Figure 4.

Effect of T on the myogenic differentiation of hSKMCs in vitro. Human SKMCs were expanded then shifted at low confluence to DM with and without T, which was supplemented daily for 3 d. Control cells were treated with vehicle alone. A, T supplementation increased AR expression levels, as shown by Western blotting. B, hSKMCs cultured with 0, 100, or 400 nm T and immunostained for MyHC; nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Scale bar, 200 μm. C, Mean myotube area was calculated for samples showed in B and expressed in μm². Fusion index and nuclear number assays were determined in the same samples, by considering two classes of myotubes, with two to four nuclei and with more than or equal to five nuclei (D–F). T supplementation increased myotube area and resulted in formation of bigger myotubes with greater number of myonuclei. Results are means ± sem of three independent experiments. **, P < 0.01; ***, P < 0.001.