Transcription profiling of mouse macrophage response to hypoxia, part 1
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ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Renee Rubio
PROVIDER: E-TIGR-90 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:We hypothesize that the culture media collected from macrophages exposed to intermittent hypoxia will induce a greater pro-inflammatory gene profile in naïve cultured macrophages than will culture media collected from macrophages exposed to sustained hypoxia. We will evaluate gene expression using microarray analysis of RNA collected from RAW 264.7 macrophages cultured for 24 hours in DMEM media obtained from 1) cells cultured with intermittent hypoxia (2 minute cycles: 90 seconds at 40 Torr and 30 seconds at 8 Torr), 2) media exposed to intermittent hypoxia, 3) cells cultured with sustained hypoxia (8 Torr), 4) media exposed to sustained hypoxia and 4) standard tissue culture conditions (fresh DMEM media; reference).
2005-01-01 | E-TIGR-91 | biostudies-arrayexpress
Project description:Here we used microarrays to characterize changes in global gene expression in the hepatopancreas of Pacific white shrimp, Litopenaeus vannamei, exposed to short term (4 h) hypoxia (H) or hypercapnic hypoxia (HH) or long term (24 h) H or HH, compared to animals in air-saturated water (normoxia). The transcriptomes of crustaceans exposed to low O2 and high CO2 contained both shared and treatment-specific signature genes (q M-bM-^IM-$ 0.01, FC M-bM-^IM-% 1.5), with shifts characteristic of metabolic depression rather than anaerobic metabolism. Down-regulated signature genes dominated the transcript profile in three of the four treatments (H 4 h, H 24 h, 4 h HH); many of these genes were involved in amino acid or RNA metabolism or in translation, including several tRNA synthetases. Unique patterns of gene expression such as increased lipid metabolism and hemocyanin synthesis (H 24 h) and initiation of apoptosis (24 h HH) were tied to specific treatments. This work contributes insight to the effects that human perturbations might have on estuarine organisms, and the importance of examining the impacts of environmentally relevant combinations of hypoxia and hypercapnia on estuarine populations. L. vannamei were exposed for 4 or 24 hours to one of the following conditions: normoxia, hypoxia or hypercapnic hypoxia. Hepatopancreas tissue from individual animals was dissected, total RNA extracted, labelled and hybridized to oligonucleotide microarrays with probes for 21,864 L. vannamei unigenes. Treatments were repeated until a total of 7 biological replicates was obtained for each time:treatment combination, except for the 24 h normoxia group, represented by 6 replicates.
2012-04-29 | E-GEOD-29029 | biostudies-arrayexpress
Project description:Transcriptional responses to hypoxia were compared between a hypoxia tolerant fish (tidepool sculpin; Oligocottus maculosus) and a hypoxia intolerant fish (silverspotted sculpin; Blepsias cirrhosus). To determine if, and how, transcriptional plasticity is associated with differences in hypoxia tolerance, each species was subjected to a hypoxic time-course and liver was sampled at normoxia, 3hr, 8hr, 24hr, 48hr and 72hr of hypoxia. The hypoxic level for each species was scaled to the species' own tolerance level (relative exposure) in order to elicit similar tissue level hypoxia between the two sculpin species. Each species was also subjected to a single environmental O2 tension (absolute exposure) and sampled at 24 hrs of hypoxia.
2014-11-01 | E-MTAB-3043 | biostudies-arrayexpress
Project description:Lactic acidosis and hypoxia are two prominent tumor microenvironmental stresses that are both known to exert important influences on gene expression and phenotypes of cancer cells. But very little is known about the cross-talk and interaction between these two stresses. We performed gene expression analysis of MCF7 cells exposed to lactic acidosis, hypoxia and combined lactic acidosis and hypoxia. We found the hypoxia response elicited under hypoxia was mostly abolished upon simultaneous exposure to lactic acidosis. The repression effects are due to loss of HIF-1α protein synthesis under lactic acidosis. In addition, we showed lactic acidosis strongly synergizes with hypoxia to activate the unfold protein response (UPR) and inflammation response which are highly similar to amino acid deprivation responses (AAR). The statistical factor analysis of hypoxia and lactic acidosis responses indicated that ATF4 locus, an important activator in the UPR/AAR pathway, is amplified in subsets of breast tumors and cancer cell lines. Varying ATF4 levels dramatically affect the ability to survive the post-stress recovery from hypoxia and lactic acidosis and may suggest its selection of ATF4 amplification in human cancers. These data suggest that lactic acidosis interacts with hypoxia by both inhibiting the canonical hypoxia response and while activating the UPR and inflammation response. Gain of ATF4 locus may offer survival advantages to allow successful adaptation to frequent fluctuations of oxygen and acidity in tumor microenvironment. Collectively, our studies have provided linkage between the short-term transcriptional responses to the long term selection of the DNA copy number alterations (CNAs) under tumor microenvironmental stresses. RNAs from MCF7 cells exposed to control condition (ambient air ~21% O2, no lactate and neutral pH), lactic acidosis (ambient air, 10 mM Lactate and pH 6.7), hypoxia (1% pO2, no lactate and neutral pH) and the combined lactic acidosis and hypoxia (1% pO2, 10 mM Lactate and pH 6.7) condition for 24 hours were extracted by miRVana kits (Ambion) and hybridized to Affymetrix Human genome 133A 2.0 arrays with standard protocol.
2012-02-27 | E-GEOD-29406 | biostudies-arrayexpress
Project description:Alternative RNA splicing analysis in Hep3B cell cultured under 21% (N1,3,5) or 1.2% (H2,4,6) oxygen Hypoxia is a common characteristic of many solid tumors. The hypoxic microenvironment stabilizes hypoxia-inducible transcription factor 1? (HIF1?) and 2? (HIF2?) to activate gene transcription, which promotes tumor cell survival. 95% of human genes are alternatively spliced, producing RNA isoforms that code functionally distinct proteins. Thus, effective hypoxia response requires increased HIF target gene transcription as well as proper RNA splicing of these HIF target genes. However, it is unclear if and how hypoxia regulates RNA splicing of HIF target genes. This study determined the effects of hypoxia on alternative splicing (AS) of HIF and non-HIF target genes in Hep3B cells and characterized the role of HIF in regulating AS of HIF induced genes. The results indicated that hypoxia generally promotes exon inclusion for hypoxia-induced, but reduces exon inclusion for hypoxia reduced genes. Mechanistically, HIF activity, but not hypoxia per se is found to be necessary and sufficient to increase exon inclusion of several HIF target genes including pyruvate dehydrogenase kinase 1 (PDK1). PDK1 splicing reporters confirmed that transcriptional activation by HIF is sufficient to increase exon inclusion of PDK1 splicing reporter. In contrast, transcriptional activation of the PDK1 minigene by other transcription factor in the absence of endogenous HIF target gene activation fails to alter PDK1 RNA splicing, demonstrating a novel role of HIF target gene(s) in regulating RNA splicing of HIF target genes. Implications:This study demonstrates a novel function of HIF in regulating RNA splicing of HIF target genes. We analyzed total RNA from Hep3B cells cultured under 21% (N1,3,5) or 1.2% (H2,4,6) oxygen using the Affymetrix Human Exon 1.0 ST platform. Array data was processed by Altanalyze software version 2.0.7. Techinical replicates were performed for Nx and Hx treated Hep3B cells
2014-05-14 | E-GEOD-57613 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2014-10-15 | E-GEOD-48983 | 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:Study of hypoxia-induced differential expression during zebrafish development at gastrulation (shield) and segmentation (8-somite) stages An 8-chip study with 2 biological replicates from +/- hypoxia at 2 different stages of development. Each sample is prepared from 12-15 dechorionated, hand-selected, developmentally synchronized embryos exposed to 1-2 hours of hypoxia.
2015-05-02 | E-GEOD-68473 | biostudies-arrayexpress
Project description:The aim of this study was to characterize the basal expression of OC, MOD and SV of newborn rats (3-5 day old) and its changes by shortâterm cultivation (24 hrs) and mild hypoxia (5 hrs). Three varieties of tissue samples were analyzed (i) Freshly prepared tissue, used as control; (ii) cultures held under normoxic conditions and (iii) cultures exposed during cultivation to 5 h hypoxia. The gene expression was analyzed using the Affymetrix GeneChip RN-U34. The functional specificity of the cochlea regions are well reflected by the gene expression pattern. For example, the basal expression of Organ of Corti is characterized by genes involved in potassium and calcium homeostasis. The MOD is characterized by genes involved in regulation of maturation of neuronal cells. The SV is characterized by genes involved in ion homeostasis and blood flow. Cultivation dramatically changes the gene expression. In contrast, mild hypoxia showed only small changes. The changes are interpreted as stress response. We used this analysis as screening approach to detect significant changes in gene expression and used the RT-PCR analyses of independent sister cultures for confirmation. Correlation analysis showed close relations of the expression levels when aliquots of the biological material were compared. However, comparisons of the expression levels received by microarray and by RT-PCR analysis of independent sister samples resulted in 70-90 % in similar predictions. This data indicate that possible discrepancies has its basis in biological sampling rather than factors of methodical errors on the level of RT-PCR. Experiment Overall Design: Pooled samples of the following cochlea preparations were studied: Experiment Overall Design: (i) Native preparations of Organ of Corti, modiolus and stria vascularis from newborn rats used as controls; (ii) Organotypic cultures of these preparations under normoxic conditions (24 h); (iii) Cultures which were exposed to 5 h hypoxia during cultivation. Samples were collected for total RNA immediately after preparation of the inner ear structures (native samples) or 24 h after onset of cultivation (normoxia, hypoxia).
2008-06-13 | E-GEOD-5446 | biostudies-arrayexpress
Project description:Studies using yeast have advanced our understanding of both replicative and chronological aging, leading to the discovery of longevity genes that have homologues in higher eukaryotes. Chronological lifespan in yeast is conventionally defined as the lifespan of a non-dividing cell. To date, this parameter has only been estimated under calorically restricted (CR) conditions, mimicked by starvation. Since post-mitotic cells in higher eukaryotes are rarely calorically-restricted, we sought to develop an alternative experimental system where non-dividing yeast would age chronologically, in the presence of excess nutrients. We report here on a system wherein alginate-encapsulated yeast are packed in a pH- and temperature-controlled bioreactor, then continuously fed non-limiting substrate for extended periods of time. We present demographic, physiological and genomic evidence indicating that after ~120 hrs, immobilized cells cease dividing, remain metabolically very active and retain >95% viability for periods of 17 days. Over the same time interval, starved planktonic cells, cultured using the same media, and also controlled for temperature and pH, retained < 1 % viability in both aerobic and anaerobic cultures,. Unlike planktonic yeast, continuously-fed immobilized cells hyper-accumulate glycogen. FACS analysis of SYTOX green-stained yeast confirms that immobilized cells completely arrest within 5 days of culture, and unlike starving planktonic cells, remain free thereafter of replicative stress and are non-apoptotic. This unusual state is supported by a global gene expression profile that is stable over time, repeatable across replicate experiments, and altogether distinct from planktonic cells cultured in the presence and absence of limiting nutrients. DNA expression profiling, performed here for the very first time on immobilized cells, reveals that glycolytic genes and their trans-acting regulatory elements are upregulated, as are genes involved in remodeling the cell wall and resisting stress; by contrast, many genes that promote cell cycle progression and carry out oxidative metabolism are repressed. Stress resistance transcription factor MSN4 and its upstream effector RIM15 are conspicuously upregulated in the immobilized state, suggesting that nutrient-sensing pathways may play a role in cell viability and longevity when yeast are immobilized and placed in prolonged culture under calorically-unrestricted conditions. The cell cycle arrest in the immobilized state is mediated by RIM 15. Over the time-course of our experiments, well-fed, non-diving immobilized cells do not appear to age. Keywords: comparison of growth states and a time course of immobilized cells Nine growth conditions or time points with two to four replicates each
2011-03-31 | E-GEOD-21187 | biostudies-arrayexpress