Figure 6

Src kinase inhibitor prevents myogenic cell fusion. Ric10 cells were cultured for 24 h in pmDM and then further cultured in pmDM supplemented with 0.1% DMSO or SU6656 for further 24 h. (A-D) The cells were fixed and subjected to immunostaining for MyHC (red). Cell nuclei were stained with DAPI (blue). Images were obtained by epifluorescence microscopy. Scale bar: 100 μm. (E) Total cell lysates (20 μg) were subjected to immunoblot analysis for the represented proteins. Flotillin, a marker of lipid rafts, was used as a loading control. (F) Differentiated cells were detected by immunostaining with anti-MyHC antibody (filled columns). Fusion indexes were calculated as percentages of nuclear numbers in multinucleated cells (open columns). Averages and standard deviations (n = 7) are shown and analyzed using Student’s t-test. *P < 0.002, **P < 0.0002. (G) Histograms represent the distribution of myogenic cells with different numbers of nuclei in unstimulated and SU6656-stimulated cultures. The cells were classified to four subpopulations: myotubes containing more than 11 nuclei (white columns), myotubes containing 2–10 nuclei (grey columns), mononucleated cells expressing MyHC (black columns), and mononucleated cells not expressing MyHC (less than 1% in the experiments).