Single Cell RNA Sequencing of fetal mouse neurosphere cultures.
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA655918 | ENA |
REPOSITORIES: ENA
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Project description:We performed single-cell RNA sequencing on neurosphere cultures derived from the dorsal telencephalic vesicles of sex segregated gestational day (GD) 12.5 C57BL/6J mouse fetuses.
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Project description:The expression of the mciroRNA, miR-153 is significantly altered in neural stem cells (NSCs) following exposure to the teratogens, alcohol and nicotine (PMIDs:22458409, 17687032). To better understand how miR-153 may mediate teratogenesis, we first needed to identify miR153 targets in fetal NSCs. Gestational day 12.5 fetal mouse cortical neural epithelium was isolated and grown as neurospheres in defined medium for studies. We overexpressed miR-153 in fetal NSCs for 24 hours in a transient transfection assay, and profiled mRNA expression using a microarray platform (codelink array) to identify potential target genes. Over expression of pre-miR-153 resulted in a ~32-fold induction of miR-153 compared to the transfection control. Transcript profiles were assessed from RNA samples (6 independent replicates in each condition) hybridized to codelink microarrays according to manufacturer's protocols. Data analysis showed that miR-153 overexpression resulted in statistically significant down-regulation of 102 genes (at a false discovery rate α=0.1, of a total of 860 transcripts down-regulated by ≥1.3-fold). These data suggested the role of this miRNA in moderately controlling expression hundreds of gene transcripts. These regulated genes may be important in controlling neural stem cell differentiation and downstream signaling pathways. Six independent replicates were performed for control and treatment groups, and twelve independent single channel intensity values were uploaded after normalization for analysis independent single channel intensity values were uploaded after normalization
2014-08-01 | E-GEOD-49684 | biostudies-arrayexpress
Project description:Recent reports have emphasized the pitfalls of iPSC technology including the potential for immunogenicity of transplanted cells. It is serious safety-related concern for iPSC-based cell therapy. However, preclinical data supporting the safety and efficacy of iPSCs are also accumulating. To address the concern of immunogenicity of ESCs/iPSCs or ESCs/iPSCs-derived neurospheres, global gene expression profiles were compared between undifferentiated mouse ESCs (EB3 line), mouse iPSCs (38C2 line), and ESC/iPSC-derived neurosphere and mouse primary culture of neurosphere obtained from fetal mouse ganglionic eminence. Mouse adult sklin fibroblast was used as a control. We used affymetrix microarrays to compare the global gene expression of neurospheres prepared several origins. Keywords: Expression profiling by array RNA extracted from neurospheres was hybridized to Affymetrix microarrays.
2012-03-01 | E-GEOD-31725 | biostudies-arrayexpress
Project description:The expression of the mciroRNA, miR-153 is significantly altered in neural stem cells (NSCs) following exposure to the teratogens, alcohol and nicotine (PMIDs:22458409, 17687032). To better understand how miR-153 may mediate teratogenesis, we first needed to identify miR153 targets in fetal NSCs. Gestational day 12.5 fetal mouse cortical neural epithelium was isolated and grown as neurospheres in defined medium for studies. We overexpressed miR-153 in fetal NSCs for 24 hours in a transient transfection assay, and profiled mRNA expression using a microarray platform (codelink array) to identify potential target genes. Over expression of pre-miR-153 resulted in a ~32-fold induction of miR-153 compared to the transfection control. Transcript profiles were assessed from RNA samples (6 independent replicates in each condition) hybridized to codelink microarrays according to manufacturer's protocols. Data analysis showed that miR-153 overexpression resulted in statistically significant down-regulation of 102 genes (at a false discovery rate α=0.1, of a total of 860 transcripts down-regulated by ≥1.3-fold). These data suggested the role of this miRNA in moderately controlling expression hundreds of gene transcripts. These regulated genes may be important in controlling neural stem cell differentiation and downstream signaling pathways.
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Project description:Neurosphere cultures were established from individually isolated E 14.5 forebrains of mouse embryos that carry a bcl2 transgene. Single-cell suspensions were prepared, seeded at 1x105 cells/ml and treated for two days with 150 nM Trichostatin A (TSA; histone deacetylase inhibitor), 500 nM 5-Aza-2-deoxycytidine (AzaC; demethylating agent) or both compounds, or left untreated. Two independent experiments were performed.
2007-12-19 | E-GEOD-587 | biostudies-arrayexpress
Project description:Significant efforts are ongoing to develop refined differentiation protocols to generate midbrain DA neurons from pluripotent stem cells (PSCs) for application in disease modeling, diagnostics, drug screening, and cell-based therapies for Parkinson’s Disease. An increased understanding of the timing and molecular mechanisms promoting the generation of distinct subtypes of midbrain DA during normal development will be essential for guiding future efforts to precisely generate molecularly defined and subtype-specific DA neurons from pluripotent stem cells. In this study, we used droplet-based single-cell RNA sequencing (scRNA-seq) to transcriptionally profile fetal DA neurons from human embryos at different stages of ventral midbrain (VM) development (6, 8, and 11 weeks post-conception) and primary fetal 3D cultures thereof that allowed differentiation and functional maturation of human DA neurons. This approach allowed us to define the molecular identities of distinct human DA progenitors and neurons at single-cell resolution and construct developmental trajectories of cell types in the developing fetal VM. Overall, these findings provide a unique transcriptional profile of developing human fetal VM and functionally mature human DA neurons, which can be used to guide stem cell-based therapies and disease modeling approaches in PD.
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