Impact of Parental Time-Restricted Feeding on Offspring Metabolic Phenotypic Traits

Abstract

Intermittent fasting (IF) is widely recognized for its numerous health benefits, yet its impact on metabolic health across generations remains relatively unexplored. This study investigates the intergenerational effects of parental IF, specifically through 8-hour daily time-restricted feeding, on the metabolic health of offspring. By examining four different combinations of parental mating groups, we demonstrate that parental IF can influence offspring metabolic health in distinct ways. Our results reveal that parental IF conferred significant metabolic advantages compared to ad libitum (AL) feeding. IF parents exhibited lower glucose, HbA1c, cholesterol, and CRP levels, and higher ketone levels compared to AL parents. Offspring of IF-exposed animals displayed sex-specific metabolic benefits when challenged with a high-fat, high-sugar, and high-salt (HFSS) diet. Notably, female offspring from IF parents were protected against HFSS-induced glucose intolerance and exhibited lower plasma glucose levels and higher ketone levels compared to offspring of ad libitum-fed parents. Additionally, female offspring from IF parents on a HFSS diet, along with both female and male offspring on a normal diet, had elevated plasma insulin levels. Furthermore, male offspring from IF parents on a normal diet exhibited a significant reduction in body weight compared to offspring from AL parents. These findings suggest that parental IF can impart enduring metabolic benefits to offspring and may serve as an effective strategy to mitigate the risks of obesity and diabetes in future generations.

Keywords: Intergenerational inheritance; Intermittent fasting; Metabolic Syndrome.

Figure 1

Experimental Design for Parental Animals and Offspring. Parental animals were divided into two groups: one received ad libitum (AL) access to food, while the other underwent daily 16-hour intermittent fasting (IF16) for 4 months before mating. All parental mice were raised under AL conditions for 6 weeks before being randomly assigned to either continue AL feeding or undergo IF16. Offspring were separated from their mothers at 4 weeks of age and further divided into male and female groups. These groups were then subdivided into those receiving a normal diet or a high-fat, high-sugar, high-salt (HFSS) diet, and were maintained under these dietary conditions for 16 weeks.

Figure 2

Impact of Intermittent Fasting on Parental Animals. (A) and (B) show the weekly body weight changes accumulated compared to the value at week 0 in male and female animals, following daily 16-hour intermittent fasting (IF) over a 16-week period. Data are presented as means ± standard deviation. Statistical analysis was conducted using two-way repeated-measures ANOVA, followed by post-hoc Šídák's multiple comparisons test. Significance levels are denoted as *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 compared to IF animals. Each group consisted of n = 32 animals. (C) and (D) display violin plots representing blood glucose levels in male and female animals, respectively. (E) and (F) depict blood ketone levels in male and female animals, while (G) and (H) show cholesterol levels. (I and J) illustrates C-reactive protein levels in male and female parental animals. Statistical analysis for these parameters was conducted using two-way ANOVA, followed by post-hoc Šídák's multiple comparisons test. Significance levels are denoted as *P < 0.05, ***P < 0.001, ****P < 0.0001 compared to ad libitum (AL) animals. Each group consisted of n = 32 animals.

Figure 3

Glucose and Insulin Response in Offspring Following Glucose Tolerance Test. Blood glucose concentrations during glucose tolerance test in normal diet male (A and B) and normal diet female (C and D) F₁ mice at 0, 15, 30, 60 and 120 minutes post intraperitoneal glucose injection. Data are presented as means ± standard deviation. Statistical analysis was conducted using two-way repeated-measure ANOVA, followed by post-hoc uncorrected Fisher's LSD comparisons test. *P < 0.05 versus normal diet F₁ (F₀ AL male X F₀ AL Female) animals. (B and D) Glucose area under the curve (AUC) for normal diet male (B) and normal diet female (D) F₁ mice. Statistical analysis was performed using one-way ANOVA, followed by Tukey's multiple comparisons test. **P < 0.01, versus F₁ (F₀ AL male X F₀ AL Female) animals. Each group consisted of n = 17-28 animals. Similar analyses were conducted for F₁ mice fed a HFSS diet. Blood glucose concentrations during glucose tolerance test in HFSS diet male (E and F) and HFSS diet female (G and H) F₁ mice after intraperitoneal glucose injection. Blood insulin levels during the glucose tolerance test in male (I and J) and female (K and L) F₁ mice fed a normal diet, measured post intraperitoneal glucose injection. Statistical analysis was conducted using two-way repeated-measure ANOVA, followed by post-hoc uncorrected Fisher's LSD comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 denote significance compared to normal diet F₁ (F₀ AL male X F₀ AL Female) animals. Insulin AUC for male (J) and female (L) F₁ mice fed a normal diet was calculated. Statistical analysis was performed using one-way ANOVA, followed by Tukey's multiple comparisons test. *P < 0.05, **P < 0.01, ****P < 0.0001 indicate significance compared to F₁ (F₀ AL male X F₀ AL Female) animals. Each group consisted of n = 15-28 animals. Similar analyses were conducted for F1 mice fed a HFSS diet. Blood insulin concentrations during the glucose tolerance test in HFSS diet male (M and N) and female (O and P) F₁ mice were measured at 0, 30, and 120 minutes post injection. Statistical significance was assessed using two-way repeated-measure ANOVA, followed by post-hoc uncorrected Fisher's LSD comparisons test. *P < 0.05, **P < 0.01 denote significance compared to F₁ (F₀ AL male X F₀ AL Female) animals. AUC were determined, with statistical analysis conducted using one-way ANOVA, followed by Tukey's multiple comparisons test. *P < 0.05 denotes significance compared to HFSS F₁ (F₀ AL male X F₀ AL Female) animals. Each group consisted of n = 18-22 animals.

Figure 4

Blood Glucose, Ketone, Cholesterol and CRP Levels of Offspring Animals. Blood glucose levels at week 16 in male offspring under normal diet (A), HFSS diet (B), as well as female offspring under normal diet (C) and HFSS diet (D). Blood ketone levels at week 16 in male offspring under normal diet (E), HFSS diet (F), as well as female offspring under normal diet (G) and HFSS diet (H). Blood cholesterol levels at week 16 in male offspring under normal diet (I), HFSS diet (J), as well as female offspring under normal diet (K) and HFSS diet (L). Blood CRP levels at week 16 in male offspring under normal diet (M), HFSS diet (N), as well as female offspring under normal diet (O) and HFSS diet (P). Statistical analysis was conducted using one-way ANOVA, followed by post-hoc Tukey's multiple comparisons test. Significance levels are denoted as *P < 0.05, **P < 0.01. Each group comprised n = 17-28 animals.

Figure 5

Body Weight of Offspring Animals. Violin plots illustrate the body weight distributions of offspring normal diet male (A), HFSS diet male (B), normal diet female (C) and HFSS diet female (D) animals at 16 weeks after weaning. Statistical analysis was performed using one-way ANOVA, followed by post-hoc Šídák's multiple comparisons test. Significance levels are denoted as *P < 0.05 compared to F₁ (F₀ AL male X F₀ AL Female) animals. Each group consisted of n = 17-28 animals.