Selection of Housekeeping Genes for Gene Expression Studies in a Rat Model of Irinotecan-Induced Mucositis

Background/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/beta-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/beta-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. Copyright (C) 2011 S. Karger AG, Basel