ABSTRACT: Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression. Two-condition experiment, Normoxic MSCs vs. Hypoxic MSCs.
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression. Overall design: Two-condition experiment, Normoxic MSCs vs. Hypoxic MSCs.
Project description:To determine whether hypoxia augments the activity of exosomes by modulating miRNAs, we use miRNA array profile to find the differential expression of miRNAs of exosomes in different culture conditions (normoxia, hypoxia, hypoxia supplemented with GW4869 which is an inhibitor of exosomes generation). Overall design: Three samples were processed for each kind of exosome that derived from normoxic-MSCs, hypoxic-MSCs, and hypoxic-MSCs suppled with GW4869 (N-EXO, H-EXO, H+G-EXO). H-Exosome was isolated from conditioned medium of MSCs under hypoxia condtion 24h. The fragmentation mixtures were hybridized to an Agilent- Mouse microRNA array 21.0
Project description:Transcriptional profiling of H1299 non-small cell lung carcinoma cells transfected with either wt p53 or mut(175) p53 driven by the 5xHRE promoter (5 repeats of hypoxia-inducible factor response elements) and treated for 16 h with normoxia (21% O2) or hypoxia(<0.1% O2). 5xHRE promoter ensures that p53 expression is induced in hypoxic conditions only. Goal was to determine the transcriptional response of p53 in hypoxia and the 175 p53 mutant was used as a control as it is DNA-binding defective and transcription-incompetent mutant. Four-condition experiment: wt p53-transfected H1299 cells treated with normoxia, mut p53-transfected H1299 cells treated with normoxia, wt p53-transfected H1299 cells treated with hypoxia, mut p53-transfected H1299 cells treated with hypoxia. Biological replicates: 1 normoxic sample with wt p53, 1 normoxic sample with mut p53, 3 hypoxic samples with wt p53, 3 hypoxic samples with mut p53.
Project description:In vivo profiling of hypoxic gene expression in gliomas using the hypoxia marker EF5 and laser-capture microdissection We have employed the hypoxia marker EF5 coupled with laser capture microdissection to isolate RNA from viable hypoxic and normoxic regions of 9L experimental gliomas. We report the identification of the hypoxic gene expression profile in a solid tumor using a prototypical hypoxia marker EF5, Laser Capture Microdissection (LCM) and Microarray Gene Expression Profiling (EF5-LCM-MGEP). We have utilized LCM to isolate total RNA from normoxic and hypoxic regions in the rat 9L glioma tumor model grown in its isogenic host (Fischer rat). Microarray analysis of the isolated RNA generated an in vivo hypoxia expression profile. For comparison, we also analyzed RNA samples isolated from rat 9L glioma cells in culture exposed to hypoxia or normoxia.
Project description:Hypoxia is a low oxygen condition that occurs in the developing tumor mass and that is associated with poor prognosis and resistance to chemo- and radio-therapy. The definition of the hypoxia gene signature is fundamental for the understanding of tumor biology, as in the case of neuroblastoma, the most common pediatric solid tumor. The issue of identifying a significant group of variables in microarray gene expression experiments is particularly difficult due to the typical high dimensional nature of the data and great effort has been spent in the development of feature selection techniques. Our main goal is to define a robust hypoxia gene signature in neuroblastoma cell lines. A set of 11 neuroblastoma cell lines were cultured under normoxic and hypoxic conditions for 18 hours, and their gene expression profiles were measured with Affymetrix GeneChip HG-U133 Plus 2.0. We used the l1-l2 regularization framework in order to select the significant probesets defining hypoxic versus normoxic cell lines. Experiment Overall Design: The expression profiles of 11 neuroblastoma cell lines under normoxia vs hypoxia were studied.
Project description:How cancer cells adapt to hypoxia during tumor development remains an important question. The hypothesis tested in the present study was that tumor cell-derived exosome vesicles (also known as microvesicles or extracellular vesicles) are mediators of hypoxia-dependent intercellular signaling in glioblastoma (GBM), i.e. highly aggressive brain tumors characterized by hypoxia and a vascular density that is among the highest of all human malignancies. In vitro hypoxia experiments and studies with patient materials reveal the enrichment in exosomes of hypoxia-regulated mRNAs and proteins, several of which were associated with poor patient prognosis. We show that cancer cell exosomes mediate hypoxia-dependent, phenotypic modulation of stromal cells in vitro and ex vivo, resulting in accelerated GBM tumor angiogenesis and growth in mice. These data suggest that exosomes constitute potent mediators of hypoxia-driven tumor development, and circulating multiparameter biomarkers of tumor hypoxia. U87 MG glioblastoma cells were grown at normoxic (21% oxygen) or hypoxic (1% oxygen) conditions for 48 hours. Conditioned media from normoxic and hypoxic cells were then used to isolate exosomes by differential centrifugation. Both cells and exosomes were lysed in Trizol reagent, and RNA was isolated.Total RNA from all samples (four types of samples in three biological repilicates) was subjected to genome-wide transcriptional analysis with Illumina HumanHT-12 V3.0 expression beadchip. Gene expression profile obtained from hypoxic U87 MG glioblastoma cells was compared to the profile of normoxic control cells. Analogically, gene expression profile obtained from hypoxic U87 MG cells was compared to the profile of exosomes secreted by normoxic U87 MG cells.
Project description:Transcriptional profiling of human amniotic fluid-derived mesenchymal stem cells (AF-MSCs) comparing normal condition-cultured AF-MSCs with hypoxia condition-culture AF-MSCs ES cells. The latter improves the proliferation and survival of AF-MSCs, and makes AF-MSCs secret more growth factors. Goal was to determine the effects of hypoxia condition on global AF-MSCs gene expression. Two-condition experiment, Nor AF-MSCs vs. Hypo AF-MSCs. Experimantal replicates: 2 (1-1 vs. 1-2; 2-1 vs. 2-2). Biological replicates: 2 normal replicates, 2 treated replicates.
Project description:Hypoxic microenvironment plays important roles in the progression of solid tumors including oral squamous cell carcinoma (OSCC). Long noncoding RNAs (lncRNAs) have gained much attention in the past few years. However, it is not clear whether lncRNAs could regulate hypoxia-adaptation of OSCC, and which lncRNAs participate in this process. Using an lncRNA microarray, we analyzed the aberrant lncRNA expression profiles in OSCC tissues compared with paired normal oral mucosa as well as in hypoxic OSCC cells compared with normoxic OSCC cells. Our microarray also identified 942 lncRNAs that were up-regulated and 507 lncRNAs that were down-regulated in cells cultured under hypoxia compared with those cultured under normoxia (Fold Change >= 2.0, P-value <= 0.05). An OSCC cell line Cal-27 was cultured under either 20% O2 (normoxic) or 1% O2 (hypoxic) conditions. Three samples from each group were obtained for microarray study.
Project description:To define ncRNA expression in hypoxic endothelial cells, we applied pro-angiogenic hypoxia to cultured endothelial cells. Afterwards, total RNA was isolated and underwent RNA-seq analysis. HUVECs were subjected to normoxic or hypoxic (0.1-0.2% O2) cell culture conditions.
Project description:Human HeLa cells were either kept in a hypoxic environment or under normoxic conditions. 137 proteins were found to be regulated, with maximum alteration of 18-fold. In particular, three clusters of differentially regulated proteins were identified, showing significant up-regulation of glycolysis and down-regulation of mitochondrial ribosomal proteins and translocases. This interaction is likely orchestrated by HIF-1. We also investigated the effect of hypoxia on the cell cycle, which is arrested at a restriction point in G1, and shows a prolonged S phase under these conditions.