CTG-Repeat Detection in Primary Human Myoblasts of Myotonic Dystrophy Type 1

Myotonic dystrophy type 1 (DM1) is an autosomal dominant multisystemic disorder caused by unstable CTG-repeat expansions in the DMPK gene. Tissue mosaicism has been described for the length of these repeat expansions. The most obvious affected tissue is skeletal muscle, making it the first target for therapy development. To date there is no approved therapy despite some existing approaches. Thus, there is the demand to further advance therapeutic developments, which will in return require several well-characterized preclinical tools and model systems. Here we describe a modified method to identify the CTG-repeat length in primary human myoblasts isolated from DM1 patients that requires less genomic DNA and avoids radioactive labeling. Using this method, we show that primary human DM1 myoblast cultures represent a population of cells with different CTG-repeat length. Comparing DNA from the identical muscle biopsy specimen, the range of CTG-repeat length in the myoblast culture is within the same range of the muscle biopsy specimen. In conclusion, primary human DM1 myoblast cultures are a well-suited model to investigate certain aspects of the DM1 pathology. They are a useful platform to perform first-line investigations of preclinical therapies

Nuclear Envelope Transmembrane Proteins in Myotonic Dystrophy Type 1

Myotonic dystrophy type 1 (DM1) is a multisystemic disorder with predominant myotonia and muscular dystrophy which is caused by CTG-repeat expansions in the DMPK gene. These repeat expansions are transcribed and the resulting mRNA accumulates RNA-binding proteins involved in splicing, resulting in a general splicing defect. We observed nuclear envelope (NE) alterations in DM1 primary myoblasts. These included invaginations of the NE as well as an altered composition of the nuclear lamina. Specifically, we investigated NE transmembrane proteins (NETs) in DM1 primary myoblasts, staining to determine if their distribution was altered compared to controls and if this could contribute to these structural defects. We also tested the expression of these NETs in muscle and how localization changes in the DM1 primary myoblasts undergoing differentiation in vitro to myotubes. We found no changes in the localization of the tested NETs, but most tended to exhibit reduced expression with increasing DMPK-repeat length. Nonetheless, the DM1 patient expression range was within the expression range of the controls. Additionally, we found a down-regulation of the possible nesprin 1 giant isoform in DM1 primary myoblasts which could contribute to the increased NE invaginations. Thus, nesprin 1 may be an interesting target for further investigation in DM1 disease pathology