Journal: International Journal of Molecular Sciences

Article Title: Molecular Analysis of a Congenital Myasthenic Syndrome Due to a Pathogenic Variant Affecting the C-Terminus of ColQ

doi: 10.3390/ijms242216217

Figure Lengend Snippet: Phenotypic characterization of hiPSC-derived skeletal muscle cells from CTRL and patient. ( A ). Representative images of hiPSC-derived skeletal muscle cells identified by Desmin (green) and Myosin Heavy Chain (Mf20, red) after 5 days of terminal differentiation. Clusters of acetylcholine receptor (green) have been detected after stimulation with Agrin. Scale bar: 20 μm. (B ). Quantification of the fusion area calculated by Mf20 immunostaining normalized on the number of nuclei. Data represent mean± SD values after normalization on the value obtained with CTRL hiSPC-derived skeletal muscle cells ( n = 3 independent experiments). The number of AChR clusters per Myotube identified by Mf20 immunostaining was determined on 3 independent experiments in triplicate. Data were analyzed with an ordinary One-Way ANOVA. Tukey’s multiple comparisons tests ( p > 0.05, ns: not significant, ** p < 0,01). ( C ). Quantification of α, β, ε and γ AchR mRNA levels as well as MUSK and RAPSYN mRNA levels by real-time RT-PCR. Level of mRNAs are represented as relative expression (2 −ΔCt versus reference gene x100). Data represent the mean ± SD values from three independent experiments in technical triplicate and were analysed with an ordinary one-way analysis of variance (ANOVA), Tukey’s multiple comparisons test compared with CTRL (* p < 0.05, ***: p ≤ 0.001, ****: p ≤ 0.0001, ns: not significant).

Article Snippet: hiPSCs skeletal muscle differentiation experiments were performed using the commercially available Stemcell Technologies®, Vancouver, Canada, skeletal muscle differentiation media and adapted from a protocol described previously [ ].

Techniques: Derivative Assay, Immunostaining, Quantitative RT-PCR, Expressing