Postnatal exercise protects offspring from high-fat diet-induced reductions in subcutaneous adipocyte beiging in C57Bl6/J mice

Abstract

Maternal low-protein and postnatal high-fat (HF) diets program offspring obesity and type 2 diabetes mellitus (T2DM) risk by epigenetically reducing beige adipocytes (BAs) via increased G9a protein expression (Histone3 Lysine9 dimethyl transferase), an inhibitor of the BA marker fibroblast growth factor 21 (FGF21). Conversely, offspring exercise reduces fat mass and white adipocytes, but the mechanisms are not yet understood. This work investigated whether exercise reduces offspring obesity and T2DM risk caused by a maternal HF diet via regulation of G9a and FGF21 expression that would convert white to BA. Two-month-old female C57Bl/6J mice (F0) were fed a 16% (normal fat; NF) or a 45% HF diet for 3 months prior to breeding, and subsequent gestation and lactation. Male offspring (F1) were fed the same NF and HF diets and further divided into either sedentary (S) or voluntary wheel running (Ex) groups for an additional 3 months yielding eight groups: NF (maternal treatment condition)-NF-S (postweaning treatment conditions), NF-HF-S, NF-NF-Ex, NF-HF-Ex, HF-NF-S, HF-HF-S, HF-NF-Ex, and HF-HF-Ex. Subcutaneous adipose tissue was collected for protein and mRNA analysis of FGF21, peroxisome proliferator-activated receptor-gamma coactivator (PGC-1 alpha, inducer of FGF21), G9a, E4BP4 (G9a coactivator), and protein expression of H3K9 demethylases (KDM4C). Postnatal HF diet decreased FGF21 positive BA numbers regardless of maternal diets and postnatal exercise. Under sedentary conditions, postnatal HF diet increased protein expression of FGF21 transcription inhibitors G9a and E4BP4 compared to NF diet resulting in decreased FGF21 expression. In contrast, postnatal HF diet and exercise decreased G9a and E4BP4 protein expression while decreasing FGF21 expression compared to NF diet. Under exercised condition, postnatal HF diet-induced KDM4C protein expression while no changes in KDM4C protein expression were induced by postnatal HF diet under sedentary conditions. These findings suggest that the postnatal diet exerts a greater impact on offspring adiposity and BA numbers than maternal diets. These data also suggest that offspring exercise induces KDM4C to counter the increase in G9a that was triggered by maternal and postnatal HF diets. Future studies need to determine whether KDM4C induces methylation status of G9a to alter thermogenic function of BA.

Keywords: Beige adipocyte; Exercise; Fibroblastic growth factor 21; High-fat diet; Histone lysin methylase; Subcutaneous adipose tissue.

Fig. 1.

Experimental design and maternal body composition. (A) Two-month-old C57Bl/6 female mice were assigned randomly into two experimental treatments: a NF diet or a HF diet. The mice were maintained on the experimental diet for 12 weeks followed by breeding and lactation maintaining diet. Offspring were then placed on either NF diet or a HF diet with a sedentary or exercise component. (B) Effect of HF diet on F1 male mice body composition.

Fig. 2.

Effect of HF diet on F1 male mice body composition and food intake. Male offspring were placed onto experimental diets for 12 weeks with measurements of body weight (panel A), fat mass (panel B), lean mass (panel C), average daily food intake (panel D), and the distance traveled in the EX group (panel E) were measured. Data are present in mean±SEM n = 8–9. Significant (P < .05) effects from three-way ANOVA are indicated in figure.

Fig. 3.

Effect of maternal HF diet on F1 male offspring HF diet and exercise glucose tolerance and AUC. The area under the curve (AUC) (panel A), HOMA-IR (panel B), and insulin concentration (panel C) were measured. Panel A and B data are present the mean±SEM, n = 4–8. Significant (P < .05) effects from three-way ANOVA are indicated in figure.

Fig. 4.

Effects of maternal HF diet on beige markers. The number of beige adipocytes were measured by taking three randomly selected areas from each mouse and averaging the numbers counted then normalizing the number of beige cells by the total mass of adipose tissue in sedentary animals (panel A) and exercised animals (panel B).

Fig. 5.

Effects of maternal HF and offspring HF and exercise on male offspring subcutaneous adipose tissue beige adipocyte expression. The FGF21 protein intensity was measured by IHC staining from taking three randomly selected areas for each mouse of adipose tissue placing the average in a bar graph (panel A), and the number of beige adipocytes were measured by taking three randomly selected areas from each mouse and averaging the numbers counted then normalizing the number of beige cells by the total mass of adipose tissue (panel B). The size of adipocytes was measured and calculated as described in the method section (section C). The protein expression of PGC1α (panel D) was measured. Data presented as mean±SEM n = 5–8. Significant (P < .05) effects from three-way ANOVA are indicated in figure.

Fig. 6.

Effects of maternal HF and offspring HF and exercise on male offspring subcutaneous adipose tissue metabolic protein expression. The mechanism of regulators of FGF21 (panel A). The protein expression of G9a (panel B) and E4BP4 (panel C) were measured. Data presented as mean±SEM n = 8–9. Significant (P < .05) effects from three-way ANOVA are indicated in figure.

Fig. 7.

Effects of maternal HF diet on male offspring subcutaneous adipose tissue metabolic protein expression. The protein expression of KDM4C4 (panel A) was measured. Data presented as mean±SEM n = 8–9. Significant (P < .05) effects from three-way ANOVA are indicated in figure.

Fig. 8.

Proposed mechanism of protective mechanism underlying F1 HF diet-induced decrease in FGF21 via regulation of KDM4C. F1 HF diet-induced reduction in beige adipocyte and FGF21 is minimized by reducing G9a by F1 exercise by increasing KDM4C expression which reduces methylation of H9K9 thereby reducing G9a, an inhibitor of FGF21.