Unknown,Transcriptomics,Genomics,Proteomics

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Perturbed gastrulation in embryos from diabetic pregnancies.

ABSTRACT: Maternal diabetes is a teratogen that can lead to neural tube closure defects in the offspring. In neurulation-stage embryos from diabetic dams, we detected abnormal tissue protruding from the open neural tube. To determine the origin of such protrusions, we compared gene expression profiles between open neural plate with normal morphology, and protrusion tissue. Neurulation-stage mouse embryos at 8.5 days of gestation were used to prepare open neural tube at the anterior aspect of neural tube closure site 1 by laser capture microdissection. For each sample, 10 sections were pooled, total RNA was extracted, and 7 ng of total RNA were used for expression profiling by Tag sequencing using an Applied Biosystems SolidSAGE kit for library construction, and an AB SOLiD 5500 XL instrument for sequencing. Protrusion tissue was prepared from whole embryos by microdissection, and 12ng of total RNA per sample was used for Tag sequencing. Sequence reads were mapped to RefSeq RNA, and count data per gene were obtained using a modified version of the Applied Biosystems SOLiDâ¢ SAGEâ¢ Analysis Software. Neural plate protrusion compared to open neural plate anterior of closure site 1 with normal morphology

ORGANISM(S): Mus musculus

SUBMITTER: J. Michael Salbaum

PROVIDER: E-GEOD-53075 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:At an incidence of approximately 1/1000 births, neural tube defects (NTDs) comprise one of the most common and devastating congenital disorders. In an attempt to enhance and expand our understanding of neural tube closure, we undertook a high-throughput gene expression analysis of the neural tube as it was forming in the mouse embryo. Open and closed sections of the developing neural tube were micro-dissected from mouse embryos, and hybridized to Affymetrix mouse expression arrays. Clustering of genes differentially regulated in open and closed sections of the developing neural tube highlighted molecular processes previously recognized to be involved in neural tube closure and neurogenesis. Analysis of the genes in these categories identified potential candidates underlying neural tube closure. In addition, we identified approximately 25 novel genes, of unknown function, that were significantly up-regulated in the closed neural tube. Based on their expression patterns in the developing neural tube, five novel genes are proposed as interesting candidates for involvement in neurogenesis. The high-throughput expression analysis of the neural tube as it forms allows for better characterization of pathways involved in neural tube closure and neurogenesis, and hopefully will strengthen the foundation for further research along the pathways dictating neural tube development. Embryos were dissected at days E8.5 and E9.5, and the neuroepithelium/ neural tube were mechanically detached from underlying tissues, and then separated into two regions: 1) M-bM-^@M-^\open neuroepitheliumM-bM-^@M-^]: neuroepithelial tissue caudal to the open/closed junction, and 2) M-bM-^@M-^\closed neural tubeM-bM-^@M-^], extending from a somiteM-bM-^@M-^Ys breadth rostral to the open/closed junction, up to the level of the fifth- or sixth-to-last somite. Samples consisted of biological triplicates of RNA extract from the above tissues (pooled by litter, and representing a total of 111 embryos): E8.5 open neuroepithelium, E8.5 closed neural tube, E9.5 open neuroepithelium, and E9.5 closed neural tube. Thus, a total of 12 samples (representing 111 embryos) were hybridized to the GeneChip Mouse Genome 430 2.0 Array (Affymetrix Inc., Santa Clara, CA, USA). One of the samples (06, closed E8.5) deviated significantly from the others in quality assessment and was therefore removed from subsequent analysis and not submitted to GEO.

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Perturbed gastrulation in embryos from diabetic pregnancies.

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