We applied RNA-seq analysis to human islet cells, received from 3 independent donors, treated with either redifferentiation cocktail + ARX shRNA, or redifferentiation cocktail + control shRNA or left untreated. Overall design: Examination of the effect of ARX inhibition on redifferentiation of ß-cell-derived (BCD) cells
Redifferentiation of expanded human islet β cells by inhibition of ARX.
No sample metadata fields
View SamplesGliomas arising in the brainstem and thalamus are devastating tumors that are difficult to surgically resect due to their proximity to eloquent brain structures. Here, we performed a comprehesive genomic and epigenomic study, using gene expression and methylation microarrays, to research on th different genomic and epigenetic signatures between brainstem, thalamic, and supratentorial gliomas.
Exome sequencing identifies somatic gain-of-function PPM1D mutations in brainstem gliomas.
Sex, Age
View SamplesThe gene expression pattern of spherical neural masses (SNM) derived from HPRT knockdown murine D3 stem cells during neuronal differentiation to final neurons was invesitigated by RNA-Seq based gene expression analysis and the results were interpreted by GO, GSEA and signaling pathway analyses with Avadis NGS and PANTHER Classification System. Overall design: For RNA-Seq experiment, total RNAs of HPRT knockdown and control SNMs were extracted time-sequentially during the SNM differentiation such as differentiation-day 0, 1, 2, 3, 4, 6, 8, 10, 12, and 14 to final neurons, and each prepared library applied to the HiSeq 2000 sequencer of Illumina for 50 cycles of single lane run.
The housekeeping gene hypoxanthine guanine phosphoribosyltransferase (HPRT) regulates multiple developmental and metabolic pathways of murine embryonic stem cell neuronal differentiation.
Specimen part, Cell line, Treatment, Subject
View SamplesBackground & Aims: MiRNAs are small (~22 nucleotide), non-coding RNA molecules that regulate gene expression through imperfect complementarity with target messenger RNAs. The function of miRNA in mammalian organogenesis is largely unknown. Conditional loss-of-function of Dicer, the enzyme that processes precursor miRNA transcripts into their mature, active form, has been shown to cause severe defects in a number of organ systems. Here we address the role of Dicer in liver development and function. Methods: Mice lacking Dicer function in hepatocytes were generated using an Afp-Cre strain to drive deletion of a floxed Dicer allele. Deletion of the flox-dicer allele was confirmed by quantitative PCR. Decreased miRNA levels detected by quantitative RT-PCR and in situ hybridization confirmed loss of Dicer function. Gene expression microarray analysis was performed on liver RNA from P28 mutant and control mice. Liver sections from mutant and control mice ranging from embryonic stages through 3-4 months of age were examined and liver function tests were performed on adult mice. Results: Mice lacking hepatocyte Dicer function were born alive at the expected frequency, and had grossly normal appearance and behavior. Despite the loss of mature miRNA, hepatic function was normal, as reflected by normal blood gludose, albumin, cholesterol, and bilirubin. However, mutant mice between 2-4 months of age exhibit progressive hepatocyte damage, elevated ALT/AST, with evidence of balanced proliferation and apoptosis in the lobule. Microarray analysis indicates large-scale changes in gene expression, with increased expression of many miRNA targets, as well as imprinted genes. Conclusions: Loss of miRNA processing in the liver at late gestation has a remarkably mild phenotype, suggesting that miRNAs do not play an essential role in hepatic physiology. However, miRNA deficiency results in hepatocyte apoptosis and balanced hepatocyte regeneration. Finally, microarray analysis of gene expression in mutant liver suggests a previously unrecognized role for Dicer in the repression of imprinted genes.
Hepatic function is preserved in the absence of mature microRNAs.
Age, Specimen part
View SamplesPurpose: using RNA-seq as a screening tool to determine candidate genes of interest within a genetically defined neural subpopulation in the zebrafish embryonic spinal cord. Results: The early embryonic spinal cord displays patterns of spontaneous activity that generate the earliest motor behavior in the zebrafish. We show the behavior and the neural activity to be inhibited by environmental levels of light. Since at these young ages the fish is blind, and since restricted illumination patterns on the trunk of the fish can elicit a photo-response, we hypothesized that the photo-inhibition is an intrinsic property of the active central pattern generator network within the spinal cord. We FACS-isolated cells from this network as well as those from a panneuronal population and sequenced mRNAs. Through differential expression analysis we identified vertebrate ancient long opsin a as a candidate and then further validated its function in the circuit through knockdown and rescue experiments. Overall design: RNA sequencing of 2 FACS purified neural populations from zebrafish spinal cord.
A spinal opsin controls early neural activity and drives a behavioral light response.
No sample metadata fields
View SamplesIn this study, we confirmed that transformed dedifferentiated astrocytes and neurons acquired a stem/progenitor cell state, although they still retained gene expression memory from their parental cell-type. Transcriptional network analysis on transformed cells revealed up-regulation of genes involved in three signaling pathways: Wnt signaling, cell cycle and focal adhesion with the gene Spp1, also known as osteopontin (OPN) serving as a key node connecting these three pathways. Inhibition of OPN blocked the formation of aggregated neurospheres, affected the proliferative capacity of transformed cell-types and reduced the expression levels of neural stem cell markers. Specific inhibition of OPN in murine glioma tumors prolonged mice survival. We conclude that OPN is an important player in dedifferentiation of cells during tumor formation, hence its inhibition can be a therapeutic target for glioblastoma. Overall design: Cortical neurons and astrocytes were derived from 11 days old SynapsinI-Cre and GFAP-Cre mice, respectively. The cells were cultured in their respective media to maintain their identity. These cells were then transduced with HRas-shp53 lentivirus with a transduction efficiency of >90%. The transduced neurons and astrocytes were later switched to neural stem cell media devoid of serum and supplemented with FGF-2 (NSC media). Within one week, these cells became proliferative and aggregated to form free-floating neurospheres. These cells, hereinafter referred to as NSynR53 and AGR53, respectively, were later harvested and mRNA collected for sequencing library generation using DP-seq. To assess the regression of these cells to an undifferentiated state along the differentiation axis, enriched populations of mESC and NSC were also grown in vitro and mRNA obtained from these cells were subjected to sequencing library preparation.
Identification of therapeutic targets for glioblastoma by network analysis.
No sample metadata fields
View SamplesPurpose: The goal of this study was to determine biological consequences during liver regeneration following partial hepatectomy in mice by next-generation sequencing. A particular interest was to compare mice with either a floxed b-PDGFR allele to mice that harbored a deletion of b-PDGFR in hepatic stellate cells (HSCs), by crossing b-PDGFR fl/fl mice with transgenic GFAP-Cre mice. Methods: b-PDGFR fl/fl mice or mice with a HSC-specific deletion of b-PDGFR underwent either sham operation or 70% partial hepatectomy. Following 72 hours, livers were collected and total RNA was extracted using tizol, followed by a purification using Quiagen spin columns including an on-column DNAse digestion step. Conclusion: Our study represents a detailed analysis of hepatic transcriptome, with biologic replicates, generated by RNA-seq technology of livers following sham operation or partial hepatectomy in b-PDGFR fl/fl mice or b-PDGFRfl/fl/GRAP-Cre mice. Overall design: Whole liver mRNA profiles of sham operated livers or livers collected 72hours after partial hepatectomy of beta-PDGFR fl/fl and beta-PDGFR fl/fl/GFAP-Cre (creating a hepatic stellate cell-specific deletion of b-PDGFR) mice were generated by deep sequencing, in duplicate, using Illumina HiSeq2000.
Induction and contribution of beta platelet-derived growth factor signalling by hepatic stellate cells to liver regeneration after partial hepatectomy in mice.
No sample metadata fields
View SamplesOptic nerves are an accessible part of the CNS, providing a source of glia without the presence of neuronal cell bodies. Therefore, an analysis was carried out of gene expression in optic nerves at P4, before myelination begins and at P10, when myelination is very actively proceeding. The goal was to obtain a profile of the changing gene expression that accompanies this transition from unmyelinated CNS nerve to myelinated nerve.
Towards resolving the transcription factor network controlling myelin gene expression.
Specimen part
View SamplesWe have established that BMP6 is an important endogenous regulator of human osteoblast differentiation. Our preliminary experiment showed that 8 hour BMP6 treatment induced early osteoblast markers in hMSC.
GAGE: generally applicable gene set enrichment for pathway analysis.
No sample metadata fields
View SamplesMeiosis is a complex developmental process that generates haploid cells from diploid progenitors. We measured mRNA abundance and protein production through yeast sporulation and found strong temporal control for most genes, achieved through both mRNA levels and translational regulation. Monitoring the timing of protein production revealed novel factors involved in recombination and helped to illuminate the molecular basis of the broad restructuring of meiotic cells. We also found a strong increase in noncanonical translation at short open reading frames (sORFs) on unannnotated transcripts and upstream regions of known transcripts (uORFs). Ribosome occupancy at near-cognate uORFs was associated with more efficient ORF translation; while some AUG uORFs, often on regulated leader extensions, acted comptetitively. This work reveals a pervasive role for meiotic translational control and great complexity in genomic coding. Overall design: Fine mapping of gene expression through meiosis reveals extensive regulation of protein synthesis and widespread non-canonical translation.
High-resolution view of the yeast meiotic program revealed by ribosome profiling.
Cell line, Subject
View Samples