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Selection of housekeeping genes for gene expression studies in a rat model of irinotecan-induced mucositis.

AbstractBACKGROUND/AIMS:
Mucositis is the term used to describe damage caused by chemotherapy to mucous membranes of the alimentary tract. RT-PCR has recently been utilised to determine the molecular events that occur in mucositis. As this method relies on the use of a validated endogenous control, this study aims to validate commonly used housekeeping genes in an irinotecan-induced mucositis model.
METHODS:
Rats were administered irinotecan and sacrificed at different time points, in particular 1, 24, 72 and 144 h following treatment. Histopathological damage was assessed by haematoxylin and eosin staining. RT-PCR was used to evaluate the expression of 11 housekeeping genes. Expression stability was determined by the Normfinder program. Matrix metalloproteinase 2 was used as a target gene to validate the appropriateness of the top-ranking housekeeping gene.
RESULTS:
For normalisation to multiple housekeeping genes, the most stable combination across all time points in the jejunum was Ywhaz/UBC and in the colon UBC/β-actin. SDHA and GAPDH were the most variable genes in the jejunum and colon where they were 4.4 and 3.2 fold upregulated following irinotecan, respectively.
CONCLUSIONS:
For normalisation of irinotecan-induced mucositis gene expression studies, a combination of Ywhaz/UBC and UBC/β-actin should be used in the jejunum and colon, respectively. UBC is the most favourable if restricted to a single housekeeping gene across all time points.
AuthorsNoor Al-Dasooqi, Joanne M Bowen, Rachel J Gibson, Richard M Logan, Andrea M Stringer, Dorothy M Keefe
JournalChemotherapy (Chemotherapy) Vol. 57 Issue 1 Pg. 43-53 ( 2011) ISSN: 1421-9794 [Electronic] Switzerland
PMID21282945 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
CopyrightCopyright © 2011 S. Karger AG, Basel.
Chemical References
  • 14-3-3 Proteins
  • Actins
  • Molecular Chaperones
  • Ubiquitin C
  • Ywhaz protein, rat
  • Irinotecan
  • Glyceraldehyde-3-Phosphate Dehydrogenases
  • Succinate Dehydrogenase
  • Matrix Metalloproteinase 2
  • Camptothecin
Topics
  • 14-3-3 Proteins (genetics, metabolism)
  • Actins (genetics, metabolism)
  • Animals
  • Camptothecin (analogs & derivatives, toxicity)
  • Colon (metabolism)
  • Disease Models, Animal
  • Gene Expression Regulation (drug effects)
  • Glyceraldehyde-3-Phosphate Dehydrogenases (genetics, metabolism)
  • Irinotecan
  • Jejunum (metabolism)
  • Male
  • Matrix Metalloproteinase 2 (genetics, metabolism)
  • Molecular Chaperones (genetics, metabolism)
  • Mucositis (chemically induced, genetics, metabolism)
  • Rats
  • Succinate Dehydrogenase (genetics, metabolism)
  • Ubiquitin C (genetics, metabolism)
  • Up-Regulation

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