After-Effects of Time-Restricted Feeding on Whole-Body Metabolism and Gene Expression in Four Different Peripheral Tissues

Abstract

Objective: Epidemiological studies show that shift workers are at increased risk for type 2 diabetes. As modern societies increasingly require shift work, it seems crucial to determine whether there are long-lasting health effects of rotational shifts.

Methods: This study examined the after-effects of 4 weeks of time-restricted feeding (TRF) during the light period (= light-fed) in rats, an animal model for shift work. This study also included a TRF-dark (= dark-fed) control group. The aligned and misaligned feeding times of light and dark feeding are associated with poor and good health outcomes, respectively. Several physiological measures were monitored continuously; blood, liver, brown adipose tissue, and soleus and gastrocnemius muscle were collected following 11 days of ad libitum (AL) feeding after ending the TRF.

Results: In the dark-fed animals, the day/night differences in food intake, activity, and respiratory exchange ratio were still enhanced at the end of the experiment. Light-fed animals displayed the smallest day/night differences for these measures, as well as for body temperature. In both the light- and dark-fed animals, rhythms in plasma glucose, nonesterified fatty acids, and gene expression had not fully recovered after 11 days of AL feeding. Importantly, the effects on gene expression were both tissue and gene dependent.

Conclusions: Our data indicate that rotational shift workers may have an increased risk of long-lasting disturbed rhythms in several physiological measures after a period of shift work. Clearly, such disturbances may harm their health.

Figure 1

Experimental design and basic physiological measures of the rats. (A) Experimental design. During the experiment, there were three different phases: (1) the baseline phase, during which all animals had ad libitum (AL) access to food; (2) the time‐restricted feeding (TRF) phase, during which the dark‐ and light‐fed groups had limited access to food; and (3) the Revert phase, during which all animals had AL access to food again and thus could recover from the TRF phase. During the TRF phase, the light‐ and dark‐fed animals had access to chow pellets for 10 hours during either the light period (Zeitgeber Time [ZT]1‐ZT11) or the dark period (ZT13‐ZT23), respectively. (B) Body weight during the experiment. Animals were weighed weekly, as well as on the day of sacrifice. (C) Body weight gain during the experiment. (D,E) Body composition of the animals during the experiments. Relative (D) fat and (E) lean body mass steadily increased and decreased, respectively, during the course of the experiments. Relative (F) fat and (G) lean tissue mass of the liver and gastrocnemius muscle of the rats after sacrifice. ^$Significant post hoc difference between dark‐fed and light‐fed animals. ^#Significant post hoc difference between AL‐fed and light‐fed animals. *P value < 0.05 after Tukey post hoc test. **P value < 0.01 after Tukey post hoc test. (B‐E) n = 9 per experimental group. (F,G) For the liver composition, n = 19, n = 15, and n = 15 for the AL, dark‐fed, and light‐fed animals, respectively. For the gastrocnemius muscle measures, n = 15, n = 11, and n = 11 for the AL, dark‐fed, and light‐fed animals, respectively. ns, nonsignificant.

Figure 2

Metabolic parameters (food intake, activity, respiratory exchange ratio [RER], and body temperature) during the three different experimental phases. Analysis of the (A‐C) daily food intake, (D‐F) locomotor activity, (G‐I) RER, and (J‐L) subcutaneous body temperature of the animals during each of the three experimental phases. Graphs at the left, middle, and right are the 24‐hour traces of the baseline, time‐restricted feeding (TRF), and Revert phases, respectively, averaged per experimental group. (M‐P) Average 24‐hour values of the metabolic parameters for the three experimental groups. (Q‐T) Difference within the metabolic parameters between light and dark period (L/D) for the ad libitum (AL), dark‐fed, and light‐fed animals in the Revert phase. All food intake, activity, and RER values are averages of the last 2 days of each experimental phase; body temperature values are averages of the last 3 days of each experimental phase. Locomotor activity is presented as arbitrary units (AU). ****P < 0.0001, ***P < 0.001, **P < 0.01, and *P < 0.05 after Tukey post hoc test. Shaded areas represent the dark period. (A‐I,M‐O,Q‐S) For the metabolic cage measures, n = 4, n = 6, and n = 6 for the AL, dark‐fed, and light‐fed animals, respectively. (J‐L,P,T) For the body temperature measures, n = 6, n = 7, and n = 7 for the AL, dark‐fed, and light‐fed animals, respectively. L/D, light/dark; ns, nonsignificant; ZT, Zeitgeber Time.

Figure 3

After‐effects of 4 weeks of time‐restricted feeding (TRF) on daily profiles of plasma metabolite levels. Plasma levels of (A) glucose, (B) insulin, (C) nonesterified fatty acids (NEFA), and (D) corticosterone in the rats 11 days after the reversion to AL feeding conditions. Shaded areas represent the dark period. N = 4 × 6 per time point and experimental group. ZT, Zeitgeber Time.

Figure 4

After‐effects of 4 weeks of time‐restricted feeding (TRF) on expression profiles of clock genes and the clock‐controlled gene albumin D‐site‐binding protein (Dbp) in liver, brown adipose tissue (BAT), and soleus and gastrocnemius tissues in rats 11 days after the reversion to ad libitum (AL) feeding conditions. Shaded areas represent the dark period. N = 4 × 6 per time point and experimental group. ^#Difference in rhythmicity compared with AL feeding conditions (gain/loss of rhythm). ^$Difference in the acrophase compared with AL feeding conditions. Bmal1, brain and muscle aryl hydrocarbon receptor nuclear translocator–like factor 1; Clock, circadian locomotor output cycles kaput; Cry, cryptochrome; Dbp, albumin D‐site‐binding protein; Reverbα, nuclear receptor subfamily 1, group D α; ZT, Zeitgeber Time.

Figure 5

After‐effects of 4 weeks of time‐restricted feeding (TRF) on expression profiles of metabolic genes in liver, brown adipose tissue (BAT), and soleus and gastrocnemius tissues in rats 11 days after the reversion to ad libitum (AL) feeding conditions. Shaded areas represent the dark period. N = 4 × 6 per time point and experimental group. ^#Difference in rhythmicity compared with AL feeding conditions (gain/loss of rhythm). ^$Difference in the acrophase compared with AL feeding conditions. Fas, fatty acid synthase; Glut, glucose transporter; Pgc, peroxisome proliferator–activated receptor γ coactivator; Pdk, pyruvate dehydrogenase kinase; Srebf, sterol regulatory element–binding protein; Tbc1d1, trehalase 2/Bub2/cell division cycle 16 domain family member 1; Ucp, uncoupling protein; ZT, Zeitgeber Time.