Practicality of intermittent fasting in humans and its effect on oxidative stress and genes related to aging and metabolism

Abstract

Caloric restriction has consistently been shown to extend life span and ameliorate aging-related diseases. These effects may be due to diet-induced reactive oxygen species acting to up-regulate sirtuins and related protective pathways, which research suggests may be partially inhibited by dietary anti-oxidant supplementation. Because caloric restriction is not sustainable long term for most humans, we investigated an alternative dietary approach, intermittent fasting (IF), which is proposed to act on similar biological pathways. We hypothesized that a modified IF diet, where participants maintain overall energy balance by alternating between days of fasting (25% of normal caloric intake) and feasting (175% of normal), would increase expression of genes associated with aging and reduce oxidative stress and that these effects would be suppressed by anti-oxidant supplementation. To assess the tolerability of the diet and to explore effects on biological mechanisms related to aging and metabolism, we recruited a cohort of 24 healthy individuals in a double-crossover, double-blinded, randomized clinical trial. Study participants underwent two 3-week treatment periods-IF and IF with anti-oxidant (vitamins C and E) supplementation. We found strict adherence to study-provided diets and that participants found the diet tolerable, with no adverse clinical findings or weight change. We detected a marginal increase (2.7%) in SIRT3 expression due to the IF diet, but no change in expression of other genes or oxidative stress markers analyzed. We also found that IF decreased plasma insulin levels (1.01 μU/mL). Although our study suggests that the IF dieting paradigm is acceptable in healthy individuals, additional research is needed to further assess the potential benefits and risks.

Trial registration: ClinicalTrials.gov NCT02132091.

Figures

FIG. 1.

Trial design and enrollment. (A) The trial consisted of two 3-week trial sequences—intermittent fasting (IF) and IF with anti-oxidant supplementation (IFAO). Each 3-week sequence was preceded by a 1-week pre-conditioning period (PC). The two trial periods were separated by a 2-week washout (WO) period. During the IF and IFAO trial sequences, participants alternated between 25% and 175% daily caloric intake. During the PC periods, participants consumed 100% of daily caloric intake. During the IFAO sequences, participants supplemented their diet with vitamin C (1000 mg) and vitamin E (400 IU) each day orally, whereas during the IF sequences, participants supplemented their diet with a placebo. Each participant completed both the IF and IFAO trial sequences, but in randomized order. Blood draws (BD) were performed at the beginning and end of each three-week trial period. Additional details are provided in the Methods section. (B) Enrollment included 24 subjects who were randomly allocated to each treatment order. Five participants withdrew before completion of the first trial sequence and were not included in the sample analysis. Analysis of samples included two subjects who withdrew during the 2-week washout period and the remaining 17 subjects who completed the trial. PBMC, peripheral blood mononuclear cells.

FIG. 2.

Participant compliance with diet and weight change. (A) The plot displays the percent of normal caloric intake consumed by each participant on a given day of the study. Each point on the plot represents the number of calories consumed by a given person on that day. Days 1–6 and 28–33 are pre-conditioning days, during which participants were provided 100% of normal caloric intake. On days 7–27 and 34–54, participants alternated between fasting (25% of normal) and feasting (175% of normal). (B) The plot shows the weight of each participant over the course of the study. Each line is the weight of a given participant and each circle represents a weight measurement. Note that part A represents day of study provided food, whereas part B represents overall days over the course of the study.

FIG. 3.

Gene expression changes. Expression of SIRT1, SIRT3, SOD2, and TFAM from PBMCs was assayed by qPCR. The height of the bars represents the average percent change from pre-study diet during the intermittent fasting (IF) (dark grey) or IF with anti-oxidant supplementation (IF+AO) (light grey) periods. The bars represent an average across the participants and error bars are the standard errors of the mean.

FIG. 4.

Nucleotide oxidation changes. Nucleotide oxidation products 8-oxo-7,8-dihydroguanosine (8-oxo-G) and 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxo-dG) were measured by high-performance liquid chromatography coupled to electrochemical detection (HPLC-ECD), normalized by non-oxidative nucleotide levels, and log-transformed before computing mean differences and confidence intervals. The height of the bars represents the (back-transformed) fold change from pre-study diet during the intermittent fasting (IF) (dark grey) or IF with anti-oxidant supplementation (IFAO) (light grey) periods. The bars represent an (back-transformed) average across the participants and error bars are the (back-transformed) 95% confidence intervals.

FIG. 5.

Plasma insulin levels. Changes in plasma insulin levels were evaluated for intermittent fasting (IF) and IF with anti-oxidant supplementation (IFAO). Each point represents a study participant. Plotted are the insulin concentration changes in μU/mL.

FIG. 6.

Dietary satisfaction survey results. Participants were administered a dietary satisfaction survey, which was graded on a five-point scale (strongly disagree, disagree, neutral, agree, and strongly agree). Results for six questions from the survey are shown. The height of the bars represents the percentage of respondents providing a given rating.