Profiling candidate housekeeping genes for data normalization in chronic ethanol treated rat liver regeneration model

Abstract

This study focuses on selecting suitable housekeeping genes (HKGs) to normalize gene expression data in the regenerating liver of the rats subjected to chronic ethanol treatment. Rats were fed a liquid diet containing 36% of total calories derived from ethanol for 5 weeks; for pair‐fed controls ethanol calories were replaced by carbohydrates. After five weeks, rats were subjected to 70% PHx and liver samples were collected at 1, 6, 12, 24, 36 and 48 h after the surgery.Our global gene expression analysis indicates that almost 50% of hepatic genes are differentially expressed within the first 24 h after PHx. The ideal HKGs should be expressed at a constant level during different stages of liver regeneration and be unaffected by chronic ethanol treatment. We analyzed expression levels of 35 candidate HKGs using a high throughput microfluidic qPCR platform. This set included both traditional HKGs (Actb, Gapdh) and novel candidates selected based on our global gene expression data. Gene expression analysis using modified geNorm algorithm showed that only five of these genes could be considered suitable HKGs in this model. Results of our gene expression data analysis were significantly affected by the choice of HKGs. Our newly characterized HKGs improved the patterns identified for the cell cycle pathway demonstrating the functional validity of our approach.Supported by AA018873, AA008714, AA014986, AA017261.