Fig. 7.

Downregulation of Foxo4 diminishes the induction of atrogin mRNA in response to TNF. Small interfering RNAs (siRNAs) 100 μM specific to 2 regions of the Foxo4 mRNA and a nonspecific siRNA control (siFoxo4-0, siFoxo4-2, and NS) were transfected into myotubes 96 h postdifferentiation. Seventy-two hours posttransfection, myotubes were treated for 2 h with 6 ng/ml TNF. A: Foxo4 mRNA, as assayed by real time PCR, and protein, as assayed by Western blot analysis, were reduced by both siFoxo4-0 and siFoxo4-2 relative to the nonspecific siRNA control (*P < 0.05, n = 3). B: TNF-induced increase in atrogin mRNA, as measured by real time PCR, was reduced by siFoxo4-0 and siFoxo4-2 relative to the nonspecific siRNA control (34% average reduction, *P < 0.05, n = 3). C: linear regression showing the relationship between changes in Foxo4 protein and changes in atrogin mRNA.