ChREBPβ is dispensable for the control of glucose homeostasis and energy balance

Figure 1. Generation of ChREBPβ-deficient mice. (A) Structure of the Mlxipl gene. sgRNA sequences are shown in red. Exon 1b sequence is shown in green. Sequence of the carbohydrate responsive element (ChoRE) is italicized and bolded. Deleted sequence is underlined. (B) mRNA levels of ChREBP isoforms in liver, perigonadal white adipose tissue (pgWAT), inguinal WAT (ingWAT), and interscapular brown adipose tissue (BAT) of refed C57BL6/J male (n = 16–21) mice. (C) Representative Western blots and quantitation of ChREBP protein levels in liver, pgWAT, and BAT of refed C57BL6/J male mice (n = 12–24). Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test. *P < 0.05, **P < 0.01, ***P < 0.001.

Figure 2. Gene expression profiles in male ChREBPβ-deficient mice. (A) Number of regulated genes in the absence of ChREBPβ (ChREBPβ KO) or of ChREBPα and β isoforms (total ChREBP KO) compared with WT in liver, perigonadal white adipose tissue (pgWAT), inguinal WAT (ingWAT), and interscapular brown adipose tissue (BAT) of C57BL6/J refed male mice (n = 6–8 per group). (B) mRNA levels of de novo lipogenesis genes in liver, pgWAT, ingWAT, and BAT of C57BL6/J mice (n = 7–21). (C) Representative Western blots and quantitation of de novo lipogenesis proteins in liver, pgWAT, ingWAT, and BAT of C57BL6/J refed male mice (n = 12–21). Representative Western blots for pgWAT here and in Figure 1C are from the same experiment. Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test. *P < 0.05, **P < 0.01, ***P < 0.001.

Figure 3. Body composition and glucose homeostasis in male ChREBPβ-deficient mice. (A) Body weight, lean and fat mass, and organ weight in male C57BL6/J (n = 7–18) mice. (B) Glucose tolerance test and blood insulin levels 15 minutes after injection during glucose tolerance tests (n = 7–18). (C) Insulin tolerance test (n = 9–18). (D) Blood glucose and insulin levels after 24 hours of fasting (left panels) and 18 hours of refeeding (right panels) (n = 11–17). (E) Blood lipid levels after 24 hours of fasting (left panels) and 18 hours of refeeding (right panels) (n = 11–17). (F) Liver triglyceride (TG) levels after 18 hours of refeeding (n = 8–9). NEFA, nonesterified fatty acid. Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test (A, B, and D–F) or 2-way ANOVA with Šídák’s post hoc test (B and C). *P < 0.05, **P < 0.01.

Figure 4. Response of ChREBPβ-deficient mice to high-fat, high-sucrose diet. (A) Body weight, lean and fat mass, and organ weight in male FVB/N (n = 7–14) mice fed high-fat, high-sucrose diet. (B) Glucose (left panels) and insulin (right panels) tolerance tests (n = 8–14). (C) Blood glucose and insulin levels after 24 hours of fasting (left panels) and 18 hours of refeeding (right panels) (n = 6–14). (D) Blood lipid levels after 24 hours of fasting (left panels) and 18 hours of refeeding (right panels) (n = 6–10). Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test (A, C, and D) or 2-way ANOVA with Šídák’s post hoc test (B). *P < 0.05.

Figure 5. Response of ChREBPβ-deficient mice to high-fat (45%) diet. (A) Body weight, lean and fat mass, and organ weight in male C57BL6/J mice (n = 9–10). (B) Insulin tolerance test (n = 9–10). (C and D) Blood glucose, insulin (C), and lipid (D) levels in fasted (left panels) and fed (right panels) conditions (n = 8–10). (E) Liver triglyceride (TG) levels after 18 hours of refeeding (n = 9). NEFA, nonesterified fatty acid. Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test (A and C–E) or 2-way ANOVA with Šídák’s post hoc test (B).

Figure 6. Response of ChREBPβ-deficient mice to thermic challenges. (A–E) Representative H&E staining (A), mRNA levels of de novo lipogenesis and thermogenic genes (B), fatty acid profile in triglycerides (TG) (C), OXPHOS and UCP1 protein levels (D), and mitochondrial DNA (mitoDNA) content (E) in interscapular brown adipose tissue of C57BL6/J male mice adapted to thermoneutrality for 6 weeks (n = 7–9). Data are mean ± SEM. Statistical analysis was performed using Mann-Whitney test. *P < 0.05, **P < 0.01.