Pathogenesis and Treatment of Muscular Dystrophy

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Description (provided by applicant): Over the last 5 years, we have been working to establish the zebrafish as a model for muscular dystrophy. In this capacity, we have published the phenotype of zebrafish lacking dystrophin and 5- sarcoglycan, completed a large genetic screen to isolate additional dystrophic mutants, and identified the mutant gene in the runzel mutant. Using our experiences in muscle research and in establishing the zebrafish as a disease model, we now propose to use the fish to investigate the pathogenesis of muscular dystrophy and evaluate cell therapy as a potential treatment option. The first aim in this project proposes to fully characterize one of our available dystrophic zebrafish models with the goal of better understanding the pathogenesis of muscular dystrophy. We have selected the emz mutant for further analysis since its mutation rough maps to a genomic interval void of any genes orthologous to those currently associated with muscular dystrophy. This mutant shows a phenotype very similar to the dystrophin mutant (sapje) suggesting that the emz phenotype of muscle degeneration is symptomatic of muscular dystrophy. We propose to identify the genetic mutation in this mutant using a traditional mapping approach and then sequencing candidate genes to identify the specific mutation. If the orthologous human gene is not currently associated with muscular dystrophy, the gene will be considered a disease candidate and sequenced in human patients for which the cause of muscular dystrophy is unknown. Since mutations in seemingly unrelated proteins can manifest as muscular dystrophy, the identification of additional genes would be helpful for establishing disease pathways. Secondly, we have established, methods to transplant cell populations in zebrafish at all developmental stages and now propose using this system to identify the cell population most capable of engrafting into and correcting the diseased muscle. Gene expression profiles of muscle engrafting cell populations will be compared with non-engrafting cells to identify genes expressed predominantly in the engrafting cells. Differentially expressed genes will be considered potential markers and used to purify analogous cell populations in mammals for future experimentation and therapy. Finally, we plan to dissect the lineage relationship of various stem cell populations by assaying the developmental potential of zebrafish muscle progenitor cells. This will be accomplished by transplanting limited populations of labeled cells early in development and then following their fate as the fish matures.