Cell lines of Duchenne Muscular dystrophy patients shows mutation-dependent delay in myogenic program

Duchenne Muscular Dystrophy is a genetic X-linked neuromuscular disorder with no cure. It is due to heterogeneous mutations in dystrophin gene, that commonly lead to the absence of the protein and to a complex cascade of events. Whether the heterogeneity of the mutations could contribute to different clinical phenotype is still an open point. Phenotype-genotype correlations have never been investigated at patient-derived muscle precursors, that may represent a simplified system to advance in personalised therapy. We used two dystrophic immortalised satellite cell lines (Myoline), HDMD1 with a stop codon mutation at exon 59 and HDMD2 with deletion of 48-52 exon. We also used a third immortalised cell line from a healthy control (Hwt). Preliminary results showed a remarkable difference in the Oxygen Consumption Rate (OCR) between HDMD2 and HDMD1 cells, assessed via mitostress assays. We observed a significant reduction of all OCR parameters (basal respiration, ATP turnover/oligomycin-sensitive respiration, maximal respiration) in HDMD1 during differentiation at 48, 96 hours and 17 days, compared to HDMD2, suggesting that the respiratory defect is exacerbated by the differentiation process in a mutation-related manner. An electrophysiological investigation by means of automated patch clamp (Patchliner-Nanion) showed a marked reduction of membrane capacitance in mutant myocytes until 6 days of differentiation and a slight delay in maturation of inward and outward currents. Molecular biology experiments confirmed mutation-specific delay in myogenic cascade, and in key players of mitochondrial function and of ion channels. This characterisation shed light into genotype-phenotype correlation at cellular level, helping to identify molecular mechanism underlying patients-specific mutations and new druggable targets.