Maternal low-protein diet alters the expression of real-time quantitative polymerase chain reaction reference genes in an age-, sex-, and organ-dependent manner in rat offspring

Altered perinatal environment, often manifested as low birth weight, is thought to contribute to greater susceptibility for hypertension, hyperlipidemia, and diabetes as a result of epigenetic modifications and alteration of transcriptional activity for key genes. Real-time polymerase chain reaction is a useful technique for the quantitative determination of differences in transcriptional activity. Real-time quantitative polymerase chain reaction data analyses require normalization of transcriptional activity of target genes to an endogenous control, usually a reference gene. In response to reports of altered expression of reference genes in various experimental models, we hypothesized that adverse perinatal environment alters reference gene expression. We examined the expression of the following reference genes in the offspring of a rodent maternal low-protein diet model: β-actin, hypoxanthine phosphoribosyltransferase 1, TATA-box-binding protein, glyceraldehyde-3-phosphate dehydrogenase, and glucuronidase-β in brain, heart, kidneys, and intestines. We found altered expression in brain, heart, and kidneys for each of the reference genes measured; these effects were age, organ, and sex dependent. Glyceraldehyde-3-phosphate dehydrogenase and glucuronidase-β were found to be the least affected by these variables, whereas hypoxanthine phosphoribosyltransferase 1 was the most inconsistent. Our findings underscore the importance of empirical determination of a reliable reference gene for real-time polymerase chain reaction studies in the low-protein diet model.