Gene expression profiling of human glioblastoma cells under culture with endothelial cell condition media using Affymetrix arrays
Description
This study investigates the gene expression changes in human glioblastoma cells subjected to endothelial cell condition media.
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Samples were collected and processed at the GENOMIC platform of Institut Cochin, Paris. RNA was extracted, labeled, and hybridized onto Affymetrix GeneChips. Data were processed using standard normalization pipelines. The study includes 4 samples divided into [control and treated groups. Each condition was replicated 2 times. The goal was to identify differentially expressed genes associated with EC coculture. Patient-derived mesenchymal GSCs were maintained as non-adherent tumorspheres in NS34 serum-free media (DMEM/F12 with N2, G5, and B27 supplements, plus Glutamax and antibiotics, Life Technologies). They were with the conditioned media harvested from human brain endothelial cells. Total RNA was extracted using RNAesay kit Qiagen, RNA integrity was assessed using Bioanalyser 200 ng of RNA was labeled using [Affymetrix GeneChip WT PLUS kit / 3' IVT kit], according to the manufacturer’s protocol. After control of fragmentation using Bioanalyzer 2100, cDNA is then hybridized to GeneChip® MTA1.0 ST (Affymetrix) at 45°C for 17 hours After overnight hybridization, chips are washed on the fluidic station FS450 following specific protocols (Affymetrix) and scanned using the GCS3000 7G The scanned images are then analyzed with Expression Console software (Affymetrix) to obtain raw data (cel files) and metrics for Quality Controls. RMA normalization is performed using TAC4.0 (Affymetrix) with default parameters Values represent log₂ normalized expression intensities