Myoblast fusion and kindlin-2 knockdown. A. C2C12 cells transfected with scrambled (scr) or kindlin-2 RNAi, incubated in differentiation media for 4 days, and then immunostained with anti-myosin heavy chain. Note that most MHC positive myoblasts treated with kindlin-2 RNAi do not form elongated, multinucleated myotubes. B. Myotube Fusion Index (MFI). The MFI was calculated using immunostained cells as in (A). The number of nuclei (labeled with DAPI) were counted in each MHC positive cell. There were significantly fewer fused myotubes in kindlin-2 knockdown cells. Results were (scrambled vs. kindlin-2): 1 nuclei 51.1+/-2.9% vs. 75.6+/-2.6% (p = 0.0001); 2–3 nuclei 25.5+/-2.6% vs. 15.4+/-1.0% (p = 0.0036); >3 nuclei 23.4+/-1.3% vs. 8.9+/-1.6% (p < 0.0001). Counts were obtained from 5 independent trials/transfections, and at least 250 cells from each condition were counted per trial.