Figure 4

FoxO1 and FoxO3a regulate Cebpb , Fos and Stat3 gene expression. (A-C) Select transcripts identified via microarray as downstream targets of FoxO in locomotor muscles during C26 cancer cachexia were validated as targets in the diaphragm via qRT-PCR analyses. Data represent mean ± SE, n = 3–5 animals/group. *p < 0.05 vs AAV9-ev control group. †p < 0.05 vs. AAV9-ev C26 group. (D) FoxO1 TM or FoxO3a TM expression plasmids were injected and electroporated into TA muscles of mice and harvested 4 days later for qRT-PCR analyses. Data represent mean ± SE, n = 5 animals/group. *p < 0.05 vs empty vector (EV). (E) C2C12 myoblasts were transfected with a pRL-TK-Renilla reporter plus a WT Cebpb promoter reporter construct or a Cebpb reporter construct which is mutated at two putative Forkhead binding elements, which prevents FoxO DNA binding. Following 3 days of differentiation, myotubes were treated with IL-6 (10 ng/mL) for 3 hours and harvested for measurement of firefly/renilla luciferase activity. Data represent mean ± SE, n = at least 9 wells/group. *p < 0.05 vs control. †p < 0.05 vs. WT Cebpb promoter.