Circadian Misalignment Augments Markers of Insulin Resistance and Inflammation, Independently of Sleep Loss

Rachel Leproult; Ulf Holmbäck; Eve Van Cauter

Disclosures

Diabetes. 2014;63(6):1860-1869. 

In This Article

Discussion

We designed the current study to determine whether circadian misalignment has adverse cardiometabolic effects independently of sleep loss. Our experimental strategy was successful, because the participants obtained nearly identical amounts of sleep irrespective of exposure to circadian alignment or misalignment. Sleep loss reduced SI and DI, but the reduction in SI was nearly twice as large when the week of sleep restriction included 4 days with bedtimes delayed by 8.5 h than when the center of the sleep period remained fixed. Despite the greater decrease in SI in participants exposed to circadian misalignment, the β-cell response was similar to that observed in participants in whom synchrony between behavioral and endogenous rhythms was maintained. These findings demonstrate that circadian misalignment can have adverse effects on insulin action and insulin release that are distinct from those imparted by sleep loss alone. Similarly, the levels of hsCRP, a marker of systemic inflammation and a predictor of cardiovascular disease risk, were increased after sleep restriction and to a greater extent in the participants who experienced circadian misalignment.

Our protocol involved restricting bedtimes to build sleep pressure and thereby achieving virtually identical amounts of sleep in both arms of the study. Thus, we controlled experimentally for total sleep time, and caloric intake was also nearly identical in both arms of the protocol. We concluded that circadian misalignment has intrinsic adverse cardiometabolic effects. A study design where bedtimes would not have been restricted would have led to a greater amount of sleep loss in the circadian misalignment group, with a need to control statistically for total sleep time in the analysis, as performed in the only previous experimental study that attempted to demonstrate adverse cardiometabolic effects of circadian misalignment.[14] In this previous study, total sleep time varied according to the degree of misalignment, and the conclusion that circadian misalignment has adverse cardiometabolic consequences relied on the statistical significance of alignment versus misalignment while controlling for sleep efficiency as a covariate in the statistical analysis. The current study provides instead a direct experimental demonstration.

The shift in circadian time could have influenced our estimations of the magnitude of the change in SI between the aligned and misaligned conditions. This issue was carefully considered when the protocol was designed. Indeed, a phase delay was used rather than a phase advance to create circadian misalignment. In healthy nonobese individuals, SI is higher in the morning than 8 to 10 h later, in the late afternoon or early evening.[29] Our participants in the circadian misalignment condition experienced a delay of the melatonin onset of about 3 h, but the clock time of the IVGTT remained fixed at 0900. Therefore, relative to internal circadian time, SI was assessed earlier—rather than later—in the biological day. In addition, there is evidence that peripheral clocks in metabolically relevant tissues shift at a slower rate than the central circadian pacemaker.[30] Thus, the delay in the diurnal variation of SI was likely less than 3 h. If our estimations of morning SI were affected by this modest shift of peripheral circadian time, it would therefore be in a direction that would result in a lower estimation of SI in the circadian misalignment than in the circadian alignment condition. Therefore, if affected at all by the shift of circadian time, the difference in the decrease of SI between the two conditions is underestimated, not overestimated.

We examined multiple putative mechanisms mediating the adverse metabolic effect of circadian misalignment. Previous studies in healthy young adults have shown that experimental reductions of sleep quality without change in sleep duration, by nearly complete suppression of SWS[31] or by severe sleep fragmentation across all sleep stages,[32] can result in decreases in SI that approximate the effect size of circadian misalignment observed in the current study. However, the macrostructure of sleep, as assessed in the current study by the total amounts of SWS and REM sleep during the week of sleep restriction, was similar in both groups. Sleep restriction did not result in an increase in SWS in either group, and REM sleep was markedly and similarly suppressed in both groups. Therefore, that alterations of sleep quality played a major role in the adverse metabolic consequences of circadian misalignment seems unlikely.

Average daily caloric intake was excessive but similar in both study groups, as were the timings of breakfast, lunch, and dinner. However, when participants in the circadian misalignment group were exposed to the four shifted nights, the overnight fast was interrupted by a small scheduled nighttime meal with continued access to snacks during the remainder of the night. Over the 7 days of sleep restriction, the proportion of daily caloric intake during the nighttime in the circadian misalignment group was threefold higher than in the circadian alignment group. The night eating syndrome in humans[33,34] and a shift of food intake from the active phase to the rest phase in laboratory rodents[35,36] have been associated with adverse metabolic consequences. Whether the disruption of dietary intake that occurred during shifted nights might have caused a further 20–30% decrease in SI compared with a normal 12-h overnight fast is an important question with major public health implications that will need to be rigorously addressed. Importantly, weight gain was similar under both conditions, and we verified that changes in body weight were not a significant predictor of changes in SI or β-cell response.

The durations of exposure to light and dark were identical in both groups, with similar levels of light intensity during periods of wakefulness and total darkness during periods of sleep. Exposure to light during the biological night during the 4 days with bedtime periods delayed by 8.5 h resulted in a delay of the DLMO of ~3 h in all but two participants. The demographics and baseline DLMO and melatonin levels of the two participants who did not shift were similar to those of the remainder of the group. Further, these two individuals experienced qualitatively and quantitatively similar changes in SI as the other participants, suggesting that the timing of the melatonin rhythm was not a major determinant of the metabolic effects of circadian misalignment.

Consistent with previous studies of partial sleep deprivation in healthy young men,[37,38] sleep restriction without circadian disruption resulted in a marked elevation of serum hsCRP levels in men. In those exposed to circadian misalignment, the relative increase was more than twice as large, revealing an adverse effect of circadian disruption on this sensitive marker of cardiovascular risk. Inflammation could be involved in the mechanisms linking sleep loss and circadian disruption to alterations in glucose metabolism.

Novel concepts regarding organization of the mammalian circadian system and its interaction with metabolism have emerged during the past 10 years.[39–43] The molecular mechanism generating circadian rhythmicity within pacemaker neurons of the SCN has been identified as a transcriptional–translational feedback loop of activators and repressors, including CLOCK and BMAL1 as positive elements and PER and CRY as negative elements. There is evidence for a direct metabolic input into the core clock mechanism. For example, REV-ERB and RORalpha, the products of two genes of the orphan nuclear hormone receptor family, repress or activate, respectively, the transcription of BMAL1 and contribute to the control of the amplitude and phase of the rhythms of clock gene expression. The same interacting circuitry of core clock and metabolic elements is present in many peripheral tissues, including muscle, liver, pancreas, and fat. Although the environmental light–dark cycle is the primary synchronizer of the central clock mechanism in the SCN, the timings of food intake and fasting have a direct effect on peripheral clocks. The central clock regulates behavioral rhythms, including the sleep–wake cycle and feeding schedule, and also entrains peripheral clocks by neural and humoral mechanisms. In the current study, we created a misalignment between the central and peripheral oscillators by imposing an 8.5-h delay of the sleep–wake and dark–light cycles on 4 of the 7 days preceding metabolic testing. Assessment of the DLMO, widely considered the most accurate marker of central circadian phase,[44] revealed that the central circadian signal had shifted by ~3 h at the end of the study.

Total sleep time and caloric intake were not affected by circadian misalignment. The timing of food intake was shifted, with a higher proportion of caloric intake occurring during the biological night and a shorter fasting period. When sleep opportunities were delayed by 8.5 h, peripheral organs were exposed to nutrients during the habitual period of overnight fast and thus received neurohormonal inputs out of phase with central circadian signals by an estimated 5 to 6 h. This misalignment between metabolic cues and central circadian signals had adverse cardiometabolic consequences that were not caused by reductions in sleep duration or quality or increases in total caloric intake.

Our study was performed under carefully controlled conditions, and the results are unequivocal. The main limitation is the sample size. A significant sex-by-group interaction emerged from the statistical analysis, but the study was not powered to examine sex differences. Findings in men were robust, with a larger-than-expected effect size of misalignment relative to alignment.

Findings from this laboratory study provide evidence in support of an intrinsic adverse effect of circadian misalignment on glucose metabolism and cardiovascular risk. The increased risk of diabetes and cardiovascular disease documented in epidemiologic studies of shift work[4–9] is thus unlikely to be solely due to sleep loss and would not be fully mitigated by strategies designed to preserve sleep duration.

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