Transcription profiling of 25 breast cancer cell lines grown in 2D and 3D tissue culture conditions
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Curry MC
PROVIDER: S-ECPF-TABM-244 | biostudies-other | 2012 Aug
REPOSITORIES: biostudies-other
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Project description:A comparison of gene expression profiles for 25 breast cancer cell lines grown in 2D and 3D tissue culture conditions
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Project description:Parvin-beta is a focal adhesion protein downregulated in human breast cancer cells. Loss of Parvin-beta contributes to increased integrin-linked kinase activity, cell-matrix adhesion, and invasion through the extracellular matrix in vitro. The effect of ectopic Parvin-beta expression on the transcriptional profile of MDA-MB-231 breast cancer cells, which normally do not express Parvin-beta was evaluated. Particular emphasis was placed upon propagating MDA-MB-231 breast cancer cells in three-dimensional culture matrices. Gene expression profiles of vector control and Parvin-beta transfected MDA-MB-231 cells cultured on (A) monomeric type I collagen coated plastic, (B) embedded in a type I collagen gel, and (C) embedded in basement membrane (growth factor reduced Matrigel), were compared. Interestingly, Parvin-beta re-expression in MDA-MB-231 cells increased the mRNA expression, serine 82 phosphorylation (mediated by CDK9), and activity of the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARgamma) and a concomitant increase in lipogenic gene expression as a downstream effector of PPARgamma. Importantly, Parvin-beta suppressed breast cancer growth in vivo with associated decreased proliferation. These data suggest that Parvin-beta might influence breast cancer progression.. Experiment Overall Design: The MDA-MB-0231 transfectants were cultured for 7 days in each growth condition and total RNA isolated using Trizol reagent (Invitrogen). Genes either upregulated or downregulated in Parvin-beta transfectants versus vector control cells were compared among the 3 different growth conditions. Venn diagrams were generated using GeneSpring software and used to produce gene lists that were unique to a particular growth condition, or shared between two or all three growth conditions. The focus was primarily on gene expression alterations observed in the 3D collagen type I and 3D Matrigel conditions.
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Project description:Live-cell assays to measure cellular function performed within 3D cultures have the potential to elucidate the underlying processes behind disease progression and tissue formation. Cells cultured in 3D interact and remodel their microenvironment and can develop into complex structures. We have developed a transcription factor (TF) activity array that uses bioluminescence imaging (BLI) of lentiviral delivered luminescent reporter constructs that allows for the non-invasive imaging of TF activity in both 2D and 3D culture. Imaging can be applied repeatedly throughout culture to capture dynamic TF activity, though appropriate normalization is necessary. We investigated in-well normalization using Gaussia or Renilla luciferase, and external well normalization using firefly luciferase. Gaussia and Renilla luciferase were each unable to provide consistent normalization for long-term measurement of TF activity. However, external well normalization provided low variability and accounted for changes in cellular dynamics. Using external normalization, dynamic TF activities were quantified for five TFs. The array captured expected changes in TF activity to stimuli, however the array also provided dynamic profiles within 2D and 3D that have not been previously characterized. The development of the technology to dynamically track TF activity within cells cultured in both 2D and 3D can provide greater understanding of complex cellular processes.
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Project description:Gene expression analysis was performed through RNA-Sequencing on samples prepared from a panel of four CRC cell lines (HT29, HCT116, LS174T, LS513) grown in either 2D or 3D cell culture conditions. Our gene expression data reveal that more than 2000 gene expression profiles are affected by the growth conditions for each cell lines even if samples prepared from each cell line still group together. Our data also demonstrate that hierarchical clustering mostly relies on the MSS/MSI phenotypes of the CRC cells.
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Project description:Parvin-beta is a focal adhesion protein downregulated in human breast cancer cells. Loss of Parvin-beta contributes to increased integrin-linked kinase activity, cell-matrix adhesion, and invasion through the extracellular matrix in vitro. The effect of ectopic Parvin-beta expression on the transcriptional profile of MDA-MB-231 breast cancer cells, which normally do not express Parvin-beta was evaluated. Particular emphasis was placed upon propagating MDA-MB-231 breast cancer cells in three-dimensional culture matrices. Gene expression profiles of vector control and Parvin-beta transfected MDA-MB-231 cells cultured on (A) monomeric type I collagen coated plastic, (B) embedded in a type I collagen gel, and (C) embedded in basement membrane (growth factor reduced Matrigel), were compared. Interestingly, Parvin-beta re-expression in MDA-MB-231 cells increased the mRNA expression, serine 82 phosphorylation (mediated by CDK9), and activity of the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARgamma) and a concomitant increase in lipogenic gene expression as a downstream effector of PPARgamma. Importantly, Parvin-beta suppressed breast cancer growth in vivo with associated decreased proliferation. These data suggest that Parvin-beta might influence breast cancer progression.. Experiment Overall Design: The MDA-MB-0231 transfectants were cultured for 7 days in each growth condition and total RNA isolated using Trizol reagent (Invitrogen). Genes either upregulated or downregulated in Parvin-beta transfectants versus vector control cells were compared among the 3 different growth conditions. Venn diagrams were generated using GeneSpring software and used to produce gene lists that were unique to a particular growth condition, or shared between two or all three growth conditions. The focus was primarily on gene expression alterations observed in the 3D collagen type I and 3D Matrigel conditions.
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