RNA-FISH of nuclear foci (red) co-stained with muscleblind-like splicing regulator 1 (green) and DAPI (blue) in DM1-716 myoblasts.
Myotonic Dystrophy Type 1 (DM1) is the most common form of adult-onset muscular dystrophy. It is characterized by progressive muscle wasting and weakness, myotonia, and multisystemic involvement affecting the heart, endocrine system, and central nervous system. DM1 is caused by an expansion of CTG trinucleotide repeats in the DMPK gene on chromosome 19.
A hallmark of DM1 disease is the formation of nuclear foci that sequester RNA binding proteins that disrupt normal RNA splicing, resulting in widespread cellular dysfunction. Two iPSC-derived DM1 patient lines harboring 473 and 716 CTG repeats (DM1-473 and DM1-716, respectively) were differentiated into myoblasts. An isogenic CRISPR-corrected control for DM1-716 was generated and validated to contain zero CTG repeats using whole genome sequencing. Both patient lines express RNA foci, while the CRISPR line is corrected of the disease phenotype.
To measure the effect of CTG repeat length on muscle function, enriched myoblasts were combined with 10% fibroblasts in a fibrin-based hydrogel to generate 3D engineered muscle tissues (EMTs). A non-related wild-type line (24-WT) was included for internal comparison. Contractile force was measured over 46 days on the Mantarray platform (n = 4 tissues averaged per data point), revealing an inverse correlation between CTG repeat length (0, 473 and 716) and muscle function, similiar to in vivo models of DM1. Extended culture times provide a large therapeutic window to assess compound efficacy and disease progression.
Myotonia is a classic disease symptom in DM1 patients. It occurs when muscle relaxation is delayed after contraction, leading to muscle stiffness and difficulty moving. Prolonged relaxation rates present in both patient disease EMTs relative to normal and isogenic, CRISPR-corrected tissues, suggesting the myotonic phenotype is preserved in DM1 model myoblasts.
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