The Influence of Melatonin on the Daily 24-h Rhythm of Putative Reference Gene Expression in White Adipose Tissues

In adipose tissue, the expression of hundreds of genes exhibits circadian oscillation, which may or may not be affected by circulating melatonin levels. Using control and pinealectomized rats, we investigated the daily expression profile ofActb,Hprt-1,B2m, andRpl37a, genes that are commonly used as reference genes for reverse transcription quantitative polymerase chain reaction (RT-qPCR), in epididymal (EP), retroperitoneal (RP), and subcutaneous (SC) adipose tissues. In control rats,Actbexpression presented a daily oscillation in all adipose tissues investigated,Hprt-1showed 24-h fluctuations in only RP and SC depots,B2mwas stable over 24 h for EP and RP but oscillated over 24 h in SC adipose tissue, andRpl37apresented a daily oscillation in only RP fat. In the absence of melatonin, the rhythmicity ofActbin all adipose depots was abolished, the daily rhythmicity ofHprt-1andB2mwas disrupted in SC fat, the peak expression ofRpl37aandHprt-1was delayed, and the amplitude ofRpl37awas reduced in RP adipose tissue. Collectively, our results demonstrate that the expression of putative reference genes displays a daily rhythm influenced by melatonin levels in a manner specific to the adipose depot. Thus, the proper standardization and daily profile expression of reference genes should be performed carefully in temporal studies using RT-qPCR analysis. (AU)