Fig. 2

Combined knockdown of SIX1, SIX2, and SIX4 does not inhibit myogenic differentiation. FSHD1 (54–2) myoblasts were transfected with siRNAs targeting SIX1, SIX2, and SIX4, and differentiated to generate multinucleated myotubes. A Multinucleated myotubes with clusters of nuclei for the individual and combined knockdown of SIX1/2/4. B Immunofluorescent staining for nuclei (blue) and MF20 (red) for si-Control and si-SIX1/2/4 conditions. C Fusion index was calculated for the conditions shown in B. D-G Myogenic markers for (D) myogenic commitment, D early and late differentiation, E fast fiber-types, F slow fiber-types and G regenerative were assessed. For fusion index, there were three fields captured with three biological replicates. Each experimental siRNA group (n = 3) was normalized to a negative si-Control with the mean and standard deviation depicted. Asterisks demonstrate statistical significance between the control vs experimental group using an unpaired two tailed t-test (ns = not significant; *p < 0.05)