Profibrotic Molecules Are Reduced in CRISPR-Edited Emery-Dreifuss Muscular Dystrophy Fibroblasts.

Emery-Dreifuss muscular dystrophy (EDMD) is caused by mutations in , , , and other related genes. The disease is characterized by joint contractures, muscle weakening and wasting, and heart conduction defects associated with dilated cardiomyopathy. Previous studies demonstrated the activation of fibrogenic molecules such as TGFbeta 2 and CTGF in preclinical models of EDMD2 and increased secretion of TGFbeta 2 in patient serum. A wide screening of patient cells suggested fibrosis, metabolism, and myogenic signaling as the most affected pathways in various EDMD forms. In this study, we show that alpha-smooth muscle actin-positive myofibroblasts are overrepresented in patient fibroblast cultures carrying , , or mutations, and profibrotic miRNA-21 is upregulated. Upon CRISPR/Cas correction of the mutated or sequence in EDMD1 or EDMD2 fibroblasts, respectively, we observe a reduced expression of fibrogenic molecules. However, in patient myoblasts, neither fibrogenic proteins nor miRNA-21 were upregulated; instead, miRNA-21-5p was downregulated along with muscle-specific miRNA-133b and miRNA-206, which have a crucial role in muscle cell homeostasis. These observations suggest that the conversion of laminopathic fibroblasts into a profibrotic phenotype is a determinant of EDMD-associated muscle fibrosis, while miRNA-206-dependent defects of laminopathic myoblasts, including altered regulation of VEGF levels, contribute to muscle cell deterioration. Notably, our study provides a proof-of-principle for the application of gene correction to EDMD1 and EDMD2 and presents EDMD1 isogenic cells that exhibit an almost complete rescue of a disease-specific miRNA signature. These cells can be used as experimental models for studying muscular laminopathies.