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Characterization of HT-29 cells cultured in NanoCulture plates

ABSTRACT: To characterize cells cultured in 3D culture system using NanoCulture plates (NCPs) (Scivax) and in 2D culture, we examined gene expression in HT-29 cells on day 1 or day 7 on NCPs or 2D monolayer culture, using DNA microarrays. From our study, tumour cells grown on NCPs were morphologically different from cells in 2D monolayers, and the cells on NCPs actively migrated and aggregated to form multicellular spheroids (MCSs). To characterize this cell migration and intercellular adhesion, we compared gene expression in HT-29 cells on day 1 on NCPs and 2D monolayer culture using DNA microarrays. We could not find any significant difference in gene expression related to cell migration or cell-cell adhesion between cells grown on NCPs and as 2D monolayers on day 1, showing that cells on NCPs migrated and aggregated without any changes in gene expression (3D Day1 vs 2D Day1 2Fold change.xls). We next characterized mature 3D MCSs on NanoCulture plates, by comparing the gene expression profile of MCSs on day 7 and day 1 (3D Day7 vs 3D Day 1 2Fold change.xls). On day 7, we found a statistically significant increase in the expression of genes related to multicellular organization, intercellular signalling and the response to hypoxia. DNA microarray analysis also showed that the expression of genes that have been reported to be target genes for the transcription factor hypoxia-inducible factor 1 (HIF-1) (Ke, Q. & Costa, M. Hypoxia-inducible factor-1 (HIF-1). Mol Pharmacol 70, 1469-1480 (2006); Semenza, G.L. Defining the role of hypoxia-inducible factor 1 in cancer biology and therapeutics. Oncogene 29, 625-634.) was increased on day 7. When compared day 7 with 2D culture cells, the same tendency was observed (3D Day7 vs 2D Day 7 2Fold change.xls). Overall design: HT-29 cells were maintained routinely as 2D monolayers in a humidified atmosphere of 5% CO2 in air at 37 °C in the growth medium recommended by suppliers. Exponentially growing cells were used in experiments, after trypsinization to detach them from the plates. Viable cells were counted using trypan blue dye-exclusion. For experiments, cells were seeded in 96-well plates at 1×10^4 cells/100 ul of NanoCulture Medium-M (Scivax) in either NCPs (Scivax), or in polystyrene plates for 2D monolayers, for 1 day or 7 days. Cells were collected and used for RNA extraction and DNA microarray analysis.

INSTRUMENT(S): Agilent-014850 Whole Human Genome Microarray 4x44K G4112F (Feature Number version)

ORGANISM(S): Homo sapiens  

SUBMITTER: Yukie Yoshii  

PROVIDER: GSE23773 | GEO | 2010-08-25



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