Arginine supplementation induces myoblast fusion via augmentation of nitric oxide production

The semi-essential amino acid, l-arginine (l-Arg), is the substrate for endogenous synthesis of nitric oxide, a molecule that is involved in myoblast proliferation and fusion. Since l-Arg supply may limit nitric oxide synthase (NOS) activity in endothelial cells, we examined l-Arg supplementation in differentiating mouse myoblasts and tested the hypothesis that l-Arg exerts direct effects on myoblast fusion via augmentation of endogenous nitric oxide production. C2C12 myoblasts in differentiation media received one of␣the␣following treatments for 120 h: 1 mM l-Arg, 0.1 mM N-nitro-l-arginine methyl ester (l-NAME), l-Arg + l-NAME, 10 mM l-Lysine, or no supplement (Control). Cultures were fixed and stained with hematoxylin and eosin for microphotometric image analysis of myotube density, nuclear density, and fusion index (% of total nuclei in myotubes). Endogenous production of nitric oxide during the treatment period peaked between 24 and 48 h. l-Arg amplified nitric oxide production between 0 and 24 h and increased myotube density, total nuclei number, and nuclear fusion index. These l-Arg effects were prevented by the NOS inhibitor, l-NAME. Further, l-Lysine, a competitive inhibitor of l-Arg uptake, repressed nitric oxide production and reduced myotube density and fusion index. In summary, l-Arg augments myotube formation and increases nitric oxide production in a process limited by cellular l-Arg uptake.