Peripheral Clock Gene Entrainment by Cortisol
The objective of this study is to assess the entrainment of peripheral blood leukocyte (PBL) clock genes by cortisol and evaluate its entraining characteristics. We demonstrate that the circadian characteristics of the peripheral entrainer (cortisol) have significant implications on the dynamics of clock genes. In the presence of a normal circadian pattern in cortisol, the PBLs are entrained to a coherent diurnal pattern. As the amplitude of cortisol's circadian rhythm is decreased slightly, the PBL oscillators remain entrained but undergo a phase shift relative to cortisol's phase. And as the amplitude is further decreased, eventually the PBLs fall out of sync and produce a flat ensemble average. Understanding the relationship between the pattern of cortisol secretion and its downstream effects on peripheral clock genes lays a foundation for studying the implications of dysregulated cortisol secretion (most notably under stress) on the function of PBLs.
To study the effect of cortisol in peripheral clock genes, we propose a unified model involving the circadian production of cortisol, cortisol signal transduction, and clock gene oscillations. We explore the “two rates” model of circadian cortisol production (Chakraborty, Krzyzanski et al. 1999) to generate the diurnal pattern in plasma cortisol concentration by switching between a high secretion rate during the morning when cortisol concentration is increasing and a low secretion rate the rest of the day when cortisol concentration decreases to a low resting level. Cortisol in plasma moves into the cytoplasm of PBLs, where it binds to glucocorticoid receptor (GR), forming an activated drug-receptor complex which subsequently translocates into the nucleus and acts as a transcription factor, regulating a wide range of genes (Yao, DuBois et al. 2006).
We further hypothesize that the peripheral circadian gene network involves a limited set of critical genes, including Per1-3 and Cry1-2, which, in collaboration with the CLOCK/BMAL1 heterocomplex, form an orchestrated feedback loop that leads to a circadian rhythm. Since Per1 has a glucocorticoid responsive element (GRE) in its promoter region and is known to be responsive to glucocorticoids (Yamamoto, Nakahata et al. 2005), we hypothesize that the primary link between cortisol's circadian rhythms and PBL circadian rhythms is the transcriptional activation of Per1 by the cortisol-GR activated complex. This circadian model includes an equation representing Per/Cry gene transcripts, which was modified by including an additive term in the Per/Cry equation representing stimulus of Per1 transcription by activated GR.
The purpose of our modeling effort is to evaluate the implications of alterations in the circadian patterns of cortisol release in the entrainment of clock genes. Our results demonstrate that under homeostatic release of cortisol, robust entrainment of clock genes is observed, as well as full synchronization of the ensemble of PBLs. As the entraining characteristics of cortisol are dysregulated, through a reduction in the amplitude of the circadian pattern (a hallmark of stress), individual cells' clock genes fall out of phase until eventually they become desynchronized. The loss of synchronization is hypothesized to play a significant role in the attenuated control effects clock genes exert on the homeostatic response.
Preliminary model predictions are depicted in Figure 1. Our model demonstrates the ability of cortisol's circadian variability to synchronize clock genes (Rsyn = 1) in an amplitude-dependent manner. It predicts that a population of cells falls out of sync in the absence of a systemic cortisol cue, whereas a circadian cortisol rhythm induces synchronization of PBL clock genes. Furthermore, as the entrainer (cortisol) loses its circadian rhythmicity, our model predicts a regime of phase shift in peripheral clock genes relative to cortisol before synchronization is lost at very low circadian amplitudes. Similar phase shifted responses have been observed in murine adipose tissues, where restricted feeding provokes a disruption of glucocorticoid rhythmicity along with phase shifts in the expression of peripheral clock genes (Zvonic, Ptitsyn et al. 2006).
Cortisol rhythmicity plays an important role in entraining peripheral clock genes and affecting the dynamics of the peripheral clock network. Our work has significant clinical implications as cortisol is a key component of the stress response and regulates the transcription of anti-inflammatory genes. In human endotoxemia experiments, cortisol is acutely elevated and peripheral clock genes in PBLs are suppressed (Haimovich, Calvano et al. 2010), which may have important effects on the inflammatory response, given the interplay between clock genes and cytokine production in peripheral immune cells (Coogan and Wyse 2008). Furthermore, in chronic stress when circadian rhythms in cortisol can be lost (Lowry and Calvano 2008), peripheral circadian rhythms may be similarly desynchronized or shifted as predicted by our model. The work presented here will serve as a foundation for studying these interconnections between circadian rhythms and the inflammatory response.
Figure 1: The degree of synchronization Rsyn (ratio of the variance of the mean field to the mean variance of each oscillator, Rsyn=1 is synchronized, 0 otherwise) and the standard deviation of the distribution of cells' phases both change as a function of cortisol's amplitude.
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