Intermittent fasting results in tissue-specific changes in bioenergetics and redox state

Abstract

Intermittent fasting (IF) is a dietary intervention often used as an alternative to caloric restriction (CR) and characterized by 24 hour cycles alternating ad libitum feeding and fasting. Although the consequences of CR are well studied, the effects of IF on redox status are not. Here, we address the effects of IF on redox state markers in different tissues in order to uncover how changes in feeding frequency alter redox balance in rats. IF rats displayed lower body mass due to decreased energy conversion efficiency. Livers in IF rats presented increased mitochondrial respiratory capacity and enhanced levels of protein carbonyls. Surprisingly, IF animals also presented an increase in oxidative damage in the brain that was not related to changes in mitochondrial bioenergetics. Conversely, IF promoted a substantial protection against oxidative damage in the heart. No difference in mitochondrial bioenergetics or redox homeostasis was observed in skeletal muscles of IF animals. Overall, IF affects redox balance in a tissue-specific manner, leading to redox imbalance in the liver and brain and protection against oxidative damage in the heart.

Fig 1. Intermittent fasting promotes lower body mass related to a mild reduction in caloric intake and lower energy conversion efficiency.

(A) Average body weight per cage in (●) AL and (☐) IF animals after one month of treatment. (B) Cumulative food intake over one month of treatment. (C) Energy conversion efficiency. Data represent averages ± SEM and were compared using t tests (n = 7 cages). ** p<0.01 vs AL. AL indicates ad libitum feeding, IF indicates intermittent fasting.

Fig 2. IF induces an increase in respiratory capacity in liver, without bioenergetic changes in other tissues.

Oxygen consumption in isolated mitochondria from (A) brain, (B) heart, (C) skeletal muscle and (D) liver in the presence of 5 mM pyruvate plus 3 mM malate (A, B and D) or 2 mM glutamate plus 2 mM malate (C). State 3 was induced by the addition of 1 mM ADP and state 4 was achieved using 0.5 μg/mL oligomycin. The insert in Panel D represents respiratory control ratios (RCR), or state 3/state 4. Data represent averages ± SEM and were compared using t tests (n = 4–6 animals). * p<0.05 vs AL in the same respiratory state.

Fig 3. IF promotes enhanced electron transport capacity in the liver.

Oxygen consumption in isolated mitochondria from livers in the presence of (A) 1 mM succinate plus 1 μM rotenone or (B) 200 μM TMPD plus 2 mM ascorbate. State 3 and state 4 were induced by ADP and oligomycin as described for Fig. 2. (C) COX-IV and NRF-1 levels measured as described in Materials and Methods. (D) Liver citrate synthase activity, determined as described in Materials and Methods. Data represent averages ± SEM and were compared using t tests (4–6 animals). * p<0.05, ** p<0.01 vs AL.

Fig 4. Reactive oxygen species production is not significantly altered by IF.

(A) H₂O₂ release by isolated mitochondria in the presence of 1 mM ADP (state 3) and 5 mM pyruvate plus 3 mM malate (brain, heart and liver) or 2 mM glutamate plus 2 mM malate (skeletal muscle). (B) Ratio between H₂O₂ production and O₂ consumption by isolated mitochondria under the same conditions as panel A. Data represent averages ± SEM and were compared using t tests (n = 4–6 animals).

Fig 5. IF changes oxidative damage to biomolecules in a tissue-specific manner.

(A) Carbonyl signals were quantified as described in Materials and Methods. (B) Malondaldehyde (MDA) levels were measured by HPLC as described in Materials and Methods. (C) Representative dot blots of NO₂-tyr signals. (D) Average densitometric results for the dot blots presented in C. Data represent averages ± SEM and were compared using t tests (n = 4–5 animals). * p<0.05, ** p<0.01 vs AL.