Role and mechanism underlying FoxO6 in skeletal muscle in vitro and in vivo

Skeletal muscle atrophy is a common feature of patients suffering with chronic infection and other systemic diseases, including acquired immunodeficiency syndrome, chronic kidney disease and cancer. Therefore, understanding the molecular basis of muscle loss is of importance. The majority of members of the forkhead box O (FoxO) family can induce skeletal muscle atrophy; however, the effect of FoxO6 on skeletal muscle is not completely understood. The present study investigated the role of FoxO6 in vitro and in vivo. Compared with the small interfering RNA (si)‑negative control (NC) group, C₂C₁₂ cell proliferation (Cell Counting Kit‑8 assay), myotube differentiation and myotube production were significantly decreased by FoxO6 knockdown, which was different from the known functions of other FoxO members. The immunofluorescence assay results demonstrated that si‑FoxO6 clearly downregulated the expression levels of myosin heavy chain (MyHC) in C₂C₁₂ myotubes compared with si‑NC. The western blotting results indicated that compared with the si‑NC group, FoxO6 knockdown induced C₂C₁₂ myotube atrophy by notably downregulating myoblast determination protein 1 (MyoD), mTOR and MyHC expression levels, and by markedly upregulating ubiquitin ligase (atrogin1) and muscle RING‑finger protein‑1 (MURF1) expression levels. Similarly, in an in vitro model of TNF‑α‑induced myotube atrophy, the western blotting results indicated that FoxO6 expression levels were decreased, whereas atrogin1, MURF1, FoxO1 and FoxO3a expression levels were increased compared with the control group. Therefore, the results indicated that, unlike FoxO1 or FoxO3a, FoxO6 maintained C₂C₁₂ myotubes and protected against atrophy. Consistent with the in vitro data, similar results were observed in vivo. Collectively, the results of the present study suggested that FoxO6 served a critical role in muscle cell metabolism in vitro and in vivo, and might serve as a promising therapeutic target for ameliorating skeletal muscle atrophy.