Unknown,Transcriptomics,Genomics,Proteomics

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Affymetrix GeneChip™ Canine Gene 1.1ST Array of Quiescent and activated canine valvular interstitial cell under TGFB1 and SB431542 treatment

ABSTRACT: Quiescent canine valvular interstitial cells (n=3) and activated valvular interstitial cells (n=3) were grown in 2% FBS. Quiescent cells were treated with either TGFB1 or vehicle for 3 days and activated cells were treated with SB431542 for 4 days. mRNA was extracted using the qiagen RNeasy minikit and assessed by the agilent 2200 bioanalsyser. all samples passed quality control and transcriptomic analysis was performed by edinburgh genomics using the Affymetrix GeneChip™ Canine Gene 1.1ST Array. Results were reported as CEL files and then analysed in Affymetrix expression console (version 1.4.1.46) using the canine 1.1ST library files from Affymetrix. Robust Multi-array Average (RMA) was used to perform gene level normalisation and signal level summarisation.Following this, quality control assessment was performed on the data set. This included assessment of hybridisation (spike in) controls, labelling (poly-A) controls and area under the curve (AUC) control, probe cell intensity boxplots, a signal histogram graph and principle component analysis (PCA) plot. Following this assessment, the datasets were exported with annotation as a text file or as a CHT file. Affymetrix transcriptome analysis console (version 3.1.0.5) was used to perform paired or unpaired one-way analysis of variance (ANOVA).

INSTRUMENT(S): GeneTitan® multi-channel instrument, RNeasy minikit

ORGANISM(S): Canis lupus familiaris

SUBMITTER: Greg Markby

PROVIDER: E-MTAB-8096 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Canine mast cell tumour proliferation depends to a large extent on the activity of KIT, a tyrosine kinase receptor. Inhibitors of the KIT tyrosine kinase have recently been introduced and successfully applied as a therapeutic agent for this tumour type. However, little is known on the downstream target genes of this signalling pathway and molecular changes after inhibition. Transcriptome analysis of the canine mast cell tumour cell line C2 treated for up to 72 hours with the tyrosine kinase inhibitor masitinib identified significant changes in the expression levels of approximately 3500 genes representing 16% of the canine genome. Approximately 40% of these genes had increased mRNA expression levels including genes associated with the pro-proliferative pathways of B- and T-cell receptors, chemokine receptors, steroid hormone receptors and EPO-, RAS and MAP kinase signalling. Proteome analysis of C2 cells treated for 72 hours identified 24 proteins with changed expression levels, most of which being involved in gene transcription, e.g. EIA3, EIA4, TARDBP, protein folding, e.g. HSP90, UCHL3, PDIA3 and protection from oxidative stress, GSTT3, SELENBP1. Transcriptome and proteome analysis of neoplastic canine mast cells treated with masitinib confirmed the strong important and complex role of KIT in these cells. Approximately 16% of the total canine genome and thus the majority of the active genes were significantly transcriptionally regulated. Most of these changes were associated with reduced proliferation and metabolism of treated cells. Interestingly, several pro-proliferative pathways were up-regulated which may represent attempts of masitinib treated cells to activate alternative pro-proliferative pathways. These pathways may contain hypothetical targets for a combination therapy with masitinib to further improve its therapeutic effect. The present study aimed at identifying the transcriptional and translational responses of neoplastic canine mast cells to the tyrosine kinase inhibitor masitinib. To this end, C2 cells, a cell line with a tandem duplication in the juxtamembrane unit and thus constitutively activated KIT, were treated with masitinib and changes in the global mRNA and protein expression levels were characterized.

Dataset Information

Affymetrix GeneChip™ Canine Gene 1.1ST Array of Quiescent and activated canine valvular interstitial cell under TGFB1 and SB431542 treatment

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