Fig. 6.

Insulin signaling in liver is impaired in female Mc4rKO relative to WT and Mc3rKO, irrespective of diet. Representative Western blots and quantified data (n = 3 per group) are shown for basal (white bar) or insulin-stimulated (black bar) PKB kinase activity (A), phosphorylation of PKB on Ser⁴⁷³ (B), phosphorylation of GSK3β on Ser⁹ (C), or phosphorylation of FoxO1 on Ser²⁵⁶ (D). Quantified data were analyzed within diet using two-way ANOVA (independent variables were genotype and insulin treatment). Phosphospecific antibody data (B–D) was normalized for total protein content. For most outcomes measured, insulin-stimulated activity was reduced in liver of Mc4rKO (MC4R), irrespective of diet, compared with Mc3rKO (MC3R) and WT mice. Significant differences between groups are indicated by lines: *, P < 0.05. For PKB activity assessed by kinase assay (A) or serine phosphorylation (B), two-way ANOVA indicated that the degree of insulin stimulation was dependent on genotype. The effects of insulin treatment were statistically significant (P < 0.05), unless indicated otherwise (n.s., not significant). AU, Arbitrary units; LFD, low-fat diet (10% kJ from fat); HFD, high-fat diet (45% kJ from fat).