SUN-020 Analyses Of Clock Gene Expressions In Peripheral Leukocytes In Hypercortisolism

Background: Hypercortisolism causes metabolic disorders such as hyperglycemia and dyslipidemia. Recent studies have shown relationships between impairment of peripheral circadian clock function and the development of metabolic diseases. However, the association between phenotypes and peripheral circadian regulations in overt hypercortisolism and subclinical hypercortisolism clinically has not been elucidated. This study examined peripheral clock-related gene expressions of patients with hypercortisolism in regulation of involvement of glucocorticoid receptors. Methods: Blood samples were obtained from four patients with hypercortisolism including one patient of Cushing syndrome due to adrenal tumor, two patients of Cushing disease, and one patient due to ectopic ACTH producing tumor and five comparatively young volunteers as normal subjects at 08:00 and 20:00. Peripheral bone mononuclear cells (PBMCs) were extracted by Ficoll density gradient methods. Transcript levels of clock genes (Clock, Bmal1, Per1,Cry1, and ROR), glucocorticoid receptor (GR) and GILZ were determined by real-time quantitative PCR. Results: Serum cortisol levels were significantly higher in samples from 08:00 than 20:00 in normal subjects (p<0.05) and these levels from patients of hypercortisolism were significantly higher than normal subjects (p<0.01). Diurnal variation of cortisol levels in the patients was not observed. The expressions of Clock and Bmal1 mRNA in normal subjects showed higher at 20:00 than at 08:00 and Per1 mRNA was significantly lower at 20:00 than 08:00 (p<0.05), exhibiting rhythmicity in the PBMCs of 5 normal subjects. The expression levels of these clock-associated mRNAs and GR were significantly (p<0.01) lower in the patients than in normal subjects at 08:00 and 20:00. Moreover, the amplitudes of mRNA expression rhythms of clock-related genes tended to diminish in the patients. Multivariate analysis showed a positive correlation between Clock and Bmal1 in normal subjects and between Per1 and Cry1 significantly (p <0.01, p<0.01), and a same correlation was observed in hypercortisolem (P <0.01, p <0.001). Clock-associated genes and GR expressions showed a significant negative correlation with cortisol levels. Conclusions: These results suggest that rhythmic mRNA expression of clock genes is impaired in PBMCs of patients of hypercortisolism. Interestingly, molecules forming dimerization at the same phase in the circadian loops showed coordinated gene expressions in hypercortisolism. The expressions of the peripheral clock genes in the patients were markedly reduced, which was thought to be caused by a remarkable cortisol excess. Expressions of peripheral clock genes may be useful markers in predicting metabolic complications and therapeutic effects as well as leading to elucidate the pathology of metabolic disorders in subclinical hypercortisolism.