Inter-subject differences in constitutive expression levels of the clock gene in man

Abstract

Circadian rhythms are generated, both at the level of the whole organism and at the cellular level, by biological clocks involving a set of clock genes. We previously performed a randomised, placebo-controlled, double-blind trial examining the effect of six months of pioglitazone (30 mg per day) therapy on neointima volume after coronary stenting in patients with coronary artery disease but without diabetes. In a subgroup of 15 patients from each group, a whole blood sample was taken at the beginning of the trial and at an eight-week clinical follow-up for isolation of total RNA using a PAXgene system. Using real time RT-PCR with relative quantitation, we investigated whether pioglitazone treatment altered clock gene expression in RNA extracted from peripheral white blood cells (PBCs). No significant changes in clock gene expression were noted in either placebo (99±45%) or pioglitazonetreated subjects (101±35%) after eight weeks. These data potentially extend previous findings that the clock gene is constitutively expressed over 24 hours to eight weeks. We observed a large range of inter-subject differences in expression levels of the clock gene. The difference between the mean value for the lowest expression individual (ΔCT 8.8) and the highest expression individual (ΔCT 3.7) revealed an approximately 34-fold difference in relative clock gene expression levels. These differences may arise through differences in light exposure, polymorphic variants of the clock gene affecting gene expression and/or post-transcriptional regulation. They could influence susceptibility to disruption of normal circadian rhythms, which occur in pathophysiological conditions pertaining to diabetes and cardiovascular disease.