Fig. 2
Promotion of muscle maturation of Healthy, LMNA-mutant, and LMNA-rescue hiPSCs using the optimized protocol. (A) Representative immunohistochemical staining for MHC in the differentiated hiPSC lines cultured for 10 days with the standard or optimized protocol. Scale bar = 100 μm. (B) The myocyte induction efficacy calculated by the percentage of nuclei in the MHC-positive area. Five images (×20 objectives) were taken from each well of the three cell lines and analyzed. (C) Images of Western blots performed on three individual samples to quantify the levels of dystrophin, MHC, and actinin in the three cell lines. (D) Graph showing the relative expression of each protein to the levels of lamin B1 expression. (E) Frequency of myotubes with specific number of nuclei as an indicator of muscle maturation. Four to five images (×10 objectives) were taken from each well of the 3 cell lines and analyzed. (F) Quantification of genes associated with muscle differentiation by RT-qPCR. Relative mRNA levels of MYH3, MYH7, MYH2, ATP2A1, CASQ1, and TMEM8C were normalized to the level of TBP, and shown as the fold increase to cells cultured using the standard protocol. Data are shown as the mean ± SD (n = 3, each group). *P < 0.05, **P < 0.01, and ***P < 0.001 versus the standard protocol.
