 were associated with susceptibility to NTDs in a Chinese Han population, and this association was affected by NTD phenotypes.

Acrania-anencephaly associated with hypospadias. Prenatal ultrasound and MRI diagnosis and molecular folate metabolism pathway analysis.

Acrania may occur as a single isolated malformation or associated with extracranial defects. Hypospadias is one of the most common congenital abnormalities of the genitalia frequently missed on prenatal sonograms. Second trimester two- and three-dimensional ultrasound and MRI diagnosis with necropsy and folate metabolism pathway analysis. The mechanisms leading to closure of both neural and urethral tubes, are far from being demonstrated, and molecular studies of this very rare association are lacking although it might be based on a common genetic mechanism, leading to a disturbed development pathway at the molecular level.

Differential gene expression in the adrenals of normal and anencephalic fetuses and studies focused on the Fras-1-related extracellular matrix protein (FREM2) gene.

PRECIS: Many genes are differentially expressed in normal compared to anencephalic human fetal adrenals (HFAs), especially the Fras-1-related extracellular matrix protein (FREM2) gene. FREM2 expression appears to be regulated by adrenocorticotrophic hormone (ACTH). CONTEXT: The expression profiles of genes responsible for cortical growth and zonation in the HFA gland are poorly characterized. The neural tube disorder anencephaly is associated with fetal adrenal hypoplasia with a large size reduction of the fetal zone of the HFA. OBJECTIVE: To determine gene expression profile differences in the adrenals of anencephalic compared to normal HFAs to identify genes that may play important roles in adrenal development. DESIGN AND METHODS: Fresh tissues were obtained at the time of autopsy from normal and anencephalic human fetuses delivered at mid-gestation. The following techniques were used: cell culture, messenger RNA (mRNA) extraction, microarray analysis, complementary DNA (cDNA) synthesis, quantitative real-time reverse transcriptase polymerase chain reaction (QT-PCR). RESULTS: We identified over 40 genes expressed at levels 4-fold or greater in the normal versus anencephalic HFAs and that 28 genes were expressed at increased levels in the anencephalic HFA. The expression of FREM2 at approximately 40-fold greater levels in the normal HFA compared to the HFA of anencephalic fetuses was confirmed by QT-PCR. Expression of FREM2 in the kidney was not significantly different between normal and anencephalic fetuses. In cultured HFA cells, ACTH treatment for 48 hours increased the expression of FREM2 and a gene responsive to ACTH, CYP17, but not tyrosine hydroxylase. CONCLUSIONS: Abnormal expression of many genes may be involved in the adrenal hypoplasia seen in anencephaly. FREM2 appears to be regulated by ACTH and is the most differentially expressed gene, which may be important in the development and function of the HFA, particularly the fetal zone of the HFA.

The human placenta expresses CYP17 and generates androgens de novo.

INTRODUCTION: The human placenta is believed to have insignificant CYP17 expression, rendering it dependent on the maternal and fetal compartments for the necessary androgenic precursors to yield the high levels of estrogens seen in pregnancy. OBJECTIVE: The aim of the study was to analyze whether the human trophoblast is capable of expressing CYP17 and producing androgens de novo. METHODS: Human trophoblasts from fresh placentas and JEG-3 cells were used for all experiments. CYP17 mRNA analysis was performed via RT-PCR, and protein detection by Western blot and immunohistochemical staining. Steroid products were quantified using RIAs. RESULTS: CYP17 mRNA was expressed in both cell types. CYP17 protein was detected by Western blotting and localized by immunostaining mainly to the cytoplasm of syncytiotrophoblasts. Measurement of 17α-hydroxyprogesterone, androstenedione, and their aromatized products in the media further demonstrated CYP17 expression and activity in the human trophoblast. Baseline levels of CYP17 steroid products were higher in primary cells and significantly increased in the presence of 22-hydroxycholesterol. CONCLUSIONS: We have demonstrated CYP17 mRNA and protein expression and activity in human trophoblasts. Considering the precursor concentration, blood flow, and mass of the placenta, we suggest that its contribution of androgens is an important source of estrogen production in pregnancy.

MicroRNAs: potential regulators involved in human anencephaly.

MicroRNAs (miRNAs) are posttranscriptional regulators of messenger RNA activity. Neural tube defects (NTDs) are severe congenital anomalies that substantially impact an infant's morbidity and mortality. The miRNAs are known to be dynamically regulated during neurodevelopment; their role in human NTDs, however, is still unknown. In this study, we show the presence of a specific miRNA expression profile from tissues of fetuses with anencephaly, one of the most severe forms of NTDs. Furthermore, we map the target genes of these miRNAs in the human genome. In comparison to healthy human fetal brain tissues, tissues from fetuses with anencephaly exhibited 97 down-regulated and 116 up-regulated miRNAs. The microarray findings were extended using real-time qRT-PCR for nine miRNAs. Specifically, of these validated miRNAs, miR-126, miR-198, and miR-451 were up-regulated, while miR-9, miR-212, miR-124, miR-138, and miR-103/107 were down-regulated in the tissues of fetuses with anencephaly. A bioinformatic analysis showed 881 potential target genes that are regulated by the validated miRNAs. Seventy-nine of these potential genes are involved in a protein interaction network. There were 6 co-occurrence annotations within the GOSlim process and 7 co-occurrence annotations within the GOSlim function found by GeneCodis 2.0. Our results suggest that miRNA dysregulation is possibly involved in the pathogenesis of anencephaly.

Abnormal folate metabolism in foetuses affected by neural tube defects.

Folic acid supplementation can prevent many cases of neural tube defects (NTDs), whereas suboptimal maternal folate status is a risk factor, suggesting that folate metabolism is a key determinant of susceptibility to NTDs. Despite extensive genetic analysis of folate cycle enzymes, and quantification of metabolites in maternal blood, neither the protective mechanism nor the relationship between maternal folate status and susceptibility are understood in most cases. In order to investigate potential abnormalities in folate metabolism in the embryo itself, we derived primary fibroblastic cell lines from foetuses affected by NTDs and subjected them to the dU suppression test, a sensitive metabolic test of folate metabolism. Significantly, a subset of NTD cases exhibited low scores in this test, indicative of abnormalities in folate cycling that may be causally linked to the defect. Susceptibility to NTDs may be increased by suppression of the methylation cycle, which is interlinked with the folate cycle. However, reduced efficacy in the dU suppression test was not associated with altered abundance of the methylation cycle intermediates, s-adenosylmethionine and s-adenosylhomocysteine, suggesting that a methylation cycle defect is unlikely to be responsible for the observed abnormality of folate metabolism. Genotyping of samples for known polymorphisms in genes encoding folate-associated enzymes did not reveal any correlation between specific genotypes and the observed abnormalities in folate metabolism. These data suggest that as yet unrecognized genetic variants result in embryonic abnormalities of folate cycling that may be causally related to NTDs.

Development of the human retina in the absence of ganglion cells.

Retinal development was studied in eyes from fetal and neonatal human anencephalic (AnC) and normal age-matched infants to determine the time of retinal ganglion cell (GC) loss and its effect on the development of other retinal neurons. At fetal week (Fwk) 14, GC loss was evident in central retina and by Fwk 19-20 almost all GC were absent, although immunocytochemical labeling for GC markers brain 3, neurofilament M and parvalbumin detected a few GC in the AnC far periphery at older ages. The inner nuclear and inner plexiform (IPL) layers showed variable amounts of thinning but all normal bipolar (BP) and horizontal cell markers were still present. The amacrine (AM) labels calbindin and calretinin were markedly reduced. Lamination for these markers in the IPL was less organized than in normal retinas, with BP and AM markers extending into the degenerated GC layer. Cone and rod photoreceptors had normal morphology and topography in AnC retina and each expressed normal phototransduction and synaptic markers. The prospective fovea was identified in AnC neonatal retina by cone packing and the absence of immunolabeled rod photoreceptors. In one AnC neonatal retina, blood vessels and astrocytes extended across the inner retina in the putative fovea and there was no evidence of a pit. In another AnC neonatal retina, blood vessels and astrocytes formed a foveal avascular zone in the inner retina and a shallow pit was present within this zone. However, both foveas showed evidence for the onset of cone elongation and packing. These findings support the model of Springer and Hendrickson [2005; Vis. Neurosci. 22, 171] in which the foveal avascular zone is critical for pit formation, but suggest that mechanisms inherent to the outer retina may be involved in early stages of foveal cone packing.

Immunocytochemical study of the GH cells in the anterior pituitary gland of human fetus II. Anencephalic fetus.

In order to elucidate the effects of hypothalamic regulation on the morphology of GH cells, light and electron microscopic immunocytochemical examinations were carried out comparing GH cells in the anterior pituitary gland of anencephalic fetus with those of normal fetuses. Three types of GH cells were identified in the anterior pituitary gland of anencephalic fetus as well as in the normal fetus. Type-I is a small, round cell containing a few small secretory granules. Type-III is a large, polygonal cell with numerous large secretory granules. Type-II is a polygonal cell with medium-sized secretory granules. The Type-II GH cell was predominant in both anencephalic and normal fetuses. The most striking difference between anencephalic and normal fetuses was the presence of atypical forms of the Type II cell. These were polygonal cells containing secretory granules, which were either immunopositive or immunonegative to anti-human GH (anti-hGH) serum. Furthermore, two other types of GH cells were identified. The somatomammotroph (SM cell) contained GH and PRL in different granules within the same cell. Also, a different type of the GH cell was noted containing two varieties of secretory granules; one was immunolabeled only with anti-hGH and the other was not immunolabeled to either anti-hGH or anti-human PRL (anti-hPRL). From these results, we suggest that an absence of hypothalamic regulation in the anencehpalic does not seriously modify GH cell morphology but induces an altered GH storage pattern in some of the cells.

Anencephaly: structural characterization of gangliosides in defined brain regions.

Gangliosides from histopathologically-defined human cerebrum-resembling remnant and cerebellum from 37 and 30 gestational week-old anencephaluses were identified using mass spectrometry and high performance thin layer chromatography combined with immunochemical analysis in comparison to respective normal newborn/fetal and adult brain regions. A novel strategy of nano-electrospray ionization quadrupole time-of-flight tandem MS has been developed for identification of ganglioside components in complex mixtures. By morphoanatomical and histological investigation the anencephalic cerebral remnant was found to be aberrant, while the anencephalic cerebellum was defined as normal. Total ganglioside concentrations in the anencephalic cerebral remnant and the cerebellum were 34% and 13% lower in relation to the age-matched controls. In the cerebral remnant, GD3, GM2 and GT1b were elevated, while GD1a was decreased in the anencephalic cerebral remnant, but enriched in anencephalic cerebellum. GQ1b was reduced in both anencephalic regions. Gg4Cer, GM1b and GD1alpha, members of the alpha-series biosynthetic pathway, and neolacto-series gangliosides were found to be present in anencephalic, as well as in normal, fetal and adult brain tissues, indicating the occurrence of these biosynthetic pathways in human brain. In both cerebral and cerebellar anencephalic tissues, GM1b, GD1alpha, nLM1 and nLD1 were expressed at a higher rate in relation to normal tissue. It can be demonstrated that the anencephalic cerebral remnant, as a primitive brain structure, represents a naturally-occurring model to study the ganglioside involvement in induction of aberrant brain development.

Reduced expression of CD99 and functional disturbance in anencephalic cortical thymocytes.

In a significant proportion of cases, anencephaly is associated with thymic enlargement, suggesting a possibility that anencephalic fetuses have a functional disturbance in thymocyte differentiation and development. In this report, we demonstrated that CD99 expression was consistently reduced in cortical thymocytes of all anencephalic fetuses. In addition, the CD99-dependent aggregation of immature cortical thymocytes was almost completely impaired and apoptosis of thymocytes was markedly reduced in several cases. These results are in agreement with previous findings that CD99 regulates the aggregation and apoptosis of various types of cells. These data strongly suggest that functional disturbance of thymocytes and thymic hyperplasia are related to the reduced expression of CD99 molecule in anencephalic fetuses.

Mid-second trimester organic acids in severe open neural tube defects.

OBJECTIVE: To investigate amnioic fluid organic concentrations at 16 weeks gestation from normal fetuses and those affected by severe neural tube defects. METHODS: Amniotic fluid collected from normal, anencephalic and severe open spina bifida fetuses was analysed for up to 24 organic acids by gas liquid chromatography. RESULTS: Significant increase in organic acids similar to those observed in defects of phenylalanine and tyrosine metabolism were identified when compared with the normal fetuses. Significant differences in organic acid concentrations were also identified within the two neural tube defect groups. CONCLUSION: A relationship between phenylalanine and tyrosine metabolism, closure rate of the neural tube and folate metabolism is proposed.

Pulmonary gastrin-releasing peptide expression in anencephaly.

Gastrin-releasing peptide (GRP) is a developmentally regulated bioactive peptide believed to function as a pulmonary growth factor. It is produced by pulmonary neuroendocrine cells, found within the conducting and respiratory epithelium, as isolated cells and in clusters known as neuroepithelial bodies (NEBs). Deficient GRP expression has been reported in pulmonary hypoplasia (PH) associated with oligohydramnios and diaphragmatic hernia. To assess further the role of GRP in maldeveloped lung we reviewed the postmortem records and histologic lung sections, stained with H&E and anti-GRP antiserum, from 11 infants with anencephaly and 11 age-matched controls. Cells immunoreactive for GRP were quantified (isolated versus NEBs) in airways and airspaces per mm2 for a standard area. PH was present in five anencephalic infants. There was no difference in the total number of GRP-positive cells, number of NEBs, size of NEBs, or number of GRP-positive cells in airways or alveoli in either group regardless of lung development. A greater proportion of the GRP-positive cells was present in the airways in anencephalic infants with PH (58%) compared with anencephalic infants without PH (40%) (P = .018). There were no differences when comparing these groups with control infants and no differences in the density of airways in each of these groups. We conclude that deficient GRP expression is not a feature of lung hypoplasia in anencephalic infants. The altered distribution of GRP-positive cells in anencephalic infants with PH may be a reflection of the structural abnormalities or accompanying altered cellular maturity.

Gangliosides of anencephalic and fetal brain--immunostaining on thin-layer chromatograms.

The expression of gangliosides of ganglio-series as well as neolacto-series gangliosides in anencephalic and in normal human fetal brain was compared with that in adult brain by immunostaining on thin-layer chromatograms. A difference in the expression of ganglio-series gangliosides with GM1a core was found between anencephalic and normal fetal brain, with less expression of GM1a and GD1a in anencephaly compared with normal fetal brain, in which these gangliosides dominate. Small amounts of GM1b were detected in fetal brain whereas only traces were found in anencephalic brain. Lactosamine-containing gangliosides were present in fetal and in anencephalic brain as alpha 2-3 as well as alpha 2-6 sialylated nLcOse4Cer structures. A heterogeneous group of neolacto-series gangliosides was expressed in anencephalic brain in both the monosialo- and presumed disialoganglioside range. These findings demonstrate a significant change in ganglioside pattern in anencephaly where the process of cell differentiation and maturation has been severely disturbed.