Expression of hemagglutinin (HA)-K63R enhances muscle differentiation in vitro . (A, B) Primary myoblast cultures were established from mice 4 to 6 weeks old and induced to differentiate by serum withdrawal. After 3 days, the cultures were stained for myosin heavy chain (MHC) and with DAPI to establish the fusion index. (C) Quantification of the fusion index from MHC-stained myotube cultures. The fusion index was established from myotubes bearing three or more nuclei using the equation: (number of nuclei in myotubes/total number of nuclei) × 100. The fusion indices were obtained from triplicate cultures of two independent animals from all genotypes. At least 200 nuclei were counted. (* P<0.008 Tg654 versus wild-type (WT), ** P<0.05 Tg3405 versus WT). (D) Differentiating cultures from both transgenic (Tg) line were monitored for the levels of MyHC, MyoD, myogenin and CyclinD1 by western blotting analysis. The figure shows results for the 3405 line; similar findings were seen for the 654 line.