Introduction:

 Ketone bodies are small molecules consisting of β-hydroxybutyrate, acetoacetate, and acetone that serve as an alternative energy source when carbohydrates are depleted ¹⁾. In particular, β-hydroxybutyrate, which accounts for approximately 80% of ketone bodies in the blood, has been shown to have potent physiological effects. It has been reported that β-hydroxybutyrate improves mitochondrial function and may suppress muscle catabolism caused by inflammation in skeletal muscle ²⁾³⁾. However, the effect on skeletal muscle protein synthesis response is not clear. Therefore, the purpose of this study was to examine the direct effects of β-hydroxybutyrate on muscle protein synthesis and anabolic signaling in C2C12 myotubes.

 

Methods:

　C2C12 myoblasts were cultured in DMEM containing 2% horse serum for 5 days to induce differentiation into myotubes. After differentiation, the myotubes were incubated with DMEM containing different concentrations of β-hydroxybutyrate (0, 0.25, 0.5, 1, 2, 5mM) for 30 minutes. For assessment of protein synthesis, myotubes were treated with 1 µM puromycin 30 min prior to cell collection. Protein synthesis and mTORC1 related proteins were quantified using Western blotting. Data were analyzed using one-way ANOVA.

 

Results:

 A significant increase in puromycin-labeled protein expression was observed after stimulation with 1mM β-hydroxybutyrate (1.20±0.06 arbitrary units) compared to 0mM (1.00±0.05, p<0.05). However, phosphorylation of mTORC1 related proteins (p70S6K^Thr389, rpS6^Ser240/244, 4E-BP1^Thr37/46) was not changed by any concentrations of β-hydroxybutyrate.

 

Conclusion:

 These results suggest that β-hydroxybutyrate might enhance muscle protein synthesis in C2C12 myotubes. In contrast, mTORC1-related signaling did not change with short-term exposure to β-hydroxybutyrate. It appears that β-hydroxybutyrate may activate muscle protein synthesis through mTORC1-independent pathways.