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Next-generation RNA sequencing of human astrocyte cultures to study the comparative levels of transmembrane proteins and their transcriptional variants

ABSTRACT: Next-generation RNA sequencing of human astrocyte cultures to study the comparative levels of transmembrane proteins and their transcriptional variants

PROVIDER: PRJNA948014 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR23949742_1.fastq.gz	Fastqsanger.gz
	SRR23949742_2.fastq.gz	Fastqsanger.gz
	SRR23949743_1.fastq.gz	Fastqsanger.gz
	SRR23949743_2.fastq.gz	Fastqsanger.gz
	SRR23949744_1.fastq.gz	Fastqsanger.gz

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Next-generation RNA sequencing of human astrocyte cultures to study the comparative levels of transmembrane proteins and their transcriptional variants

Project description:The aim of this study was to perform Next-generation RNA sequencing to analyse the expression of transmembrane proteins in human astrocytes and compare the prevalance of transcriptional variants thereof.

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Rat Astrocyte-Neuron Co-cultures versus purified co-cultured Astrocytes versus Astrocyte alone

Project description:Profiling of transcriptional changes in rat astrocytes when co-cultured with neurons: comparison of astrocytes cultured alone with astrocytes co-cultured with mouse hippocampal neurons. Co-cultured astrocytes are isolated using cold jet, a novel tool for these neuron-glia cultures. Over the last decade, the importance of astrocyte-neuron communication in neuronal development and synaptic plasticity has become increasingly clear. Since neuron-astrocyte interactions represent highly dynamic and reciprocal processes, we hypothesized that at least part of the involved astrocyte genes may be regulated as a consequence of their interactions with maturing neurons. In order to identify such neuron-induced astrocyte genes in vitro, we tested the effectiveness of the ‘cold jet’, a new method for separation of neurons from co-cultured astrocytes. The cold jet method is performed under ice-cold conditions and avoids protease-mediated isolation of astrocytes or time-consuming centrifugation, yielding intact astrocyte mRNA with approximately 90% of neuronal RNA removed. Using this method, we executed genome-wide profiling in which RNA derived from astrocyte-only cultures was compared with astrocyte RNA derived from differentiating neuron-astrocyte co-cultures. Data analysis revealed changes in expression of a large number of mRNAs and biological processes, including novel findings. Thus, cold jet is an efficient method to separate astrocytes from neurons in co-culture, and in this study reveals that neurons induce robust gene-expression changes in co-cultured astrocytes.

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Ethanol induced transcriptional and translational changes in Aldh1l1-EGFP/Rpl10a cortical astrocyte cultures

Project description:The role astrocytes play in brain development and function has come under increased focus as the diversity of roles they are involved in has become apparent. We have previously shown ethanol exposed astrocytes alter neuronal neurite outgrowth in an in vitro co-culture system and that ethanol alters the astrocyte produced extracellular matrix (ECM) in vitro, with similar alterations in vivo. Here we utilize the translating ribosome affinity purification (TRAP) procedure in Aldh1l1-EGFP/Rpl10a transgenic mouse primary cortical astrocyte cultures to transcriptionally and translationally profile the astrocyte response to ethanol. We found large differences in the total RNA pool in comparison to the translating RNA pool while the ethanol response within each pool showed a large degree of overlap. Comparisons to published datasets indicate the in vitro model used here are most similar to PD1-PD7 in vivo cortical astrocytes and the ethanol regulated genes showed significant overlap with models of chronic ethanol exposure in astrocytes and a model of third-trimester ethanol exposure in the hippocampus and cerebellum. These findings will further our understanding of the effects of ethanol on astrocyte gene expression and protein translation and how these changes alter brain development.

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Project description:Inflammation features in diverse central nervous system disorders such as stroke, trauma, neurodegeneration, infection and autoimmunity. To better understand how inflammatory mediators may alter astrocyte functions, we examined the effects of transforming growth factor-Î²1 (TGF-Î²1), lipopolysaccharide (LPS) and interferon-gamma (IFNÎ³) on purified, murine, primary cortical astrocyte cultures. We used microarrays to conduct whole genome expression profiling, and measured calcium signaling, which is implicated in mediating dynamic astrocyte functions. Combinatorial exposure to TGF-Î²1, LPS and IFNÎ³ significantly modulated astrocyte expression of over 6,800 genes and resulted in both additive and synergistic changes compared with individual stimuli alone. Bioinformatic analysis revealed that combinatorial treatment significantly and markedly up regulated molecular networks and pathways associated with immune signaling and with regulation of cell compromise, death, growth and proliferation. These findings provide databases of astrocyte transcriptome changes elicited by the inflammatory stimuli, TGF-Î²1, LPS and IFNÎ³ alone and in combination, and show that these stimuli up regulate astrocyte molecular networks associated with immune- and injury-related functions and significantly alter astrocyte calcium signaling evoked by multiple GPCR. We used microarrays to examine the effects of transforming growth factor-Î²1 (TGF-Î²1), lipopolysaccharide (LPS) and interferon-gamma (IFNÎ³) on purified, murine, primary cortical astrocyte cultures.

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Wildtype and Gde3 knockout conditioned media from primary astrocyte culture

Project description:Primary astrocyte cultures were prepared from wildtype and Gde3 knockout mouse brain. Conditioned media was collected, concentrated and analyzed using TMT-MS.

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Expression profile of primary murine astrocyte-enriched cultures (AEC) exposed to oxygen-glucose deprivation

Project description:Expression profile of primary murine astrocyte-enriched cultures (AEC) exposed to oxygen-glucose deprivation

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Identification of genes associated with the astrocyte-specific gene Gfap during astrocyte differentiation

Project description:Chromosomes and genes are non-randomly arranged within the mammalian cell nucleus. Clustering of genes is of great significance in transcriptional regulation. However, the relevance of gene clustering in their expression during differentiation of neural precursor cells (NPCs) into astrocytes remains unclear. We performed a genome-wide enhanced circular chromosomal conformation capture (e4C) to screen genes associated with an astrocyte-specific gene, glial fibrillary acidic protein (Gfap), during astrocyte differentiation. We identified 13 genes that were specifically associated with Gfap and expressed in NPC-derived astrocytes. These results provide evidence for functional significance of gene clustering in transcriptional regulation during NPCs differentiation. comparison of NPCs vs LIF+ vs LIF- cells.

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