Fig. 6

p-FoxO3Ser413 and p-AMPKThr172 levels increase in muscles from cachectic mice. a Left panel: representative western blot of p-AMPK (Thr172) and total AMPK, p-FoxO3 (Ser413), p-FoxO3 (Ser253), total FoxO3 levels in TA extracts from control animals and tumor-bearing mice treated with vehicle, (−)-JQ1, and (+)-JQ1. Vinculin serves as loading control. Middle panel: average normalized values of band intensity for p-AMPK (Thr172). Right panel: average normalized values of band intensity for p-FoxO3 (Ser413) (animals per group: n = 4–6). Data represent means ± SD. Statistical analysis was performed by using one-way ANOVA followed by Tukey’s post hoc test. *p < 0.05; **p < 0.01; ***p < 0.001. “a” indicates statistical significance compared to control; “b” indicates statistical significance compared to C26 + vehicle; “c” indicates statistical significance compared to C26 + (−)-JQ1 20 mg/kg/day; “d” indicates statistical significance compared to C26 + (−)-JQ1 50 mg/kg/day. b Left panel: representative western blot analysis of p-FoxO3 (Ser413), p-FoxO3 (Ser253), total FoxO3, p-AMPK (Thr172) and total AMPK in nuclear/cytoplasmic extracts of TAs from control and tumor-bearing mice treated with vehicle or (+)-JQ1 (20 mg/ml/day). Three animals were used for each experimental group. Middle panel: average normalized values of band intensity for p-FoxO3 (Ser413) in nuclear fraction. Right panel: average normalized values of band intensity for p-AMPK (Thr172) in nuclear extracts. Statistical analysis was performed by using one-way ANOVA followed by Tukey’s post hoc test. *p < 0.05; **p < 0.01. “a” indicates statistical significance compared to control; “b” indicates statistical significance compared to C26 + vehicle. c Frozen TA sections (animals per group: n = 3) were stained with antibodies raised against p-FoxO3 (Ser413) antibodies. Arrowheads point at nuclei positive for p-FoxO3. Scale bar: 50 μm