Transcriptomic analysis of human astrocytes in vitro reveals hypoxia-induced mitochondrial dysfunction, modulation of metabolism and dysregulation of the immune response
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE145935 | GEO | 2020/02/26
REPOSITORIES: GEO
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Project description:Oxygen is vital for the development and survival of mammals. In response to hypoxia, the brain initiates numerous adaptive responses at the organ level as well as at the molecular and cellular levels, including the alteration of gene expression. Astrocytes play critical roles in the proper functioning of the brain; thus the manner in which astrocytes respond to hypoxia is likely important in determining the outcome of brain hypoxia. Here, we used microarray gene expression profiling and data-analysis algorithms to identify and analyze hypoxia-responsive genes in primary human astrocytes. We also compared gene expression patterns in astrocytes with those in human HeLa cells and pulmonary artery endothelial cells (ECs). Remarkably, in astrocytes, five times as many genes were induced as suppressed, whereas in HeLa and pulmonary ECs, as many as or more genes were suppressed than induced. More genes encoding hypoxia-inducible functions, such as glycolytic enzymes and angiogenic growth factors, were strongly induced in astrocytes compared with HeLa cells. Furthermore, gene ontology and computational algorithms revealed that many target genes of the EGF and insulin signaling pathways and the transcriptional regulators Myc, Jun, and p53 were selectively altered by hypoxia in astrocytes. Indeed, Western blot analysis confirmed that two major signal transducers mediating insulin and EGF action, Akt and MEK1/2, were activated by hypoxia in astrocytes. These results provide a global view of the signaling and regulatory network mediating oxygen regulation in human astrocytes.. This SuperSeries is composed of the following subset Series:; GSE3045: Astrocytic response to hypoxia; GSE3051: HeLa response to hypoxia Experiment
2007-10-26 | E-GEOD-4483 | biostudies-arrayexpress
Project description:We used cortical
human fetal astrocytes, density 8-15000 cells/cm2 (ScienCell, San Diego, CA, USA seeded separately in Astrocyte
medium (ScienCell, San Diego, CA, USA) P0.
Methamphetamine (Sigma-Aldrich, St Louis, MO, USA) was prepared at a stock concentration of 10mM in distilled water. Methamphetamine dilutions 50 micromolar were made in Endo and Neuro/Astro/Peri media respectively and applied to the cells for 26 hrs. Further info in "Proteomic and metabolomic characterization of human neurovascular unit cells in response to methamphetamine" Herland et al Adv Biosystems 2020 [doi:10.25345/C55F3P] [dataset license: CC0 1.0 Universal (CC0 1.0)]
2020-07-21 | MSV000085795 | MassIVE
Project description:Oxygen is vital for the development and survival of mammals. In response to hypoxia, the brain initiates numerous adaptive responses at the organ level as well as at the molecular and cellular level, including the alteration of gene expression. Astrocytes play critical roles in the proper functioning of the brain, thus the manner in which astrocytes respond to hypoxia is likely important in determining the outcome of brain hypoxia. We used microarray gene expression profiling and data analysis algorithms to identify and analyze hypoxia-responsive genes in primary human astrocytes. We also compared the gene expression patterns in astrocytes with those in human HeLa cells and pulmonary artery endothelial cells (ECs). Remarkably, in astrocytes, 5 times as many genes were up-regulated as down-regulated, whereas in HeLa and pulmonary ECs, as many or more genes were down-regulated as up-regulated. More genes encoding hypoxia-inducible functions, such as glycolytic enzymes and angiogenic growth factors, were strongly induced in astrocytes compared to HeLa cells. The extent of induction was also greater than in HeLa cells. Further, gene ontology and computational algorithms revealed that many target genes of the EGF and insulin signaling pathways and the transcriptional regulators Myc, Jun and p53 were selectively altered by hypoxia in astrocytes and HeLa cells to a varying degree. These results provide a global view of the signaling and regulatory network mediating oxygen regulation in astrocytes, in comparison with that in HeLa cells. Experiment Overall Design: Gene expression profiles of three independent batches of astrocytes grown under normoxia were compared to gene expression profiles of three independent batches of astrocytes grown under 1% oxygen for 24 hours. Affymetrix U133 plus 2.0 microarrays were used.
2008-06-12 | E-GEOD-3045 | biostudies-arrayexpress
Project description:Cortical human fetal astrocytes, density 8-15000 cells/cm2 ( ScienCell, San Diego, CA, USA) seeded in Astrocyte medium (ScienCell, San Diego, CA, USA). Further info in "Proteomic and metabolomic characterization of human neurovascular unit cells in response to methamphetamine" Herland et al Adv Biosystems 2020
2020-07-23 | MSV000085817 | GNPS
Project description:Regarding astrocytes and microglia, we are looking for pathways related with inflammatory activation, phagocytosis and vesicular/exosome trafficking. Regarding motor neurons, we are searching for pathways related with mitochondrial dynamics, Endoplasmic Reticulum stress, cell death, axonal transport, synaptic vesicles transport and oxidative stress. The general aim is to identify putative hits in ALS that relates with miR-146a expression.
2021-04-06 | PXD024294 | Pride
Project description:Transcriptional profiling of mouse primary astrocytes comparing control untreated astrocytes with astrocytes treated with recombinant LCN2 protein (10 micro gram/ml). Goal was to determine the effects of LCN2 treatment on global gene expression in astrocytes. A secreted protein lipocalin-2 (LCN2) has been implicated in diverse cellular processes including cell morphology and migration. We have previously demonstrated that lcn2 mediates reactive astrocytosis. In order to further understand the role of lcn2 in the CNS, astrocyte transcriptome was analyzed following LCN2 treatment. Chemokines were the major group of genes upregulated by LCN2. Two-condition experiment, control untreated astrocytes vs. LCN2 protein treated astrocytes. Biological replicates: 1 control replicates, 1 treated replicates.
2012-06-29 | E-GEOD-15667 | biostudies-arrayexpress
Project description:The central nervous system normally functions at O2 levels which would be regarded as hypoxic by most other tissues. However, most in vitro studies of neurons and astrocytes are conducted under hyperoxic conditions without consideration of O2-dependent cellular adaptation. We analyzed the reactivity of astrocytes to 1, 4 and 9% O2 tensions compared to the cell culture standard of 20% O2, to investigate their ability to sense and translate this O2 information to transcriptional activity. Variance of ambient O2 tension for rat astrocytes resulted in profound changes in ribosomal activity, cytoskeletal and energy-regulatory mechanisms and cytokine-related signaling. Clustering of transcriptional regulation patterns revealed four distinct response pattern groups that directionally pivoted around the 4% O2 tension, or demonstrated coherent ascending/decreasing gene expression patterns in response to diverse oxygen tensions. Immune response and cell cycle/cancer-related signaling pathway transcriptomic subsets were significantly activated with increasing hypoxia, whilst hemostatic and cardiovascular signaling mechanisms were attenuated with increasing hypoxia. Our data indicate that variant O2 tensions induce specific and physiologically-focused transcript regulation patterns that may underpin important physiological mechanisms that connect higher neurological activity to astrocytic function and ambient oxygen environments. These strongly defined patterns demonstrate a strong bias for physiological transcript programs to pivot around the 4% O2 tension, while uni-modal programs that do not, appear more related to pathological actions. The functional interaction of these transcriptional ‘programs’ may serve to regulate the dynamic vascular responsivity of the central nervous system during periods of stress or heightened activity. Cerebral cortices were removed from 6-8-day-old Wistar rats, cortical astrocytes were isolated and these cells were maintained in a humidified incubator at 37°C (95% air; 5% CO2). This resulted in a culture of cortical astrocytes, as confirmed by visual inspection the following day and later by glial fibrillary acidic protein immunohistochemistry. Any cortical astrocyte culture that was not homogenous was disposed of and not used in this study. Culture medium was exchanged every 7 days and cells were grown in culture for up to 14 days. 24hr prior to experimentation cells were transferred to an hypoxic workstation equilibrated with either 1%, 4%, 9% or 21% O2 and 5% CO2, with the remaining percentage gas being N2. Once cortical astrocytes had reached approximately 90% confluence (75cm2 flask) they were subjected to hypoxia as above, washed with PBS, removed from the flask base with 0.05% trypsin-EDTA (Gibco) and then gently centrifuged (500xg). The cell pellet was re-suspended in PBS, centrifuged twice more to remove any traces of media, then triturated in 8-10 volumes of RNAlater (Applied Biosystems), frozen and stored at -80oC. Messenger RNA was amplified and labeled with biotin using the standard Illumina protocol (Illumina TotalPrep RNA Amplification Kit Ambion; Austin, TX, cat # IL1791), and hybridized to Illumina's Sentrix Rat Ref-12,v1 Expression BeadChips (Illumina, San Diego, CA). Three replicates from each treatment group (1%, 21%, 4%, and 9% O2) were hybridized and the data was extracted using the Illumina BeadStudio software(v3.4).
2012-09-20 | E-GEOD-29296 | biostudies-arrayexpress
Project description:Astrocytes were purified from mouse cortices and human cortex tissue by immunopanning with HepaCAM. All treatments were proceeded on 3DIV in serum-free medium. Astrocytes were treated by TNFα for 48hr, by Poly I:C for 72hr, or by hypoxia for 72hr. Acutely purified atrocytes RNA was harvest immediately after HepaCAM immunopanning. Serum selected astrocytes RNA was collected after 4-6 days culturing in serum medium. Serum-free cultured astrocytes RNA was collected on 5-6 DIV. Serum-selection purified human astrocytes were transplanted into P2-11 Rag2 immunodeficient mouse brains, and after about 8 months, transplanted human and host mouse astocytes RNA was collected acutely on HepaCAM immunopanning dish.
2021-04-21 | GSE147870 | GEO
Project description:We used cortical
human fetal astrocytes, density 8-15000 cells/cm2 (ScienCell, San Diego, CA, USA seeded separately in Astrocyte
medium (ScienCell, San Diego, CA, USA) P0.
Methamphetamine (Sigma-Aldrich, St Louis, MO, USA) was prepared at a stock concentration of 10mM in distilled water. Methamphetamine dilutions 50 micromolar were made in Endo and Neuro/Astro/Peri media respectively and applied to the cells for 26 hrs. Further info in "Proteomic and metabolomic characterization of human neurovascular unit cells in response to methamphetamine" Herland et al Adv Biosystems 2020 [doi:10.25345/C55F3P] [dataset license: CC0 1.0 Universal (CC0 1.0)]
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Project description:Glaucoma leads to vision loss due to retinal ganglion cell death. Astrocyte reactivity contributes to neurodegeneration. Our recent study found that lipoxin B4 (LXB4), produced by retinal astrocytes, has direct neuroprotective actions on retinal ganglion cells. In this study, we aimed to investigate how the autacoid LXB4 influences astrocyte reactivity in the retina under inflammatory cytokine-induced activation and ocular hypertension conditions. The protective activity of LXB4 was investigatedin vivousing the mouse silicone-oil model of chronic ocular hypertension (n=40). By employing a range of analytical techniques, including bulk RNA-seq, RNAscope in-situhybridization, qPCR, and lipidomic analyses, we discovered the formation of neuroprotective lipoxins in rodents (including the retina and optic nerve), primates (optic nerve), and human brain astrocytes, indicating their presence across various species. Our findings in the mouse retina demonstrated significant dysregulation of the lipoxin pathway in response to chronic ocular hypertension, leading to an increase in 5-lipoxygenase (5-LOX) activity and a decrease in 15-lipoxygenase activity. This dysregulation was coincident with a marked upregulation of astrocyte reactivity. Reactive human brain astrocytes also showed a significant increase in 5-LOX. Administration of LXB4 regulated the lipoxin pathway, restored and amplified LXA4 generation (another lipoxin with distinct bioactions), and mitigated astrocyte reactivity in mouse retinas and human brain astrocytes. In conclusion, the lipoxin pathway is functionally expressed in rodents, primates, and human astrocytes, and is a resident neuroprotective pathway that is downregulated in reactive astrocytes. Novel cellular targets for LXB4’s neuroprotective action are inhibition of astrocyte reactivity and restoration of lipoxin generation. Amplifying the lipoxin pathway is a potential target to disrupt or prevent astrocyte reactivity in neurodegenerative diseases.
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